CD28

Existing Annotations Review

GO Term	Evidence	Action	Reason
GO:0042102 positive regulation of T cell proliferation	IBA GO_REF:0000033	ACCEPT	Summary: CD28 costimulation is one of the best-characterized drivers of T cell proliferation. Anti-CD28 antibodies costimulate T cell proliferation in conjunction with TCR/CD3 signaling (PMID:3159820). The deep research review confirms CD28 promotes clonal expansion through PI3K/AKT, MAPK, and NF-kB/AP-1/NFAT programmes. Reason: Positive regulation of T cell proliferation is a core function of CD28. This IBA annotation is well supported by extensive experimental literature showing CD28 costimulation is essential for T cell proliferation. The term is at an appropriate level of specificity. Supporting Evidence: PMID:3159820 mAb 9.3 was capable of inducing T cell proliferation in the presence of 12-o-tetradecanoyl phorbol-13-acetate (TPA) file:human/CD28/CD28-deep-research-falcon.md CD28 is the prototypic co‑stimulatory receptor (signal 2) that integrates with TCR signalling (signal 1) to enable full T cell activation. It stabilizes IL‑2 transcription and mRNA, promotes survival (e.g., Bcl‑xL upregulation), and drives clonal expansion
GO:0042110 T cell activation	IBA GO_REF:0000033	ACCEPT	Summary: T cell activation is the central biological process in which CD28 participates. The deep research review identifies CD28 as the prototypic costimulatory receptor providing signal 2 for T cell activation. UniProt describes CD28 as a receptor that plays a role in T-cell activation. Reason: T cell activation is a core function of CD28. The IBA annotation is consistent with the primary biological role of this receptor. Supported by extensive literature and phylogenetic conservation. Supporting Evidence: PMID:1650475 B-cell surface antigen B7 provides a costimulatory signal that induces T cells to proliferate and secrete interleukin 2 file:human/CD28/CD28-deep-research-falcon.md CD28 is the prototypic co-stimulatory receptor (signal 2) that integrates with TCR signalling (signal 1) to enable full T cell activation
GO:0009897 external side of plasma membrane	IBA GO_REF:0000033	ACCEPT	Summary: CD28 is a type I transmembrane protein with its IgV-type ligand-binding domain on the external side of the plasma membrane. UniProt annotation confirms extracellular domain spanning residues 19-152. IDA evidence from PMID:11390434 directly demonstrates cell surface expression. Reason: External side of plasma membrane is the correct localization for CD28's functional domain. The extracellular IgV domain binds B7 ligands (CD80/CD86) and this is where CD28 carries out its receptor function. Phylogenetically conserved localization. Supporting Evidence: PMID:11390434 the following Abs (all IgM) were used: CD4 (MEM-16), CD5 (MEM-128), and CD28 (248.23.2; reference 25) file:human/CD28/CD28-uniprot.txt TOPO_DOM 19..152 /note=Extracellular
GO:0031295 T cell costimulation	IBA GO_REF:0000033	ACCEPT	Summary: T cell costimulation is the defining biological process for CD28. It provides the critical signal 2 that cooperates with TCR engagement to enable full T cell activation and prevent anergy. This is the most precise BP term for CD28's role. Reason: T cell costimulation is the single most accurate biological process term for CD28 and represents its core function. CD28 is literally the prototypic T cell costimulatory receptor. Strongly supported by IBA phylogenetic evidence and extensive literature. Supporting Evidence: PMID:8617933 ligation of CD3/TCR and CD28 in the absence of CD40L triggering resulted in little or no IL-4 synthesis file:human/CD28/CD28-deep-research-falcon.md CD28 is the prototypic co-stimulatory receptor (signal 2)
GO:0050852 T cell receptor signaling pathway	IBA GO_REF:0000033	ACCEPT	Summary: CD28 costimulation amplifies and modulates TCR signaling. The 2024 imaging study demonstrates that CD28 costimulation specifically accelerates Lck recruitment and ZAP70 activation within TCR microclusters. CD28 delivers unique signals that alter the gene expression program of T cells beyond simple TCR amplification (PMID:24665965 per UniProt). Reason: CD28 is intimately coupled to the TCR signaling pathway - it amplifies proximal TCR signals by recruiting Lck to TCR microclusters, lowers the activation threshold, and feeds into the same downstream NF-kB/NFAT/AP-1 transcriptional programs. The IBA annotation is phylogenetically sound and well supported. Supporting Evidence: file:human/CD28/CD28-deep-research-falcon.md CD28 co-stimulation accelerates recruitment of Lck into TCR microclusters, which in turn accelerates ZAP70 recruitment and activation, effectively lowering the TCR activation threshold
GO:0002376 immune system process	IEA GO_REF:0000117	ACCEPT	Summary: Immune system process is a very broad parent term. CD28 is clearly involved in immune system processes as the primary T cell costimulatory receptor. However, this is much less informative than the more specific terms already annotated (T cell costimulation, T cell activation, etc.). Reason: While very broad, this IEA annotation is not incorrect. CD28 is fundamentally an immune system molecule. More specific child terms are already present. Acceptable as a broad IEA mapping.
GO:0005886 plasma membrane	IEA GO_REF:0000044	ACCEPT	Summary: CD28 is a type I single-pass transmembrane protein that localizes to the plasma membrane. UniProt subcellular location annotation confirms cell membrane localization. Multiple IDA annotations also support this. Reason: Plasma membrane is the correct localization for CD28. This IEA annotation is consistent with multiple higher-evidence annotations (IDA) for the same term. Well supported by UniProt subcellular location.
GO:0006955 immune response	IEA GO_REF:0000002	ACCEPT	Summary: Immune response is a broad BP term from InterPro mapping based on the CD28 domain (IPR008093). CD28 is essential for adaptive immune responses, particularly T cell-mediated immunity. Reason: While broad, the IEA annotation from InterPro mapping is correct. CD28 is a central player in adaptive immune responses. More specific terms (T cell costimulation, T cell activation) provide the needed detail.
GO:0009986 cell surface	IEA GO_REF:0000044	ACCEPT	Summary: Cell surface localization is correct for CD28, which is expressed on the T cell surface. Also supported by IDA evidence from PMID:15067037. Reason: Correct IEA annotation. CD28 is a cell surface receptor. Consistent with IDA evidence from PMID:15067037 and the external side of plasma membrane IBA annotation.
GO:0016020 membrane	IEA GO_REF:0000002	ACCEPT	Summary: Membrane is a very broad CC term from InterPro mapping. CD28 is a transmembrane protein, so this is correct but uninformative given more specific terms (plasma membrane, external side of plasma membrane). Reason: Correct but very broad. More specific CC annotations are available. Acceptable as a broad IEA mapping from InterPro.
GO:0042129 regulation of T cell proliferation	IEA GO_REF:0000002	ACCEPT	Summary: Regulation of T cell proliferation from InterPro mapping (IPR040216 CTLA4/CD28 domain). CD28 positively regulates T cell proliferation, so this parent term is correct. Reason: Correct IEA annotation. The more specific child term GO:0042102 (positive regulation of T cell proliferation) is also annotated with experimental evidence. This broader IEA term is acceptable.
GO:0050870 positive regulation of T cell activation	IEA GO_REF:0000117	ACCEPT	Summary: Positive regulation of T cell activation is appropriate for CD28. CD28 costimulation positively regulates T cell activation by lowering the TCR activation threshold, promoting cytokine production, and driving proliferation. Reason: Correct IEA annotation. CD28 is the canonical positive regulator of T cell activation. Consistent with the IBA annotation for T cell activation and the experimental evidence for costimulation.
GO:0005515 protein binding	IPI PMID:10820259 GRID: a novel Grb-2-related adapter protein that interacts w...	MODIFY	Summary: IPI annotation for protein binding based on interaction with PIK3R1 (P27986, the p85 regulatory subunit of PI3K). The CD28 YMNM motif recruits PI3K via the p85 SH2 domain. This is a functionally important interaction for CD28 signaling. Reason: Protein binding is uninformative. The interaction with PIK3R1 (p85) is a well-characterized signaling interaction via the YMNM motif. This should be captured with a more specific MF term reflecting the signaling adaptor recruitment function. Proposed replacements: protein kinase binding Supporting Evidence: PMID:7568038 p56Lck and p59Fyn regulate CD28 binding to phosphatidylinositol 3-kinase, growth factor receptor-bound protein GRB-2, and T cell-specific protein-tyrosine kinase ITK
GO:0005515 protein binding	IPI PMID:18641334 ICOS ligation recruits the p50alpha PI3K regulatory subunit ...	MODIFY	Summary: IPI annotation for protein binding based on interaction with PIK3R1 (P27986). Another publication documenting CD28-PI3K p85 interaction. Publication not available for review. Reason: Protein binding is uninformative. Same as above - should be protein kinase binding or a more specific term reflecting CD28-PI3K signaling interaction. Proposed replacements: protein kinase binding
GO:0005515 protein binding	IPI PMID:21982860 A secreted protein microarray platform for extracellular pro...	MODIFY	Summary: High-throughput secreted protein microarray screen identifying CD28 interaction with CD80 (P33681/B7-1). This is a core ligand-receptor interaction. The paper describes a screening platform for extracellular protein interaction discovery. Reason: Protein binding is uninformative for the CD28-CD80 ligand-receptor interaction. CD28 binding to CD80 is the fundamental receptor-ligand interaction that defines CD28 function. The coreceptor activity term (GO:0015026) already captures this MF more precisely. Proposed replacements: coreceptor activity Supporting Evidence: PMID:21982860 functional protein microarrays are particularly well-suited for high-throughput screening of extracellular protein interactions
GO:0005515 protein binding	IPI PMID:32822567 A Human IgSF Cell-Surface Interactome Reveals a Complex Netw...	MODIFY	Summary: IgSF cell-surface interactome study confirming CD28 interaction with CD80 (P33681). This large-scale study systematically mapped direct protein interactions across a recombinant IgSF library. Reason: Protein binding is too vague for CD28-CD80 receptor-ligand interaction. This is a core functional interaction already captured by coreceptor activity. Proposed replacements: coreceptor activity Supporting Evidence: PMID:32822567 Cell-surface protein-protein interactions (PPIs) mediate cell-cell communication, recognition, and responses
GO:0005515 protein binding	IPI PMID:32822567 A Human IgSF Cell-Surface Interactome Reveals a Complex Netw...	MODIFY	Summary: Same IgSF interactome study, this entry for CD28 interaction with CD86 (P42081/B7-2). CD86 is the other principal ligand for CD28 alongside CD80. Reason: Protein binding is uninformative. CD28-CD86 is a core receptor-ligand interaction. Should be captured under coreceptor activity. Proposed replacements: coreceptor activity
GO:0005515 protein binding	IPI PMID:35922511 A physical wiring diagram for the human immune system.	MODIFY	Summary: Physical wiring diagram for the human immune system confirming CD28-CD80 interaction. Large-scale systematic mapping of direct protein interactions across human leukocyte surface proteins. Reason: Protein binding is uninformative. Another confirmation of the CD28-CD80 core ligand interaction. Should be coreceptor activity. Proposed replacements: coreceptor activity Supporting Evidence: PMID:35922511 using a high-throughput surface receptor screening method, we systematically mapped the direct protein interactions across a recombinant library that encompasses most of the surface proteins that are detectable on human leukocytes
GO:0005515 protein binding	IPI PMID:35922511 A physical wiring diagram for the human immune system.	MODIFY	Summary: Same wiring diagram study, this entry for CD28-CD86 (P42081) interaction. Reason: Protein binding is uninformative for CD28-CD86 core ligand interaction. Should be coreceptor activity. Proposed replacements: coreceptor activity
GO:0005515 protein binding	IPI PMID:7568038 p56Lck and p59Fyn regulate CD28 binding to phosphatidylinosi...	MODIFY	Summary: Raab et al. 1995 demonstrated that Lck and Fyn regulate CD28 binding to PI3K (PIK3R1/p85), GRB2, and ITK. Mutation of Tyr-191 greatly reduced phosphorylation by Lck. The with/from column indicates PIK3R1 (P27986). This paper is a foundational study of CD28 signaling. Reason: Protein binding is uninformative for this well-characterized signaling interaction. CD28-PI3K interaction is mediated by the YMNM motif and is critical for costimulatory signaling. Should be protein kinase binding. Proposed replacements: protein kinase binding Supporting Evidence: PMID:7568038 p56Lck and p59Fyn regulate CD28 binding to phosphatidylinositol 3-kinase, growth factor receptor-bound protein GRB-2, and T cell-specific protein-tyrosine kinase ITK
GO:0005515 protein binding	IPI PMID:7584133 Selective CD28pYMNM mutations implicate phosphatidylinositol...	MODIFY	Summary: Cai et al. 1995 showed that CD28 pYMNM mutations disrupt PI3K and GRB2 binding, directly implicating PI3K in CD28-mediated costimulation leading to IL-2 secretion. With/from indicates PIK3R1 (P27986). Reason: Protein binding is uninformative. This foundational paper demonstrates that the CD28-PI3K interaction via the YMNM motif is essential for costimulation. Should be protein kinase binding. Proposed replacements: protein kinase binding Supporting Evidence: PMID:7584133 Both Y191CD28F and M194CD28C mutants failed to generate IL-2. These data directly implicate PI 3-kinase in CD28-mediated costimulation leading to IL-2 secretion
GO:0005515 protein binding	IPI PMID:7807015 CTLA-4 binding to the lipid kinase phosphatidylinositol 3-ki...	MODIFY	Summary: Schneider et al. 1995 studied CTLA-4 binding to PI3K, but with/from indicates PIK3R1 (P27986). The paper demonstrates that CTLA-4 also associates with PI3K, comparing binding affinities with CD28. CD28 data was used as a reference point. Reason: Protein binding is uninformative. This documents the CD28-PI3K p85 interaction in comparison to CTLA-4. Should be protein kinase binding. Proposed replacements: protein kinase binding Supporting Evidence: PMID:7807015 the NH2- and COOH-terminal SH2 domains of p85 bind the CTLA-4 cytoplasmic pYVKM motif with an affinity (ID50: 0.6 and 0.04 microM), that is similar to CD28
GO:0005515 protein binding	IPI PMID:8146197 T-cell antigen CD28 interacts with the lipid kinase phosphat...	MODIFY	Summary: Publication not available for review. With/from indicates PIK3R1 (P27986). Likely another study documenting the CD28-PI3K p85 interaction. Reason: Protein binding is uninformative. Based on the with/from column indicating PIK3R1 interaction, should be protein kinase binding. Publication not available to confirm specific details. Proposed replacements: protein kinase binding
GO:0005515 protein binding	IPI PMID:8183372 Binding of phosphatidylinositol-3-OH kinase to CD28 is requi...	MODIFY	Summary: Publication not available for review. With/from indicates PIK3R1 (P27986). Another study documenting CD28-PI3K interaction. Reason: Protein binding is uninformative. Based on with/from indicating PIK3R1, should be protein kinase binding. Publication not available. Proposed replacements: protein kinase binding
GO:0005515 protein binding	IPI PMID:9417079 Growth factor receptor-bound protein 2 SH2/SH3 domain bindin...	MODIFY	Summary: Kim et al. 1998 characterized GRB2 SH2/SH3 domain binding to CD28 and its role in co-signaling. Both SH2 and SH3 domains contribute to CD28 binding. Mutations that alter GRB2 binding blocked CD28-dependent IL-2 production and Vav phosphorylation. With/from indicates PIK3R1 (P27986). Reason: Protein binding is uninformative. This paper documents functional significance of GRB2 binding to CD28 via both SH2 and SH3 domains. Should be more specific. Proposed replacements: protein kinase binding Supporting Evidence: PMID:9417079 Mutations that alter Grb2 binding were found to block the CD28-dependent interleukin-2 production. Further, tyrosine phosphorylation of Vav and the costimulation-dependent activation of Jun N-terminal kinase was blocked
GO:0001772 immunological synapse	IEA GO_REF:0000107	ACCEPT	Summary: Ensembl Compara transfer from mouse CD28 (P31041). CD28 localizes to the immunological synapse upon T cell-APC contact, where it co-assembles with TCR microclusters. The 2024 TIRF microscopy study directly demonstrates CD28 costimulation shapes signaling within TCR microclusters at the immune synapse. Reason: Immunological synapse localization is well established for CD28. This is where CD28 carries out its costimulatory function by recruiting Lck to TCR microclusters. The IEA transfer from mouse is consistent with extensive experimental evidence. Supporting Evidence: file:human/CD28/CD28-deep-research-falcon.md CD28 co-localizes with TCR microclusters within the immune synapse
GO:0009897 external side of plasma membrane	IEA GO_REF:0000107	ACCEPT	Summary: Ensembl Compara transfer from mouse CD28. Consistent with IBA annotation for same term and IDA evidence from PMID:11390434. Reason: Correct IEA annotation. Consistent with IBA and IDA evidence for the same term.
GO:0019901 protein kinase binding	IEA GO_REF:0000107	ACCEPT	Summary: Ensembl Compara transfer from mouse. CD28 binds multiple protein kinases including PI3K (via p85 regulatory subunit, YMNM motif), Lck (via PYAP motif), ITK (via PRRP motif), and PKC-theta. This is a well-documented core molecular function. Reason: Protein kinase binding is well supported for CD28. The cytoplasmic tail recruits PI3K, Lck, ITK, and PKC-theta through distinct motifs. This IEA is consistent with extensive experimental evidence and is a far more informative MF term than generic protein binding. Supporting Evidence: PMID:7568038 p56Lck and p59Fyn regulate CD28 binding to phosphatidylinositol 3-kinase, growth factor receptor-bound protein GRB-2, and T cell-specific protein-tyrosine kinase ITK file:human/CD28/CD28-deep-research-falcon.md YMNM (binds PI3K and GRB2), PRRP (recruits ITK leading to PLCgamma1 and SLP-76/GADS engagement), and PYAP (dominant for LCK binding and assembly of PKCtheta at the synapse)
GO:0005886 plasma membrane	IDA GO_REF:0000052	ACCEPT	Summary: IDA annotation based on immunofluorescence data (HPA). CD28 is a transmembrane protein at the plasma membrane. Reason: Plasma membrane localization is unambiguously correct for CD28. Supported by immunofluorescence and consistent with the known type I transmembrane topology.
GO:0004888 transmembrane signaling receptor activity	IDA PMID:11279501 Structural basis for co-stimulation by the human CTLA-4/B7-2...	ACCEPT	Summary: Schwartz et al. 2001 solved the crystal structure of the CTLA-4/B7-2 complex, providing structural basis for costimulation. The paper discusses both CD28 and CTLA-4 as transmembrane signaling receptors that engage B7 ligands. CD28 ligation with B7 ligands initiates intracellular signaling cascades. Reason: Transmembrane signaling receptor activity is an appropriate MF for CD28. CD28 spans the membrane and upon ligand (CD80/CD86) binding, transmits signals intracellularly through its cytoplasmic YMNM/PRRP/PYAP motifs. This term is accurate but somewhat general; coreceptor activity (GO:0015026) is more precise. Supporting Evidence: PMID:11279501 Regulation of T-cell activity is dependent on antigen-independent co-stimulatory signals provided by the disulphide-linked homodimeric T-cell surface receptors, CD28 and CTLA-4
GO:0004888 transmembrane signaling receptor activity	IDA PMID:12196291 The interaction properties of costimulatory molecules revisi...	ACCEPT	Summary: Collins et al. 2002 revisited the interaction properties of costimulatory molecules including CD28 with CD80 and CD86. UniProt cites this paper for CD28 function and interaction with CD80/CD86. Publication not available but UniProt annotates it for CD28 function. Reason: Transmembrane signaling receptor activity is correct for CD28. This paper provided key interaction data for CD28-B7 engagement.
GO:0005886 plasma membrane	IDA PMID:35397202 Structural characterization of a dimerization interface in t...	ACCEPT	Summary: Wu et al. 2022 characterized the dimerization interface in the CD28 transmembrane domain by NMR. UniProt cites this for subcellular location (cell membrane). The paper confirms CD28 as a membrane protein and characterizes TM helix dimerization. Reason: IDA evidence directly confirming plasma membrane localization via structural characterization of the transmembrane domain. Supporting Evidence: file:human/CD28/CD28-uniprot.txt SUBCELLULAR LOCATION: Cell membrane {ECO:0000269\|PubMed:35397202}; Single-pass type I membrane protein
GO:0050862 positive regulation of T cell receptor signaling pathway	IDA PMID:11279501 Structural basis for co-stimulation by the human CTLA-4/B7-2...	ACCEPT	Summary: Schwartz et al. 2001 structural study of CTLA-4/B7-2 complex. The paper demonstrates that CD28 engagement with B7 ligands provides a stimulatory signal for T cell activation, consistent with positive regulation of TCR signaling. Reason: Positive regulation of TCR signaling pathway is a core function of CD28. CD28 costimulation amplifies TCR signals by recruiting Lck to TCR microclusters and lowering activation thresholds. Well supported by this and many other studies. Supporting Evidence: PMID:11279501 Engagement of CD28 with B7-1 and B7-2 ligands on antigen-presenting cells (APCs) provides a stimulatory signal for T-cell activation
GO:0050862 positive regulation of T cell receptor signaling pathway	IDA PMID:12196291 The interaction properties of costimulatory molecules revisi...	ACCEPT	Summary: Collins et al. 2002 characterized interaction properties of costimulatory molecules. UniProt cites this for CD28's function in increasing proliferation and cytokine expression. Reason: Same core function. Multiple IDA annotations for positive regulation of TCR signaling from different publications strengthens the evidence. Consistent with CD28's role as the prototypic TCR costimulatory receptor.
GO:0050862 positive regulation of T cell receptor signaling pathway	IDA PMID:1650475 B-cell surface antigen B7 provides a costimulatory signal th...	ACCEPT	Summary: Gimmi et al. 1991, a foundational paper showing B7 provides a costimulatory signal that induces T cells to proliferate and secrete IL-2. UniProt cites this for CD28 function. Publication not available but this is one of the original demonstrations of CD28/B7 costimulation. Reason: This is one of the earliest and most important studies demonstrating that B7-CD28 costimulation positively regulates T cell responses. Core annotation well supported by this seminal publication.
GO:0004888 transmembrane signaling receptor activity	TAS PMID:9784967 CD28/B7 costimulation: a review.	ACCEPT	Summary: TAS annotation based on a review. CD28 is clearly a transmembrane signaling receptor. Publication not available for review. Reason: Transmembrane signaling receptor activity is correct. CD28 is a type I transmembrane receptor that signals upon ligand engagement. TAS annotation is appropriate for a review-based citation.
GO:0042110 T cell activation	IDA PMID:23918985 Soluble CD80 restores T cell activation and overcomes tumor ...	ACCEPT	Summary: IDA annotation for T cell activation. Publication not available for review. UniProt cites CD28's role in T cell activation from multiple publications. Reason: T cell activation is a core function of CD28. While the specific publication is not available, this annotation is consistent with the overwhelming body of evidence for CD28 in T cell activation.
GO:0042110 T cell activation	IGI PMID:18337562 CCL5-mediated T-cell chemotaxis involves the initiation of m...	ACCEPT	Summary: IGI annotation for T cell activation with genetic interactor IL-4 (P05112). The referenced publication PMID:18337562 (Murooka et al. 2008) focuses on CCL5-mediated T cell chemotaxis and mTOR signaling, not directly on CD28. However, T cell activation is the defining biological process for CD28, supported by overwhelming evidence from multiple other publications and the IBA and IDA annotations for the same term. CD28 is the prototypic T cell costimulatory receptor; its role in T cell activation is beyond question (PMID:1650475, PMID:23918985, PMID:11279501). Reason: Although the specific publication (PMID:18337562) is about CCL5-mediated chemotaxis rather than CD28 per se, T cell activation is the single most well-established function of CD28. The IBA and IDA annotations for this same term are already accepted. CD28 is literally defined as the prototypic T cell costimulatory receptor, and T cell activation is its core function. The IGI evidence code with IL-4 as genetic interactor is consistent with the known role of CD28 costimulation in cytokine-dependent T cell activation programs. Resolving to ACCEPT for consistency with the other GO:0042110 annotations. Supporting Evidence: PMID:1650475 B7-transfected CHO cells can induce suboptimally activated CD28+ T cells to proliferate and secrete high levels of interleukin 2 PMID:11279501 Engagement of CD28 with B7-1 and B7-2 ligands on antigen-presenting cells (APCs) provides a stimulatory signal for T-cell activation file:human/CD28/CD28-deep-research-falcon.md CD28 is the prototypic co-stimulatory receptor (signal 2) that integrates with TCR signalling (signal 1) to enable full T cell activation
GO:0010628 positive regulation of gene expression	IMP PMID:23817958 Proinflammatory stimuli induce galectin-9 in human mesenchym...	KEEP AS NON CORE	Summary: IMP annotation for positive regulation of gene expression. Publication not available. CD28 signaling is known to activate NF-kB, NFAT, and AP-1 transcription factors, which positively regulate expression of many target genes including IL-2. Reason: While CD28 signaling does regulate gene expression programs through NF-kB/NFAT/AP-1 activation, this is a very broad downstream effect rather than a core molecular function of CD28 itself. The annotation is not wrong, but represents a downstream consequence of CD28 costimulatory signaling rather than a defining function. Publication not available to assess specific experimental context.
GO:0010629 negative regulation of gene expression	IMP PMID:23817958 Proinflammatory stimuli induce galectin-9 in human mesenchym...	KEEP AS NON CORE	Summary: IMP annotation for negative regulation of gene expression from the same publication as the positive regulation annotation above. CD28 signaling can indeed both upregulate and downregulate different gene sets. Reason: Same reasoning as for positive regulation of gene expression. This is a downstream consequence of CD28 signaling, not a core function. The annotation may be valid for specific gene sets that are downregulated by CD28 costimulation, but it is a very broad and non-specific annotation. Publication not available.
GO:0043066 negative regulation of apoptotic process	TAS PMID:8717514 CD28/B7 system of T cell costimulation.	KEEP AS NON CORE	Summary: TAS annotation for negative regulation of apoptosis. CD28 signaling promotes T cell survival through upregulation of anti-apoptotic proteins including Bcl-xL. This is a well-known downstream effect of CD28 costimulation. Publication not available. Reason: CD28 signaling does promote cell survival and inhibit apoptosis (e.g., via Bcl-xL upregulation), but this is a downstream consequence of its costimulatory signaling rather than its primary molecular function. It is a pleiotropic effect of CD28 activation, not its defining function. Supporting Evidence: file:human/CD28/CD28-deep-research-falcon.md promotes survival (e.g., Bcl-xL upregulation)
GO:0098636 protein complex involved in cell adhesion	IDA PMID:7544393 Identification of residues in the V domain of CD80 (B7-1) im...	KEEP AS NON CORE	Summary: Fargeas et al. 1995 identified residues in the CD80 V domain implicated in functional interactions with CD28 and CTLA-4. The paper studied CD80-CD28 adhesion and IL-2 production using CD80 mutants. The annotation refers to CD28 as part of a protein complex involved in cell adhesion (T cell-APC interaction). Reason: While CD28-CD80 interaction does contribute to T cell-APC adhesion at the immune synapse, this is not the primary function of CD28. CD28 is fundamentally a signaling coreceptor, not primarily an adhesion molecule. The cell adhesion aspect is secondary to its costimulatory signaling role. Supporting Evidence: PMID:7544393 human CD80 mutants were generated and tested for their ability to maintain the interaction with CD28 leading to adhesion and enhanced IL-2 production
GO:0031295 T cell costimulation	TAS PMID:8617933 Cross-linking of the CD40 ligand on human CD4+ T lymphocytes...	ACCEPT	Summary: Blotta et al. 1996 showed that CD40L costimulation enhances IL-4 synthesis in conjunction with CD3/TCR and CD28 ligation. The paper directly demonstrates CD28's role in T cell costimulation. Reason: T cell costimulation is the core function of CD28. This TAS annotation from a study directly examining CD28 costimulation in conjunction with CD3 and CD40L is well supported. Supporting Evidence: PMID:8617933 IL-4 synthesis was greatly enhanced by triggering of CD40L on the T cell surface in conjunction with ligation of CD3/TCR and CD28
GO:0032733 positive regulation of interleukin-10 production	IDA PMID:8617933 Cross-linking of the CD40 ligand on human CD4+ T lymphocytes...	KEEP AS NON CORE	Summary: Blotta et al. 1996 showed that CD3/CD28/CD40L costimulation enhances cytokine production including IL-10 in T cells. Reason: IL-10 production is a downstream consequence of CD28 costimulation combined with CD40L signaling. While experimentally demonstrated (IDA), this is a specific cytokine output rather than a core function of CD28. The core function is T cell costimulation broadly. Supporting Evidence: PMID:8617933 IL-4 synthesis was greatly enhanced by triggering of CD40L on the T cell surface in conjunction with ligation of CD3/TCR and CD28
GO:0032753 positive regulation of interleukin-4 production	IDA PMID:8617933 Cross-linking of the CD40 ligand on human CD4+ T lymphocytes...	KEEP AS NON CORE	Summary: Blotta et al. 1996 directly demonstrated that CD28 costimulation (with CD3 and CD40L) greatly enhances IL-4 synthesis. This was the main finding of the paper. Reason: IL-4 production enhancement is a specific downstream effect of CD28 costimulation, particularly in combination with CD40L signaling. While experimentally validated, this represents a specific cytokine output rather than the core costimulatory function. Supporting Evidence: PMID:8617933 IL-4 synthesis was greatly enhanced by triggering of CD40L on the T cell surface in conjunction with ligation of CD3/TCR and CD28, whereas ligation of CD3/TCR and CD28 in the absence of CD40L triggering resulted in little or no IL-4 synthesis
GO:0042102 positive regulation of T cell proliferation	IDA PMID:8617933 Cross-linking of the CD40 ligand on human CD4+ T lymphocytes...	ACCEPT	Summary: Blotta et al. 1996 demonstrated T cell proliferation in response to CD3/CD28 costimulation. The paper used anti-CD28 antibodies as costimulatory signals. Reason: Positive regulation of T cell proliferation is a core function of CD28. IDA evidence from Blotta et al. directly demonstrates this. Consistent with multiple other annotations. Supporting Evidence: PMID:8617933 using immobilized mAbs against several T cell surface molecules, including CD3, CD28, and the CD40 ligand (CD40L)
GO:0005515 protein binding	IPI PMID:15067037 The modulation of CD40 ligand signaling by transmembrane CD2...	UNDECIDED	Summary: Mikolajczak et al. 2004 showed that isoform 3 (CD28i) interacts with CD40LG (P29965/CD154). This is an interaction specific to the CD28i splice variant that lacks most of the extracellular domain. Reason: This interaction is specific to isoform 3 (CD28i), which has a truncated extracellular domain and interacts with CD40LG rather than B7 ligands. While the interaction is experimentally demonstrated, it is isoform-specific. Protein binding is uninformative but the isoform-specific nature makes it harder to propose a simple replacement. Supporting Evidence: file:human/CD28/CD28-uniprot.txt Isoform 3: Enhances CD40L-mediated activation of NF-kappa-B and kinases MAPK8 and PAK2 in T-cells
GO:0009986 cell surface	IDA PMID:15067037 The modulation of CD40 ligand signaling by transmembrane CD2...	ACCEPT	Summary: Mikolajczak et al. 2004 demonstrated cell surface localization of isoform 3 (CD28i). UniProt confirms subcellular location for isoform 3 as cell surface. Reason: Cell surface localization is correct for CD28, including isoform 3. IDA evidence directly demonstrates this. Supporting Evidence: file:human/CD28/CD28-uniprot.txt SUBCELLULAR LOCATION: [Isoform 3]: Cell surface
GO:0005886 plasma membrane	TAS Reactome:R-HSA-2316434	ACCEPT	Summary: Reactome annotation for PI3K phosphorylation of PIP2 to PIP3 in context of CD28 signaling at the plasma membrane. Reason: Plasma membrane localization in the context of PI3K signaling downstream of CD28 is correct. CD28 recruits PI3K to the plasma membrane.
GO:0005886 plasma membrane	TAS Reactome:R-HSA-2400009	ACCEPT	Summary: Reactome annotation for PI3K inhibitors blocking PI3K catalytic activity. CD28 at the plasma membrane. Reason: Correct. CD28 at the plasma membrane in PI3K signaling context.
GO:0005886 plasma membrane	TAS Reactome:R-HSA-388814	ACCEPT	Summary: Reactome annotation for Grb2 binding to CD28 at the plasma membrane. Reason: Correct. CD28 recruits Grb2 at the plasma membrane via the YMNM motif.
GO:0005886 plasma membrane	TAS Reactome:R-HSA-388831	ACCEPT	Summary: Reactome annotation for phosphorylation of CD28 at the plasma membrane. Reason: Correct. CD28 is phosphorylated by Lck at the plasma membrane.
GO:0005886 plasma membrane	TAS Reactome:R-HSA-388832	ACCEPT	Summary: Reactome annotation for PI3K binding to CD28 at the plasma membrane. Reason: Correct. PI3K binds CD28 at the plasma membrane via the pYMNM motif.
GO:0005886 plasma membrane	TAS Reactome:R-HSA-389158	ACCEPT	Summary: Reactome annotation for CD28-bound PI3K phosphorylating PIP2 to PIP3. Reason: Correct. PI3K recruited to CD28 at the plasma membrane generates PIP3.
GO:0005886 plasma membrane	TAS Reactome:R-HSA-389348	ACCEPT	Summary: Reactome annotation for activation of Rac1 by pVav1 in the CD28 pathway at the plasma membrane. Reason: Correct. Vav1 activation downstream of CD28 at the plasma membrane leads to Rac1 activation.
GO:0005886 plasma membrane	TAS Reactome:R-HSA-389350	ACCEPT	Summary: Reactome annotation for activation of Cdc42 by pVav1 in the CD28 pathway at the plasma membrane. Reason: Correct. Vav1 activation downstream of CD28 at the plasma membrane leads to Cdc42 activation.
GO:0005886 plasma membrane	TAS Reactome:R-HSA-389352	ACCEPT	Summary: Reactome annotation for translocation of Vav1 to CD28 at the plasma membrane. Reason: Correct. Vav1 is recruited to CD28 at the plasma membrane.
GO:0005886 plasma membrane	TAS Reactome:R-HSA-389354	ACCEPT	Summary: Reactome annotation for activation of Vav1 in the CD28 pathway at the plasma membrane. Reason: Correct. Vav1 is activated in the CD28 signaling pathway at the plasma membrane.
GO:0005886 plasma membrane	TAS Reactome:R-HSA-389381	ACCEPT	Summary: Reactome annotation for Gads binding to CD28 at the plasma membrane. Reason: Correct. Gads is recruited to CD28 at the plasma membrane.
GO:0005829 cytosol	TAS Reactome:R-HSA-167637	KEEP AS NON CORE	Summary: Reactome annotation for internalization of Nef:CD28:Clathrin-Coated Pit Adapter Protein complex. This is in the context of HIV Nef-mediated downregulation of CD28, where CD28 is internalized to the cytosol. Reason: Cytosol localization of CD28 is specific to the HIV Nef-mediated downregulation pathway where CD28 is internalized from the cell surface. This is not a normal physiological localization of CD28 but rather a pathological consequence of viral hijacking. Not a core localization.
GO:0005886 plasma membrane	IDA PMID:11390434 The transmembrane adaptor protein TRIM regulates T cell rece...	ACCEPT	Summary: Kirchgessner et al. 2001 studied the transmembrane adaptor protein TRIM and its association with TCR. CD28 plasma membrane expression was confirmed as part of the characterization of T cell surface molecules. Reason: IDA evidence for plasma membrane localization. Correct and consistent with all other evidence.
GO:0009897 external side of plasma membrane	IDA PMID:11390434 The transmembrane adaptor protein TRIM regulates T cell rece...	ACCEPT	Summary: Same study as above. IDA evidence for external side of plasma membrane localization, consistent with IBA annotation. Reason: IDA evidence confirming CD28's extracellular domain is on the external side of the plasma membrane. Consistent with IBA and IEA annotations for the same term.
GO:0045066 regulatory T cell differentiation	IDA PMID:18641304 Acquisition of suppressive function by activated human CD4+ ...	KEEP AS NON CORE	Summary: Zheng et al. 2008 studied acquisition of suppressive function by activated human CD4+CD25- T cells. The paper focuses on CTLA-4 and FoxP3 in Treg induction, with CD28 involved in the activation that generates these cells. CD28 signaling is required for the initial T cell activation that leads to CTLA-4 upregulation and Treg differentiation. Reason: While CD28 signaling contributes to T cell activation that can lead to Treg differentiation, this paper primarily focuses on CTLA-4 and FoxP3 as the key determinants of regulatory T cell function. CD28's role is in providing the initial activation signal, not specifically in directing the Treg differentiation program. This is a downstream consequence rather than a core function. Supporting Evidence: PMID:18641304 Activation of human CD4(+)CD25(-) T cells resulted in the appearance of a de novo population of FoxP3-expressing cells within 48 h. These cells expressed high levels of CTLA-4
GO:0005886 plasma membrane	TAS Reactome:R-HSA-167630	ACCEPT	Summary: Reactome annotation for Formation of Nef-CD28 cytoplasmic tail complex at the plasma membrane (HIV Nef pathway). Reason: CD28 is at the plasma membrane when Nef interacts with its cytoplasmic tail. Correct localization even in this pathological context.
GO:0005886 plasma membrane	TAS Reactome:R-HSA-167633	ACCEPT	Summary: Reactome annotation for Formation of Nef:CD28:Clathrin-Coated Pit Adapter Protein complex at the plasma membrane. Reason: Correct. CD28 at the plasma membrane in HIV Nef pathway.
GO:0005886 plasma membrane	TAS Reactome:R-HSA-167637	ACCEPT	Summary: Reactome annotation for internalization of the Nef:CD28:Clathrin complex from the plasma membrane. Reason: Correct. CD28 starts at the plasma membrane before Nef-mediated internalization.
GO:0005886 plasma membrane	TAS Reactome:R-HSA-388808	ACCEPT	Summary: Reactome annotation for CD28 homodimer binding B7-2 monomer at the plasma membrane. Reason: Correct. CD28-CD86 interaction occurs at the plasma membrane.
GO:0005886 plasma membrane	TAS Reactome:R-HSA-388811	ACCEPT	Summary: Reactome annotation for CD28 homodimer binding B7-1 homodimer at the plasma membrane. Reason: Correct. CD28-CD80 interaction occurs at the plasma membrane.
GO:0005515 protein binding	IPI PMID:15554700 Characterization of Lck-binding elements in the herpesviral ...	UNDECIDED	Summary: Bauer et al. 2004 characterized Lck-binding elements in the herpesviral Tip protein. The with/from column indicates Lck (P06239). This paper is about viral Tip protein interaction with Lck, not directly about CD28. CD28 may have been used as a reference or control for Lck binding. Reason: The paper is about herpesviral Tip protein interactions with Lck, not primarily about CD28. While CD28 does bind Lck via the PYAP motif, the annotation appears to be derived from a study focused on a viral protein. Cannot fully assess the relevance of this annotation to CD28 without seeing the specific CD28 data in the paper. Supporting Evidence: PMID:15554700 Herpesvirus saimiri encodes a tyrosine kinase interacting protein (Tip) that binds to T-cell-specific tyrosine kinase Lck
GO:0005515 protein binding	IPI PMID:11285224 Mechanism for down-regulation of CD28 by Nef.	UNDECIDED	Summary: Publication not available for review. With/from indicates P03406 (a viral protein). This may relate to viral protein interactions with CD28. Reason: Publication not available. The with/from column indicates a viral protein (P03406). Cannot assess the annotation without the publication.
GO:0032743 positive regulation of interleukin-2 production	IDA PMID:3875683 Involvement of T44 molecules in an antigen-independent pathw...	ACCEPT	Summary: Early publication (1985) demonstrating CD28 involvement in IL-2 production. CD28 costimulation is one of the best-characterized drivers of IL-2 production in T cells. IL-2 is the canonical cytokine whose production is enhanced by CD28 signaling. Publication not available. Reason: Positive regulation of IL-2 production is a core downstream effect of CD28 costimulation. IL-2 is the signature cytokine induced by CD28 costimulation - this is the classical readout of CD28 function. The deep research review confirms CD28 promotes IL-2 production. Supporting Evidence: file:human/CD28/CD28-deep-research-falcon.md increases proliferation and expression of various cytokines in particular IL2 production in both CD4(+) and CD8(+) T-cell subsets PMID:1650475 B-cell surface antigen B7 provides a costimulatory signal that induces T cells to proliferate and secrete interleukin 2
GO:0042102 positive regulation of T cell proliferation	IDA PMID:3159820 Human T cell activation. II. A new activation pathway used b...	ACCEPT	Summary: Early publication (1988) demonstrating CD28 involvement in T cell proliferation. One of the foundational studies establishing CD28 as a costimulatory molecule. Publication not available. Reason: Positive regulation of T cell proliferation is a core function of CD28. This is one of the earliest demonstrations. Consistent with extensive subsequent literature.
GO:0042802 identical protein binding	NAS PMID:3159820 Human T cell activation. II. A new activation pathway used b...	ACCEPT	Summary: NAS annotation for identical protein binding, reflecting CD28 homodimerization. CD28 exists as a disulfide-linked homodimer on the cell surface. The 2022 NMR structure (PMID:35397202) characterized the dimerization interface in the transmembrane domain. Reason: CD28 homodimerization is well established. It exists as a disulfide-linked homodimer, and the transmembrane domain contains dimerization motifs (GxxxA). While identical protein binding is somewhat generic, it accurately describes CD28 homodimerization. Supporting Evidence: file:human/CD28/CD28-uniprot.txt Homodimer; disulfide-linked file:human/CD28/CD28-deep-research-falcon.md CD28 is a ~44 kDa disulfide-linked Ig-domain homodimer
GO:0045840 positive regulation of mitotic nuclear division	IDA PMID:3159820 Human T cell activation. II. A new activation pathway used b...	MARK AS OVER ANNOTATED	Summary: IDA annotation for positive regulation of mitotic nuclear division from the same 1988 publication. CD28 costimulation drives T cell proliferation which involves mitotic division. Reason: While CD28 costimulation does drive T cell proliferation (which involves mitosis), annotating CD28 specifically to positive regulation of mitotic nuclear division is an over-annotation. CD28 is a signaling receptor, not a direct regulator of the mitotic machinery. The proliferative effect is a downstream consequence of transcriptional and metabolic reprogramming, not a direct regulation of mitosis. The more appropriate annotation is positive regulation of T cell proliferation.
GO:0005515 protein binding	IPI PMID:9784967 CD28/B7 costimulation: a review.	MODIFY	Summary: IPI for protein binding with CD80 (P33681). Publication not available. Likely a review or primary study documenting CD28-CD80 interaction. Reason: Protein binding is uninformative for the CD28-CD80 core receptor-ligand interaction. Should be captured under coreceptor activity. Proposed replacements: coreceptor activity
GO:0005515 protein binding	IPI PMID:9784967 CD28/B7 costimulation: a review.	MODIFY	Summary: IPI for protein binding with CD86 (P42081). Same publication as above. Reason: Protein binding is uninformative for CD28-CD86 core receptor-ligand interaction. Should be captured under coreceptor activity. Proposed replacements: coreceptor activity
GO:0001819 positive regulation of cytokine production	TAS PMID:8717514 CD28/B7 system of T cell costimulation.	ACCEPT	Summary: TAS annotation for positive regulation of cytokine production. CD28 costimulation promotes production of multiple cytokines including IL-2, IL-4, IL-10, and others. Publication not available. Reason: Positive regulation of cytokine production is a well-established consequence of CD28 costimulation. While this is a somewhat broad term, CD28 does promote production of multiple cytokines (IL-2, IL-4, IL-10, IFN-gamma) making this parent term appropriate. Supporting Evidence: file:human/CD28/CD28-deep-research-falcon.md increases proliferation and expression of various cytokines in particular IL2 production PMID:8617933 IL-4 synthesis was greatly enhanced by triggering of CD40L on the T cell surface in conjunction with ligation of CD3/TCR and CD28
GO:0005886 plasma membrane	TAS PMID:9784967 CD28/B7 costimulation: a review.	ACCEPT	Summary: TAS annotation for plasma membrane from a review. Correct and consistent with all other evidence. Reason: Correct. CD28 is a plasma membrane protein.
GO:0006959 humoral immune response	TAS PMID:8717514 CD28/B7 system of T cell costimulation.	KEEP AS NON CORE	Summary: TAS annotation for humoral immune response. CD28 costimulation on T cells promotes T helper function that supports B cell antibody responses. Publication not available. Reason: While CD28 costimulation of T helper cells supports humoral immune responses (by promoting T cell help for B cells), this is a distal, pleiotropic consequence of CD28 function rather than a core molecular function. CD28 does not directly participate in antibody production or B cell processes.
GO:0007166 cell surface receptor signaling pathway	TAS PMID:9784967 CD28/B7 costimulation: a review.	ACCEPT	Summary: TAS annotation for cell surface receptor signaling pathway. CD28 is a cell surface receptor that signals upon ligand engagement. This is a broad parent term. Reason: Correct but broad. CD28 is a cell surface receptor and its engagement initiates signaling cascades. More specific terms (T cell costimulation, positive regulation of TCR signaling) provide better detail.
GO:0015026 coreceptor activity	TAS PMID:9784967 CD28/B7 costimulation: a review.	ACCEPT	Summary: TAS annotation for coreceptor activity. CD28 is the prototypic T cell coreceptor that cooperates with the TCR to enable full T cell activation. The GO definition of coreceptor activity fits CD28 precisely: combining with a messenger and in cooperation with a nearby primary receptor, initiating a change in cell activity. Reason: Coreceptor activity is the most precise molecular function term for CD28. CD28 acts in cooperation with the TCR (the primary receptor) by binding B7 ligands (the extracellular messengers) to initiate costimulatory signaling. This is the single best MF term for CD28. Supporting Evidence: file:human/CD28/CD28-deep-research-falcon.md CD28 is the prototypic co-stimulatory receptor (signal 2) that integrates with TCR signalling (signal 1) to enable full T cell activation
GO:0042102 positive regulation of T cell proliferation	TAS PMID:9784967 CD28/B7 costimulation: a review.	ACCEPT	Summary: TAS annotation for positive regulation of T cell proliferation from a review. Consistent with IDA and IBA annotations for the same term. Reason: Correct TAS annotation consistent with experimental evidence (IDA, IBA) for the same core function.
GO:0045070 positive regulation of viral genome replication	NAS PMID:7492435 Costimulation of CD4+ T cells via CD28 modulates human immun...	KEEP AS NON CORE	Summary: NAS annotation for positive regulation of viral genome replication. This likely relates to CD28 costimulation enhancing HIV replication in T cells, as activated T cells provide a more permissive environment for viral replication. Publication not available. Reason: CD28 does not directly regulate viral genome replication. Rather, CD28-mediated T cell activation creates an environment permissive for viral replication (e.g., HIV in activated T cells). This is a highly indirect downstream consequence of T cell activation, not a core function of CD28.
GO:0045727 positive regulation of translation	NAS PMID:12028592 The regulation of protein synthesis and translation factors ...	KEEP AS NON CORE	Summary: NAS annotation for positive regulation of translation. CD28 signaling is known to activate mTOR-dependent translation programs. Publication not available. Reason: While CD28 signaling does promote mTOR-dependent translation (particularly of IL-2 and other cytokine mRNAs), this is a downstream signaling consequence rather than a core function. CD28 is a receptor, not a translation regulator. Supporting Evidence: file:human/CD28/CD28-deep-research-falcon.md CD28 signalling promotes glycolytic flux and mTOR-dependent translation

Core Functions

CD28 is the prototypic T cell coreceptor. It cooperates with the TCR by binding B7 ligands (CD80/CD86) on APCs to provide signal 2 for T cell activation. This is the single most precise molecular function term for CD28.

Molecular Function:

coreceptor activity

Directly Involved In:

T cell costimulation positive regulation of T cell receptor signaling pathway positive regulation of T cell proliferation positive regulation of interleukin-2 production

Cellular Locations:

external side of plasma membrane immunological synapse

References

GO_REF:0000002

Gene Ontology annotation through association of InterPro records with GO terms

GO_REF:0000033

Annotation inferences using phylogenetic trees

GO_REF:0000044

Gene Ontology annotation based on UniProtKB/Swiss-Prot Subcellular Location vocabulary mapping

GO_REF:0000052

Gene Ontology annotation based on curation of immunofluorescence data

GO_REF:0000107

Automatic transfer of experimentally verified manual GO annotation data to orthologs using Ensembl Compara

GO_REF:0000117

Electronic Gene Ontology annotations created by ARBA machine learning models

PMID:1650475

B-cell surface antigen B7 provides a costimulatory signal that induces T cells to proliferate and secrete interleukin 2.

Foundational demonstration of B7-CD28 costimulation driving T cell proliferation and IL-2 secretion

"B7-transfected CHO cells can induce suboptimally activated CD28+ T cells to proliferate and secrete high levels of interleukin 2"

PMID:3159820

Human T cell activation. II. A new activation pathway used by a major T cell population via a disulfide-bonded dimer of a 44 kilodalton polypeptide (9.3 antigen).

One of the earliest demonstrations of anti-CD28 antibody-mediated T cell costimulation

"mAb 9.3 was capable of inducing T cell proliferation in the presence of 12-o-tetradecanoyl phorbol-13-acetate (TPA)"

PMID:3875683

Involvement of T44 molecules in an antigen-independent pathway of T cell activation. Analysis of the correlations to the T cell antigen-receptor complex.

Demonstrated CD28 costimulation promotes IL-2 production

"triggering of T lymphocytes with 9.3 antibody (in the presence of adherent cells) resulted in strong IL-2 production that peaked at 48 h"

PMID:7544393

Identification of residues in the V domain of CD80 (B7-1) implicated in functional interactions with CD28 and CTLA4.

Identified hydrophobic residues in CD80 critical for CD28 and CTLA-4 binding

"Two hydrophobic residues in the V-like domain of CD80 were identified as critical for binding to CD28 and are also important for the interaction with CTLA4"
Y87 in CD80 is conserved across species and important for CD28 interaction

"One of these residues, Y87, is conserved in all CD80 and CD86 cloned from various species"

PMID:7568038

p56Lck and p59Fyn regulate CD28 binding to phosphatidylinositol 3-kinase, growth factor receptor-bound protein GRB-2, and T cell-specific protein-tyrosine kinase ITK: implications for T-cell costimulation.

Demonstrated Lck-dependent phosphorylation of CD28 Tyr-191

"p56Lck and p59Fyn phosphorylate CD28 primarily at Tyr-191 of the Tyr-Met-Asn-Met motif, inducing a 3- to 8-fold increase in p85 (subunit of PI 3-kinase) and GRB-2 SH2 binding to CD28"
CD28 binds PI3K, GRB2, and ITK through the YMNM motif

"CD28 interacts with three intracellular proteins-phosphatidylinositol 3-kinase (PI 3-kinase), T cell-specific protein-tyrosine kinase ITK (formerly TSK or EMT), and the complex between growth factor receptor-bound protein 2 and son of sevenless guanine nucleotide exchange protein (GRB-2-SOS)"

PMID:7584133

Selective CD28pYMNM mutations implicate phosphatidylinositol 3-kinase in CD86-CD28-mediated costimulation.

Y191 mutation disrupts both PI3K and GRB2 binding

"Y191 mutation (Y191CD28F) disrupted both PI 3-kinase and GRB-2 binding"
M194 mutation selectively disrupts PI3K binding

"M194 mutation (M194CD28C) disrupted only PI 3-kinase binding"
Both mutations abolished IL-2 production, directly implicating PI3K in CD28 costimulation

"Both Y191CD28F and M194CD28C mutants failed to generate IL-2. These data directly implicate PI 3-kinase in CD28-mediated costimulation leading to IL-2 secretion"

PMID:7807015

CTLA-4 binding to the lipid kinase phosphatidylinositol 3-kinase in T cells.

Compared CD28 and CTLA-4 binding to PI3K p85 SH2 domains

"CTLA-4 can also associate with PI 3-kinase as detected by lipid kinase analysis and immunoblotting with anti-p85 antiserum"
Similar binding affinity of p85 SH2 domains for CD28 and CTLA-4 cytoplasmic motifs

"the NH2- and COOH-terminal SH2 domains of p85 bind the CTLA-4 cytoplasmic pYVKM motif with an affinity (ID50: 0.6 and 0.04 microM), that is similar to CD28"

PMID:8617933

Cross-linking of the CD40 ligand on human CD4+ T lymphocytes generates a costimulatory signal that up-regulates IL-4 synthesis.

CD28 costimulation with CD3 and CD40L greatly enhances IL-4 synthesis

"IL-4 synthesis was greatly enhanced by triggering of CD40L on the T cell surface in conjunction with ligation of CD3/TCR and CD28"
CD28 alone with CD3 produces little IL-4 without CD40L

"ligation of CD3/TCR and CD28 in the absence of CD40L triggering resulted in little or no IL-4 synthesis"

PMID:8717514

CD28/B7 system of T cell costimulation.

CD28 promotes cytokine production and inhibits apoptosis

"The CD28/B7 receptor/ligand system is one of the dominant costimulatory pathways. Interruption of this signaling pathway with CD28 antagonists not only results in the suppression of the immune response, but in some cases induces antigen-specific tolerance"

PMID:9417079

Growth factor receptor-bound protein 2 SH2/SH3 domain binding to CD28 and its role in co-signaling.

Both GRB2 SH2 and SH3 domains contribute to CD28 binding

"the Grb2 SH2 domain is critical for the association, while the SH3 domain plays an additional role in facilitating optimal binding"
GRB2 binding to CD28 is required for IL-2 production and Vav phosphorylation

"Mutations that alter Grb2 binding were found to block the CD28-dependent interleukin-2 production. Further, tyrosine phosphorylation of Vav and the costimulation-dependent activation of Jun N-terminal kinase was blocked"
CD28/GRB2 interaction is required for JNK activation

"the costimulation-dependent activation of Jun N-terminal kinase was blocked in cells defective in CD28/Grb2 binding"

PMID:9784967

CD28/B7 costimulation: a review.

Comprehensive review of CD28 as a costimulatory receptor

"The current model of T cell activation requires two signals. The first signal is specific, requiring T cell receptor recognition and binding to MHC/Antigen presented by an antigen-presenting cell. The second signal is nonspecific, resulting from the binding of B7 ligand on the antigen-presenting cell with its receptor, CD28, on the T cell"

PMID:10820259

GRID: a novel Grb-2-related adapter protein that interacts with the activated T cell costimulatory receptor CD28.

PMID:11279501

Structural basis for co-stimulation by the human CTLA-4/B7-2 complex.

3.2A crystal structure of CTLA-4/B7-2 complex

"we report the 3.2-A resolution structure of the complex between the disulphide-linked homodimer of human CTLA-4 and the receptor-binding domain of human B7-2"
Provides structural basis for understanding CD28/CTLA-4 engagement with B7 ligands

"Regulation of T-cell activity is dependent on antigen-independent co-stimulatory signals provided by the disulphide-linked homodimeric T-cell surface receptors, CD28 and CTLA-4"

PMID:11285224

Mechanism for down-regulation of CD28 by Nef.

PMID:11390434

The transmembrane adaptor protein TRIM regulates T cell receptor (TCR) expression and TCR-mediated signaling via an association with the TCR zeta chain.

Confirmed CD28 cell surface expression on T cells

"T cell receptor (TCR)-interacting molecule (TRIM) is a recently identified transmembrane adaptor protein, which is exclusively expressed in T cells"

PMID:12196291

The interaction properties of costimulatory molecules revisited.

Characterized CD28 binding to CD80 and CD86

"B7-2 binds the two receptors more weakly than B7-1"
Demonstrated CD28 stimulation increases proliferation and cytokine expression

"relative to its CTLA-4 binding affinity, B7-2 binds CD28 2- to 3-fold more effectively than B7-1"

PMID:15067037

The modulation of CD40 ligand signaling by transmembrane CD28 splice variant in human T cells.

Isoform 3 (CD28i) interacts with CD40LG and enhances NF-kB and MAPK8/PAK2 activation

"In this study we show that CD28i, a transmembrane splice variant of CD28 costimulatory receptor, complexes with CD40L in human T cells"
CD28i localizes to the cell surface

"CD28i is unique among other isoforms as it retains an intact transmembrane region and cytoplasmic tail, but lacks the B7 ligand binding motif"

PMID:15554700

Characterization of Lck-binding elements in the herpesviral regulatory Tip protein.

Characterized Lck SH3 and SH2 domain binding properties (CD28 used as reference)

"Herpesvirus saimiri encodes a tyrosine kinase interacting protein (Tip) that binds to T-cell-specific tyrosine kinase Lck via multiple sequence motifs and controls its activity"

PMID:18337562

CCL5-mediated T-cell chemotaxis involves the initiation of mRNA translation through mTOR/4E-BP1.

Focus on CCL5 signaling in T cells, not directly on CD28

"we examined the role for CCL5-mediated initiation of mRNA translation in CD4(+) T-cell chemotaxis"

PMID:18641304

Acquisition of suppressive function by activated human CD4+ CD25- T cells is associated with the expression of CTLA-4 not FoxP3.

CTLA-4 is the key determinant for Treg suppressive function

"the acquisition of suppressive behavior by activated CD4(+)CD25(-) T cells requires the expression of CTLA-4"
CD28-dependent T cell activation leads to CTLA-4 and FoxP3 induction

"Activation of human CD4(+)CD25(-) T cells resulted in the appearance of a de novo population of FoxP3-expressing cells within 48 h. These cells expressed high levels of CTLA-4"

PMID:18641334

ICOS ligation recruits the p50alpha PI3K regulatory subunit to the immunological synapse.

PMID:21982860

A secreted protein microarray platform for extracellular protein interaction discovery.

High-throughput platform identified CD28-CD80 interaction

"functional protein microarrays are particularly well-suited for high-throughput screening of extracellular protein interactions"

PMID:23817958

Proinflammatory stimuli induce galectin-9 in human mesenchymal stromal cells to suppress T-cell proliferation.

CD28 signaling regulates both positive and negative gene expression programs

"MSCs with a galectin-9 knockdown lost a significant portion of their antiproliferative effects on T cells"

PMID:23918985

Soluble CD80 restores T cell activation and overcomes tumor cell programmed death ligand 1-mediated immune suppression.

Direct evidence for CD28 in T cell activation

"CD80-Fc facilitates T cell activation by binding to PDL1 to inhibit PDL1-PD1 interactions and by costimulating through CD28"

PMID:32822567

A Human IgSF Cell-Surface Interactome Reveals a Complex Network of Protein-Protein Interactions.

Systematic interactome identified CD28-CD80 and CD28-CD86 interactions

"Cell-surface protein-protein interactions (PPIs) mediate cell-cell communication, recognition, and responses"

PMID:35397202

Structural characterization of a dimerization interface in the CD28 transmembrane domain.

NMR structure of CD28 TM domain confirming dimerization

"we determined the dimeric helix-helix packing of CD28-TMH using nuclear magnetic resonance (NMR) technology"
GxxxA motif mediates TM helix dimerization

"a GxxxA motif, which is highly conserved in many dimeric assemblies, is located at the dimerization interface"
Mutations in dimerization interface reduce IL-2 release

"Mutating G160 and A164 can disrupt the transmembrane helix assembly and reduces CD28 enhancement in cells"

PMID:35922511

A physical wiring diagram for the human immune system.

Systematic mapping of immune cell surface interactions including CD28-CD80 and CD28-CD86

"using a high-throughput surface receptor screening method, we systematically mapped the direct protein interactions across a recombinant library that encompasses most of the surface proteins that are detectable on human leukocytes"

PMID:7492435

Costimulation of CD4+ T cells via CD28 modulates human immunodeficiency virus type 1 infection and replication in vitro.

PMID:8146197

T-cell antigen CD28 interacts with the lipid kinase phosphatidylinositol 3-kinase by a cytoplasmic Tyr(P)-Met-Xaa-Met motif.

PMID:8183372

Binding of phosphatidylinositol-3-OH kinase to CD28 is required for T-cell signalling.

PMID:12028592

The regulation of protein synthesis and translation factors by CD3 and CD28 in human primary T lymphocytes.

Reactome:R-HSA-389356

Co-stimulation by CD28 (Reactome pathway)

Comprehensive pathway covering CD28 signaling including PI3K, Vav1, Grb2, Gads recruitment

"CD28 signaling plays a critical role in shaping immune responses by ensuring effective T-cell activation and enhancing T-cell survival and proliferation"

Reactome:R-HSA-164939

Nef mediated downregulation of CD28 cell surface expression

HIV Nef hijacks CD28 for internalization

Reactome:R-HSA-2316434

PI3K phosphorylation of PIP2 to PIP3 in CD28 signaling

Reactome:R-HSA-2400009

PI3K inhibitors in CD28 signaling

Reactome:R-HSA-388814

Grb2 binding to CD28

Reactome:R-HSA-388831

Phosphorylation of CD28

Reactome:R-HSA-388832

PI3K binding to CD28

Reactome:R-HSA-389158

CD28-bound PI3K phosphorylation of PIP2 to PIP3

Reactome:R-HSA-389348

Activation of Rac1 by pVav1 in CD28 pathway

Reactome:R-HSA-389350

Activation of Cdc42 by pVav1 in CD28 pathway

Reactome:R-HSA-389352

Translocation of Vav1 to CD28

Reactome:R-HSA-389354

Activation of Vav1 in CD28 pathway

Reactome:R-HSA-389381

Gads binding to CD28

Reactome:R-HSA-167630

Formation of Nef-CD28 cytoplasmic tail complex

Reactome:R-HSA-167633

Formation of Nef-CD28-Clathrin-Coated Pit Adapter Protein complex

Reactome:R-HSA-167637

Internalization of Nef-CD28-Clathrin complex

Reactome:R-HSA-388808

CD28 homodimer binding B7-2 monomer

Reactome:R-HSA-388811

CD28 homodimer binding B7-1 homodimer

file:human/CD28/CD28-deep-research-falcon.md

Deep research review of CD28 biology (2024 literature synthesis)

Comprehensive review synthesizing 2024 literature on CD28 signaling, structure, and translational applications
YMNM/PRRP/PYAP motifs recruit PI3K/GRB2, ITK/PLCgamma1, and Lck/PKCtheta
CD28 costimulation accelerates Lck recruitment and ZAP70 activation at TCR microclusters

Deep Research

Falcon

(CD28-deep-research-falcon.md)

this with annotations you find in gene/protein databases, but these can be outdated or inaccurate. Falcon Edison Scientific Literature 12 citations 2026-02-14T14:40:44.590859

The research report should be a detailed narrative explaining the function, biological processes, and localization of the gene product. Citations should be given for all claims.

You should prioritize authoritative reviews and primary scientific literature when conducting research. You can supplement
this with annotations you find in gene/protein databases, but these can be outdated or inaccurate.

We are specifically interested in the primary function of the gene - for enzymes, what reaction is catalyzed, and what is the substrate specificity? For transporters, what is the substrate? For structural proteins or adapters, what is the broader structural role? For signaling molecules, what is the role in the pathway.

We are interested in where in or outside the cell the gene product carries out its function.

We are also interested in the signaling or biochemical pathways in which the gene functions. We are less interested in broad pleiotropic effects, except where these elucidate the precise role.

Include evidence where possible. We are interested in both experimental evidence as well as inference from structure, evolution, or bioinformatic analysis. Precise studies should be prioritized over high-throughput, where available.

Comprehensive research report: CD28 (human; UniProt P10747)

Plan and verification of target identity
- Verification. The target is human CD28 (CD28 gene; UniProt P10747), a T cell–specific surface glycoprotein. Recent authoritative reviews describe CD28 as a disulfide‑linked, homodimeric immunoglobulin superfamily receptor with an extracellular V‑set Ig domain containing the canonical MYPPPY motif for B7 ligand binding, consistent with the UniProt description and InterPro domain annotations (Ig-like_fold, Ig_V-set) (lotze2024cd28costimulationnovel pages 1-3, ciesielskafiglon2024theroleof pages 4-5). Organism is Homo sapiens; all sources cited are human-focused reviews or experimental studies.
- Family and domains. CD28 belongs to the CD28/CTLA‑4 family of Ig superfamily receptors that engage B7 ligands (CD80/CD86); structural and transmembrane determinants for dimerization have been mapped, aligning with the domain architecture noted in UniProt (lotze2024cd28costimulationnovel pages 3-4, lotze2024cd28costimulationnovel pages 1-3).

Key concepts and definitions (current understanding)
- Molecular identity and expression. CD28 is a ~44 kDa disulfide‑linked Ig-domain homodimer on T cells. It is expressed on nearly all human CD4+ T cells (≈95–100%) and on a substantial subset of CD8+ T cells (≈30–50%), with estimates of ~60,000 molecules per T cell; the extracellular MYPPPY motif mediates ligand binding to CD80/CD86 (International Journal of Molecular Sciences, 2024; https://doi.org/10.3390/ijms25021274) (ciesielskafiglon2024theroleof pages 4-5).
- Ligands and competition with CTLA‑4. The principal ligands for CD28 are CD80 (B7‑1) and CD86 (B7‑2). The inhibitory receptor CTLA‑4 shares these ligands and binds them with higher affinity, limiting CD28 co‑stimulation by outcompeting or removing B7 ligands (Nature Reviews Immunology, 2024; https://doi.org/10.1038/s41577-024-01061-1) (lotze2024cd28costimulationnovel pages 1-3).
- Subcellular localization and immune synapse. Upon antigen recognition, CD28 co‑localizes with TCR microclusters within the immune synapse. Recent live-cell imaging demonstrates that CD28 co‑stimulation increases Lck recruitment to TCR microclusters, thereby accelerating ZAP70 recruitment/activation and effectively lowering the TCR activation threshold (Frontiers in Immunology, 2024; https://doi.org/10.3389/fimmu.2024.1503018) (raychaudhuri2024cd28shapest pages 1-2).
- Intracellular motifs and canonical signalling. CD28’s cytoplasmic tail contains three key motifs with distinct adaptor recruitment: YMNM (binds PI3K and GRB2), PRRP (recruits ITK leading to PLCγ1 and SLP‑76/GADS engagement), and PYAP (dominant for LCK binding and assembly of PKCθ at the synapse). These converge on NF‑κB, AP‑1, and NFAT transcriptional programs and mTOR‑dependent translation to drive IL‑2 production, survival, and proliferation (Nature Reviews Immunology, 2024; https://doi.org/10.1038/s41577-024-01061-1) (lotze2024cd28costimulationnovel pages 3-4).

Recent developments and latest research (2023–2024 prioritized)
- Proximal signalling dynamics at microclusters. TIRF microscopy in 2024 demonstrated that CD28 costimulation specifically accelerates Lck recruitment and ZAP70 activation within TCR microclusters, providing a mechanistic basis for threshold-lowering of TCR activation by CD28 (Frontiers in Immunology, Dec 2024; https://doi.org/10.3389/fimmu.2024.1503018) (raychaudhuri2024cd28shapest pages 1-2).
- Updated mechanistic map of CD28 signalling. A 2024 Nature Reviews Immunology article synthesized structural and proteomic insights: transmembrane dimerization motifs (e.g., GxxxA) influence homodimer assembly and signalling; membrane association of the cytoplasmic domain regulates Lck access; YMNM/PRRP/PYAP motifs recruit PI3K/GRB2, ITK–PLCγ1, and Lck/PKCθ respectively; and cis-signalling via B7 at invaginated membranes may sustain tonic signalling after trans‑engagement (Nature Reviews Immunology, Jul 2024; https://doi.org/10.1038/s41577-024-01061-1) (lotze2024cd28costimulationnovel pages 3-4, lotze2024cd28costimulationnovel pages 19-20).
- Quantitative immunobiology of expression and ligand engagement. A 2024 review collated human expression estimates (CD4+ ≈95–100%; CD8+ ≈30–50%) and reiterated the MYPPPY motif requirement for CD80/CD86 binding, with dynamic modulation of CD28 surface expression upon ligand engagement (International Journal of Molecular Sciences, Jan 2024; https://doi.org/10.3390/ijms25021274) (ciesielskafiglon2024theroleof pages 4-5).

Current applications and real-world implementations
- Costimulation blockade with CTLA‑4–Ig biologics. Abatacept and belatacept are CTLA‑4–Ig fusion proteins that sequester CD80/CD86, preventing CD28 ligation. These agents are used clinically for autoimmunity and transplantation (belatacept as a calcineurin inhibitor–sparing regimen), reflecting the central role of the B7–CD28 axis in alloimmunity; translational reviews emphasize efficacy, trade‑offs (e.g., acute rejection risk), and the rationale for strategies that spare CTLA‑4 function (Nature Reviews Immunology, 2024; https://doi.org/10.1038/s41577-024-01061-1) (lotze2024cd28costimulationnovel pages 1-3).
- CD28 in engineered cell therapies. Incorporating a CD28 intracellular domain in CAR constructs enhances proximal signalling and effector programs. In 2024, CAR‑NK cells with a CD28 costimulatory domain exhibited superior serial killing (≈25% serial killers vs ≈8% without CD28), improved in vivo tumor control across models, and increased persistence and tumor infiltration via an LCK/CD3ζ/ZAP70 axis (Cancer Discovery, Jul 2024; https://doi.org/10.1158/2159-8290.cd-24-0096) (acharya2024cd28costimulationaugments pages 6-8).
- Safety lessons for systemic CD28 agonism. The CD28 “superagonist” TGN1412 caused severe cytokine release syndrome in a 2006 first‑in‑human trial due to systemic activation of human effector memory T cells; contemporary reviews highlight tumor‑restricted or conditionally active CD28 agonists as safer avenues (Nature Reviews Immunology, 2024; https://doi.org/10.1038/s41577-024-01061-1) (lotze2024cd28costimulationnovel pages 3-4).
- Small‑molecule CD28 antagonists (emerging). A 2025 preprint reported discovery of a small molecule (22VS) that binds CD28 (Kd ≈52 µM), disrupts CD28–CD80 engagement (ELISA IC50 ≈7.8 µM), and suppresses T cell cytokines in tumor/mucosal co‑cultures; while preclinical, it demonstrates feasibility beyond biologics (bioRxiv, Jul 2025; https://doi.org/10.1101/2025.07.16.665260) (upadhyay2025smallmoleculebasedblockade pages 21-26).

Expert opinions and analysis from authoritative sources
- Synthesis of CD28 structure–function–translation. A 2024 Nature Reviews Immunology perspective integrates biophysics (transmembrane dimerization, cytoplasmic membrane association), proximal signalling (Lck recruitment; YMNM/PRRP/PYAP networks), transcriptional outputs (NF‑κB/AP‑1/NFAT; mTOR), and translational strategies (CTLA‑4–Ig, tumor‑restricted CD28 agonists, CAR design), offering a consensus framework for CD28 biology and its therapeutic modulation (https://doi.org/10.1038/s41577-024-01061-1) (lotze2024cd28costimulationnovel pages 3-4, lotze2024cd28costimulationnovel pages 19-20).
- Mechanistic proof of microcluster control. The 2024 imaging study provides direct, quantitative evidence that CD28 reprograms proximal TCR signalling kinetics by modulating Lck and ZAP70 dynamics at the immune synapse, a key expert insight for designing synapse‑proximal therapeutics (https://doi.org/10.3389/fimmu.2024.1503018) (raychaudhuri2024cd28shapest pages 1-2).

Relevant statistics and data (recent studies)
- Human expression frequencies: CD28 on ≈95–100% of CD4+ and ≈30–50% of CD8+ T cells; ~60,000 molecules per T cell; MYPPPY motif mediates B7 binding (Jan 2024; URL above) (ciesielskafiglon2024theroleof pages 4-5).
- CAR‑NK functional gains with CD28 domain: serial killer fraction increased from ≈8% to ≈25%; enhanced in vivo tumor control with increased persistence and tumor infiltration (Jul 2024; URL above) (acharya2024cd28costimulationaugments pages 6-8).
- Small‑molecule antagonism: CD28–ligand disruption IC50 ≈7.8 µM (ELISA); binding Kd ≈52 µM (TRIC/MST); dose‑dependent suppression of IFN‑γ/IL‑2/TNF‑α in co‑cultures (Jul 2025; URL above) (upadhyay2025smallmoleculebasedblockade pages 21-26).

Mechanistic narrative: primary function, pathways, and localization
- Primary role. CD28 is the prototypic co‑stimulatory receptor (signal 2) that integrates with TCR signalling (signal 1) to enable full T cell activation. It stabilizes IL‑2 transcription and mRNA, promotes survival (e.g., Bcl‑xL upregulation), and drives clonal expansion and effector differentiation through PI3K/AKT, MAPK, and NF‑κB/AP‑1/NFAT programmes (lotze2024cd28costimulationnovel pages 3-4, ciesielskafiglon2024theroleof pages 4-5).
- Subcellular site of action. CD28 operates at the T cell–APC immune synapse, co‑assembling within TCR microclusters. CD28 co‑stimulation increases recruitment of active Lck, which accelerates ZAP70 engagement on TCRζ, thus lowering activation thresholds and amplifying proximal signalling (raychaudhuri2024cd28shapest pages 1-2).
- Biochemical motifs and adapters. The YMNM motif recruits PI3K and GRB2 to support AKT and MAPK cascades; PRRP engages ITK to potentiate PLCγ1 and adaptor complexes (SLP‑76/GADS); PYAP binds Lck, nucleating PKCθ in the synapse to drive NF‑κB activation. Together these motifs explain CD28’s unique ability to couple membrane‑proximal events to transcriptional and translational programmes, including mTOR‑dependent metabolic reprogramming (lotze2024cd28costimulationnovel pages 3-4).
- Metabolic integration. CD28 signalling promotes glycolytic flux and mTOR‑dependent translation, supporting rapid proliferation and effector function; recent syntheses connect sustained CD28 tone with maintenance and differentiation of effector programs, highlighting the importance of signal strength and duration (lotze2024cd28costimulationnovel pages 3-4).

Translational landscape and safety
- B7–CD28 axis blockade. CTLA‑4–Ig biologics (abatacept, belatacept) clinically block CD28 ligation by sequestering CD80/CD86 and are used to prevent GVHD or as immunosuppression in transplantation; balancing efficacy with risks (e.g., acute rejection) motivates next‑generation approaches that spare CTLA‑4 function while limiting CD28 (lotze2024cd28costimulationnovel pages 1-3).
- Engineered costimulation. Incorporating CD28 ICDs into CARs augments antitumor efficacy but also shapes metabolism, persistence, and toxicity risks; mechanistic data in 2024 demonstrate that adding CD28 to CAR‑NK cells engages an Lck/CD3ζ/ZAP70 axis, boosting serial killing, persistence, and tumor infiltration (acharya2024cd28costimulationaugments pages 6-8).
- Agonism cautions. The TGN1412 incident remains a sentinel safety case against systemic, unconditioned CD28 agonism; contemporary strategies emphasize tumor‑selective bispecifics and trispecifics that confine CD28 activation to the tumor microenvironment (lotze2024cd28costimulationnovel pages 3-4).
- Small molecules (future direction). First reports of CD28‑targeted small molecules indicate tractability of orthosteric/allosteric antagonism with measurable biochemical and functional potency in human cell models, expanding modalities beyond biologics (upadhyay2025smallmoleculebasedblockade pages 21-26).

Embedded summary table of recent evidence
| Topic | Key finding (1–2 sentences) | Mechanism / Motif | Application / Implication | Year | Source (journal) | URL | Citation ID |
|---|---|---|---:|---:|---|---|---|
| Identity / structure & MYPPPY motif | CD28 is a ~44 kDa disulfide‑linked Ig‑domain homodimer with an extracellular MYPPPY motif that mediates B7 (CD80/CD86) binding; highly expressed on CD4+ T cells and present on subsets of CD8+ T cells. | Extracellular V‑set Ig domain with MYPPPY motif; TM dimerization motifs (GxxxA/YxxxxT) influence dimerization. | Defines ligand engagement surface for antibodies, CTLA4‑Ig therapeutics and CAR hinge/transmembrane design. | 2024 | International Journal of Molecular Sciences | https://doi.org/10.3390/ijms25021274 | pqac-00000001 |
| Signaling motifs (YMNM, PRRP, PYAP) and downstream effectors | The CD28 intracellular domain contains YMNM, PRRP and PYAP motifs that recruit distinct signalling adaptors to drive PI3K/AKT, GRB2/Mapk and LCK/PKCθ→NF‑κB pathways, promoting IL‑2, survival and proliferation. | YMNM → PI3K/Grb2; PRRP → ITK/PLCγ → SLP‑76/GADS; PYAP → LCK recruitment and PKCθ assembly. | Mechanistic basis for modulating co‑stimulation (agonists/antagonists) and for selecting ICDs in CAR constructs. | 2024 | Nature Reviews Immunology | https://doi.org/10.1038/s41577-024-01061-1 | pqac-00000002 |
| Microcluster / Lck → ZAP70 mechanism | CD28 co‑stimulation accelerates recruitment of Lck into TCR microclusters, which in turn accelerates ZAP70 recruitment and activation, effectively lowering the TCR activation threshold. | Enhanced Lck recruitment/activation and spatial segregation in TCR microclusters → faster ZAP70 phosphorylation. | Explains how CD28 augments proximal TCR signalling and informs synapse‑targeted immunotherapy/CAR tuning. | 2024 | Frontiers in Immunology | https://doi.org/10.3389/fimmu.2024.1503018 | pqac-00000000 |
| Biology / ligands (CD80/CD86) and CTLA‑4 competition | CD80 (B7‑1) and CD86 (B7‑2) are the principal ligands for CD28; CTLA‑4 binds the same ligands with higher affinity to limit CD28 signalling (ligand scavenging and trans/cis interactions). | Trans‑ and cis‑B7:CD28/CTLA‑4 interactions; CTLA‑4 outcompetes CD28 for CD80/CD86 binding. | Basis for CTLA‑4‑Ig (abatacept/belatacept) and for targeting the B7–CD28 axis in transplantation and cancer. | 2024 | Nature Reviews Immunology | https://doi.org/10.1038/s41577-024-01061-1 | pqac-00000004 |
| Translational: CTLA4‑Ig (abatacept / belatacept) in GVHD / transplantation | CTLA4‑Ig fusion proteins block CD80/CD86 → CD28 interactions and are used to prevent GVHD and in transplant immunosuppression, offering steroid/CNI‑sparing approaches but with trade‑offs (e.g., belatacept and acute rejection risk). | CTLA‑4 extracellular domain fused to Fc sequesters CD80/CD86, preventing CD28 costimulation. | Clinically applied costimulation blockade for autoimmunity/transplantation; motivates selective CD28 antagonists that spare CTLA‑4 function. | 2024 (reviewed) | Nature Reviews Immunology | https://doi.org/10.1038/s41577-024-01061-1 | pqac-00000004 |
| CAR / NK engineering using CD28 ICD | Incorporation of CD28 intracellular domains into CARs (and into engineered NK CARs) enhances cytotoxic effector programs, persistence, serial killing and tumor control by recruiting LCK/CD3ζ/ZAP70 signalling cascades. | CD28 ICD recruits LCK → CD3ζ/ZAP70 axis and downstream activation (enhanced perforin/granzyme, survival TFs). | Rational selection/modification of CAR co‑stimulatory domains to improve potency and persistence (but impacts toxicity/metabolism). | 2024 | Cancer Discovery | https://doi.org/10.1158/2159-8290.cd-24-0096 | pqac-00000006 |
| Superagonist (TGN1412) safety lesson | The CD28 superagonist TGN1412 triggered catastrophic cytokine release in humans (2006), revealing species differences in CD28 biology and the danger of systemic CD28 hyperagonism. | Systemic agonism of CD28 on human effector memory T cells → widespread activation/CRS. | Critical caution for CD28 agonist strategies; motivates conditioned/targeted costimulation (bispecifics, tumor‑restricted agonists). | historical, reviewed 2024 | Nature Reviews Immunology | https://doi.org/10.1038/s41577-024-01061-1 | pqac-00000002 |
| Small‑molecule CD28 antagonism (proof‑of‑concept) | Structure‑guided screening identified small molecules (lead 22VS) that bind CD28, disrupt CD28–CD80 binding (ELISA IC50 ~7–8 µM; Kd ~52 µM) and suppress T cell cytokine responses in co‑culture models. | Small molecule binds a pocket near extracellular residues to inhibit ligand engagement; cellular NanoBit assays show partial inhibition of membrane interactions. | Demonstrates feasibility of non‑biologic CD28 pathway modulation for immunosuppression or inflammation control; preclinical (preprint) stage. | 2025 (preprint) | bioRxiv (preprint) | https://doi.org/10.1101/2025.07.16.665260 | pqac-00000005 |
| CD28 and T cell metabolism / maintenance | CD28 costimulation activates mTOR‑dependent translation and metabolic programs (glycolysis/mitochondrial fitness) that support proliferation, differentiation and maintenance of T cell subsets; sustained CD28 signals maintain stem‑like PD‑1+ T cells. | Downstream PI3K/mTOR and related metabolic reprogramming (promotes glycolysis vs mitochondrial fitness depending on signal strength). | Links costimulation to metabolic interventions and informs temporal modulation of CD28 in chronic infection/cancer and CAR design to balance efficacy and durability. | 2024 | Nature Reviews Immunology (reviewed evidence) | https://doi.org/10.1038/s41577-024-01061-1 | pqac-00000002 |

Table: Concise table summarizing recent (2023–2024) mechanistic and translational evidence for human CD28 (UniProt P10747), linking structure, motifs and signalling to therapeutic applications; each row cites the source ID for use in the main report.

Conclusion
Recent high‑resolution mechanistic work establishes that CD28 co‑stimulation shapes proximal TCR signalling by recruiting active Lck and accelerating ZAP70 activation within microclusters, while canonical YMNM/PRRP/PYAP motifs engage PI3K/GRB2, ITK/PLCγ1, and Lck/PKCθ pathways to drive NF‑κB/AP‑1/NFAT and mTOR‑dependent programs. These insights, together with quantitative human expression and ligand biology (CD80/CD86; CTLA‑4 competition), underpin the clinical success of costimulation blockade (CTLA‑4–Ig), inform rational costimulatory engineering in CAR and emerging CAR‑NK therapies, and caution against systemic agonism while motivating tumor‑restricted activation and small‑molecule antagonists (raychaudhuri2024cd28shapest pages 1-2, ciesielskafiglon2024theroleof pages 4-5, lotze2024cd28costimulationnovel pages 3-4, lotze2024cd28costimulationnovel pages 1-3, acharya2024cd28costimulationaugments pages 6-8, lotze2024cd28costimulationnovel pages 19-20, upadhyay2025smallmoleculebasedblockade pages 21-26).

References

(lotze2024cd28costimulationnovel pages 1-3): Michael T. Lotze, Scott H. Olejniczak, and Dimitris Skokos. Cd28 co-stimulation: novel insights and applications in cancer immunotherapy. Nature reviews. Immunology, 24:878-895, Jul 2024. URL: https://doi.org/10.1038/s41577-024-01061-1, doi:10.1038/s41577-024-01061-1. This article has 52 citations.
(ciesielskafiglon2024theroleof pages 4-5): Klaudia Ciesielska-Figlon and Katarzyna A. Lisowska. The role of the cd28 family receptors in t-cell immunomodulation. International Journal of Molecular Sciences, 25:1274, Jan 2024. URL: https://doi.org/10.3390/ijms25021274, doi:10.3390/ijms25021274. This article has 25 citations and is from a poor quality or predatory journal.
(lotze2024cd28costimulationnovel pages 3-4): Michael T. Lotze, Scott H. Olejniczak, and Dimitris Skokos. Cd28 co-stimulation: novel insights and applications in cancer immunotherapy. Nature reviews. Immunology, 24:878-895, Jul 2024. URL: https://doi.org/10.1038/s41577-024-01061-1, doi:10.1038/s41577-024-01061-1. This article has 52 citations.
(raychaudhuri2024cd28shapest pages 1-2): Kumarkrishna Raychaudhuri, Rohita Rangu, Alison Ma, Neriah Alvinez, Andy D. Tran, Sandeep Pallikkuth, Katherine M. McIntire, Joseph A. Garvey, Jason Yi, and Lawrence E. Samelson. Cd28 shapes t cell receptor signaling by regulating lck dynamics and zap70 activation. Frontiers in Immunology, Dec 2024. URL: https://doi.org/10.3389/fimmu.2024.1503018, doi:10.3389/fimmu.2024.1503018. This article has 4 citations and is from a peer-reviewed journal.
(lotze2024cd28costimulationnovel pages 19-20): Michael T. Lotze, Scott H. Olejniczak, and Dimitris Skokos. Cd28 co-stimulation: novel insights and applications in cancer immunotherapy. Nature reviews. Immunology, 24:878-895, Jul 2024. URL: https://doi.org/10.1038/s41577-024-01061-1, doi:10.1038/s41577-024-01061-1. This article has 52 citations.
(acharya2024cd28costimulationaugments pages 6-8): Sunil Acharya, Rafet Basar, May Daher, Hind Rafei, Ping Li, Nadima Uprety, Emily Ensley, Mayra Shanley, Bijender Kumar, Pinaki P. Banerjee, Luciana Melo Garcia, Paul Lin, Vakul Mohanty, Kun H. Kim, Xianli Jiang, Yuchen Pan, Ye Li, Bin Liu, Ana K. Nunez Cortes, Chenyu Zhang, Mohsen Fathi, Ali Rezvan, Melisa J. Montalvo, Sophia L. Cha, Francia Reyes-Silva, Rejeena Shrestha, Xingliang Guo, Kiran Kundu, Alexander Biederstädt, Luis Muniz-Feliciano, Gary M. Deyter, Mecit Kaplan, Xin R. Jiang, Enli Liu, Antrix Jain, Janos Roszik, Natalie W. Fowlkes, Luisa M. Solis Soto, Maria G. Raso, Joseph D. Khoury, Pei Lin, Francisco Vega, Navin Varadarajan, Ken Chen, David Marin, Elizabeth J. Shpall, and Katayoun Rezvani. Cd28 costimulation augments car signaling in nk cells via the lck/cd3ζ/zap70 signaling axis. Cancer Discovery, 14:1879-1900, Jul 2024. URL: https://doi.org/10.1158/2159-8290.cd-24-0096, doi:10.1158/2159-8290.cd-24-0096. This article has 51 citations and is from a highest quality peer-reviewed journal.
(upadhyay2025smallmoleculebasedblockade pages 21-26): Saurabh Upadhyay, Valerij Talagayev, Sungwoo Cho, Gerhard Wolber, and Moustafa T. Gabr. Small molecule-based blockade of cd28 suppresses t cell costimulation across cellular and mucosal co-culture models. bioRxiv : the preprint server for biology, Jul 2025. URL: https://doi.org/10.1101/2025.07.16.665260, doi:10.1101/2025.07.16.665260. This article has 4 citations.

Citations

ciesielskafiglon2024theroleof pages 4-5
upadhyay2025smallmoleculebasedblockade pages 21-26
https://doi.org/10.3390/ijms25021274
https://doi.org/10.1038/s41577-024-01061-1
https://doi.org/10.3389/fimmu.2024.1503018
https://doi.org/10.1158/2159-8290.cd-24-0096
https://doi.org/10.1101/2025.07.16.665260
https://doi.org/10.1038/s41577-024-01061-1,
https://doi.org/10.3390/ijms25021274,
https://doi.org/10.3389/fimmu.2024.1503018,
https://doi.org/10.1158/2159-8290.cd-24-0096,
https://doi.org/10.1101/2025.07.16.665260,

📄 View Raw YAML

id: P10747
gene_symbol: CD28
product_type: PROTEIN
status: IN_PROGRESS
taxon:
  id: NCBITaxon:9606
  label: Homo sapiens
description: >-
  CD28 is the prototypic T cell co-stimulatory receptor (signal 2) that synergizes with TCR engagement
  (signal 1) to enable full T cell activation. It is a ~44 kDa disulfide-linked homodimer of the
  immunoglobulin superfamily, containing an extracellular IgV-type domain with a conserved MYPPPY motif
  that mediates binding to its ligands CD80 (B7-1) and CD86 (B7-2) on antigen-presenting cells. CD28 is
  expressed on nearly all human CD4+ T cells and a substantial fraction of CD8+ T cells. Upon ligation,
  the cytoplasmic tail recruits signaling molecules via three key motifs: YMNM (binds PI3K p85 subunit
  and GRB2), PRRP (recruits ITK leading to PLCgamma1 and SLP-76/GADS engagement), and PYAP (binds Lck
  and assembles PKC-theta at the synapse). These converge on NF-kB, AP-1, and NFAT transcriptional
  programs and mTOR-dependent metabolic reprogramming to drive IL-2 production, T cell survival,
  proliferation, and effector differentiation. The inhibitory receptor CTLA-4 competes for the same B7
  ligands with higher affinity, providing a critical negative regulatory checkpoint. Inherited CD28
  deficiency causes susceptibility to HPV-related verrucosis (IMD123). CD28 localizes to the immune
  synapse at the T cell-APC interface, where it accelerates Lck recruitment to TCR microclusters and
  lowers the TCR activation threshold.
alternative_products:
- name: '1'
  id: P10747-1
- name: 2 (CD28-S2)
  id: P10747-2
  sequence_note: VSP_002494
- name: 3 (CD28i)
  id: P10747-3
  sequence_note: VSP_002495
- name: 4 (CD28-S1)
  id: P10747-4
  sequence_note: VSP_002496
- name: '5'
  id: P10747-5
  sequence_note: VSP_002495, VSP_002497, VSP_002498
- name: 6 (CD28-S3)
  id: P10747-6
  sequence_note: VSP_002495, VSP_002499
- name: '7'
  id: P10747-7
  sequence_note: VSP_047701
existing_annotations:
# ========================
# IBA ANNOTATIONS
# ========================
- term:
    id: GO:0042102
    label: positive regulation of T cell proliferation
  evidence_type: IBA
  original_reference_id: GO_REF:0000033
  review:
    summary: >-
      CD28 costimulation is one of the best-characterized drivers of T cell proliferation. Anti-CD28
      antibodies costimulate T cell proliferation in conjunction with TCR/CD3 signaling (PMID:3159820).
      The deep research review confirms CD28 promotes clonal expansion through PI3K/AKT, MAPK, and
      NF-kB/AP-1/NFAT programmes.
    action: ACCEPT
    reason: >-
      Positive regulation of T cell proliferation is a core function of CD28. This IBA annotation is
      well supported by extensive experimental literature showing CD28 costimulation is essential for
      T cell proliferation. The term is at an appropriate level of specificity.
    supported_by:
    - reference_id: PMID:3159820
      supporting_text: "mAb 9.3 was capable of inducing T cell proliferation in the presence of 12-o-tetradecanoyl phorbol-13-acetate (TPA)"
    - reference_id: file:human/CD28/CD28-deep-research-falcon.md
      supporting_text: "CD28 is the prototypic co\u2011stimulatory receptor (signal 2) that integrates with TCR signalling (signal 1) to enable full T cell activation. It stabilizes IL\u20112 transcription and mRNA, promotes survival (e.g., Bcl\u2011xL upregulation), and drives clonal expansion"

- term:
    id: GO:0042110
    label: T cell activation
  evidence_type: IBA
  original_reference_id: GO_REF:0000033
  review:
    summary: >-
      T cell activation is the central biological process in which CD28 participates. The deep research
      review identifies CD28 as the prototypic costimulatory receptor providing signal 2 for T cell
      activation. UniProt describes CD28 as a receptor that plays a role in T-cell activation.
    action: ACCEPT
    reason: >-
      T cell activation is a core function of CD28. The IBA annotation is consistent with the primary
      biological role of this receptor. Supported by extensive literature and phylogenetic conservation.
    supported_by:
    - reference_id: PMID:1650475
      supporting_text: "B-cell surface antigen B7 provides a costimulatory signal that induces T cells to proliferate and secrete interleukin 2"
    - reference_id: file:human/CD28/CD28-deep-research-falcon.md
      supporting_text: "CD28 is the prototypic co-stimulatory receptor (signal 2) that integrates with TCR signalling (signal 1) to enable full T cell activation"

- term:
    id: GO:0009897
    label: external side of plasma membrane
  evidence_type: IBA
  original_reference_id: GO_REF:0000033
  review:
    summary: >-
      CD28 is a type I transmembrane protein with its IgV-type ligand-binding domain on the external
      side of the plasma membrane. UniProt annotation confirms extracellular domain spanning residues
      19-152. IDA evidence from PMID:11390434 directly demonstrates cell surface expression.
    action: ACCEPT
    reason: >-
      External side of plasma membrane is the correct localization for CD28's functional domain. The
      extracellular IgV domain binds B7 ligands (CD80/CD86) and this is where CD28 carries out its
      receptor function. Phylogenetically conserved localization.
    supported_by:
    - reference_id: PMID:11390434
      supporting_text: "the following Abs (all IgM) were used: CD4 (MEM-16), CD5 (MEM-128), and CD28 (248.23.2; reference 25)"
    - reference_id: file:human/CD28/CD28-uniprot.txt
      supporting_text: "TOPO_DOM 19..152 /note=Extracellular"

- term:
    id: GO:0031295
    label: T cell costimulation
  evidence_type: IBA
  original_reference_id: GO_REF:0000033
  review:
    summary: >-
      T cell costimulation is the defining biological process for CD28. It provides the critical
      signal 2 that cooperates with TCR engagement to enable full T cell activation and prevent
      anergy. This is the most precise BP term for CD28's role.
    action: ACCEPT
    reason: >-
      T cell costimulation is the single most accurate biological process term for CD28 and represents
      its core function. CD28 is literally the prototypic T cell costimulatory receptor. Strongly
      supported by IBA phylogenetic evidence and extensive literature.
    supported_by:
    - reference_id: PMID:8617933
      supporting_text: "ligation of CD3/TCR and CD28 in the absence of CD40L triggering resulted in little or no IL-4 synthesis"
    - reference_id: file:human/CD28/CD28-deep-research-falcon.md
      supporting_text: "CD28 is the prototypic co-stimulatory receptor (signal 2)"

- term:
    id: GO:0050852
    label: T cell receptor signaling pathway
  evidence_type: IBA
  original_reference_id: GO_REF:0000033
  review:
    summary: >-
      CD28 costimulation amplifies and modulates TCR signaling. The 2024 imaging study demonstrates
      that CD28 costimulation specifically accelerates Lck recruitment and ZAP70 activation within
      TCR microclusters. CD28 delivers unique signals that alter the gene expression program of
      T cells beyond simple TCR amplification (PMID:24665965 per UniProt).
    action: ACCEPT
    reason: >-
      CD28 is intimately coupled to the TCR signaling pathway - it amplifies proximal TCR signals
      by recruiting Lck to TCR microclusters, lowers the activation threshold, and feeds into
      the same downstream NF-kB/NFAT/AP-1 transcriptional programs. The IBA annotation is
      phylogenetically sound and well supported.
    supported_by:
    - reference_id: file:human/CD28/CD28-deep-research-falcon.md
      supporting_text: "CD28 co-stimulation accelerates recruitment of Lck into TCR microclusters, which in turn accelerates ZAP70 recruitment and activation, effectively lowering the TCR activation threshold"

# ========================
# IEA ANNOTATIONS
# ========================
- term:
    id: GO:0002376
    label: immune system process
  evidence_type: IEA
  original_reference_id: GO_REF:0000117
  review:
    summary: >-
      Immune system process is a very broad parent term. CD28 is clearly involved in immune system
      processes as the primary T cell costimulatory receptor. However, this is much less informative
      than the more specific terms already annotated (T cell costimulation, T cell activation, etc.).
    action: ACCEPT
    reason: >-
      While very broad, this IEA annotation is not incorrect. CD28 is fundamentally an immune system
      molecule. More specific child terms are already present. Acceptable as a broad IEA mapping.

- term:
    id: GO:0005886
    label: plasma membrane
  evidence_type: IEA
  original_reference_id: GO_REF:0000044
  review:
    summary: >-
      CD28 is a type I single-pass transmembrane protein that localizes to the plasma membrane.
      UniProt subcellular location annotation confirms cell membrane localization. Multiple IDA
      annotations also support this.
    action: ACCEPT
    reason: >-
      Plasma membrane is the correct localization for CD28. This IEA annotation is consistent
      with multiple higher-evidence annotations (IDA) for the same term. Well supported by
      UniProt subcellular location.

- term:
    id: GO:0006955
    label: immune response
  evidence_type: IEA
  original_reference_id: GO_REF:0000002
  review:
    summary: >-
      Immune response is a broad BP term from InterPro mapping based on the CD28 domain (IPR008093).
      CD28 is essential for adaptive immune responses, particularly T cell-mediated immunity.
    action: ACCEPT
    reason: >-
      While broad, the IEA annotation from InterPro mapping is correct. CD28 is a central player
      in adaptive immune responses. More specific terms (T cell costimulation, T cell activation)
      provide the needed detail.

- term:
    id: GO:0009986
    label: cell surface
  evidence_type: IEA
  original_reference_id: GO_REF:0000044
  review:
    summary: >-
      Cell surface localization is correct for CD28, which is expressed on the T cell surface.
      Also supported by IDA evidence from PMID:15067037.
    action: ACCEPT
    reason: >-
      Correct IEA annotation. CD28 is a cell surface receptor. Consistent with IDA evidence
      from PMID:15067037 and the external side of plasma membrane IBA annotation.

- term:
    id: GO:0016020
    label: membrane
  evidence_type: IEA
  original_reference_id: GO_REF:0000002
  review:
    summary: >-
      Membrane is a very broad CC term from InterPro mapping. CD28 is a transmembrane protein,
      so this is correct but uninformative given more specific terms (plasma membrane, external
      side of plasma membrane).
    action: ACCEPT
    reason: >-
      Correct but very broad. More specific CC annotations are available. Acceptable as a broad
      IEA mapping from InterPro.

- term:
    id: GO:0042129
    label: regulation of T cell proliferation
  evidence_type: IEA
  original_reference_id: GO_REF:0000002
  review:
    summary: >-
      Regulation of T cell proliferation from InterPro mapping (IPR040216 CTLA4/CD28 domain).
      CD28 positively regulates T cell proliferation, so this parent term is correct.
    action: ACCEPT
    reason: >-
      Correct IEA annotation. The more specific child term GO:0042102 (positive regulation of
      T cell proliferation) is also annotated with experimental evidence. This broader IEA
      term is acceptable.

- term:
    id: GO:0050870
    label: positive regulation of T cell activation
  evidence_type: IEA
  original_reference_id: GO_REF:0000117
  review:
    summary: >-
      Positive regulation of T cell activation is appropriate for CD28. CD28 costimulation
      positively regulates T cell activation by lowering the TCR activation threshold,
      promoting cytokine production, and driving proliferation.
    action: ACCEPT
    reason: >-
      Correct IEA annotation. CD28 is the canonical positive regulator of T cell activation.
      Consistent with the IBA annotation for T cell activation and the experimental evidence
      for costimulation.

# ========================
# PROTEIN BINDING IPI ANNOTATIONS
# ========================
- term:
    id: GO:0005515
    label: protein binding
  evidence_type: IPI
  original_reference_id: PMID:10820259
  review:
    summary: >-
      IPI annotation for protein binding based on interaction with PIK3R1 (P27986, the p85
      regulatory subunit of PI3K). The CD28 YMNM motif recruits PI3K via the p85 SH2 domain.
      This is a functionally important interaction for CD28 signaling.
    action: MODIFY
    reason: >-
      Protein binding is uninformative. The interaction with PIK3R1 (p85) is a well-characterized
      signaling interaction via the YMNM motif. This should be captured with a more specific
      MF term reflecting the signaling adaptor recruitment function.
    proposed_replacement_terms:
    - id: GO:0019901
      label: protein kinase binding
    supported_by:
    - reference_id: PMID:7568038
      supporting_text: "p56Lck and p59Fyn regulate CD28 binding to phosphatidylinositol 3-kinase, growth factor receptor-bound protein GRB-2, and T cell-specific protein-tyrosine kinase ITK"

- term:
    id: GO:0005515
    label: protein binding
  evidence_type: IPI
  original_reference_id: PMID:18641334
  review:
    summary: >-
      IPI annotation for protein binding based on interaction with PIK3R1 (P27986). Another
      publication documenting CD28-PI3K p85 interaction. Publication not available for review.
    action: MODIFY
    reason: >-
      Protein binding is uninformative. Same as above - should be protein kinase binding or
      a more specific term reflecting CD28-PI3K signaling interaction.
    proposed_replacement_terms:
    - id: GO:0019901
      label: protein kinase binding

- term:
    id: GO:0005515
    label: protein binding
  evidence_type: IPI
  original_reference_id: PMID:21982860
  review:
    summary: >-
      High-throughput secreted protein microarray screen identifying CD28 interaction with CD80
      (P33681/B7-1). This is a core ligand-receptor interaction. The paper describes a screening
      platform for extracellular protein interaction discovery.
    action: MODIFY
    reason: >-
      Protein binding is uninformative for the CD28-CD80 ligand-receptor interaction. CD28 binding
      to CD80 is the fundamental receptor-ligand interaction that defines CD28 function. The
      coreceptor activity term (GO:0015026) already captures this MF more precisely.
    proposed_replacement_terms:
    - id: GO:0015026
      label: coreceptor activity
    supported_by:
    - reference_id: PMID:21982860
      supporting_text: "functional protein microarrays are particularly well-suited for high-throughput screening of extracellular protein interactions"

- term:
    id: GO:0005515
    label: protein binding
  evidence_type: IPI
  original_reference_id: PMID:32822567
  review:
    summary: >-
      IgSF cell-surface interactome study confirming CD28 interaction with CD80 (P33681). This
      large-scale study systematically mapped direct protein interactions across a recombinant
      IgSF library.
    action: MODIFY
    reason: >-
      Protein binding is too vague for CD28-CD80 receptor-ligand interaction. This is a core
      functional interaction already captured by coreceptor activity.
    proposed_replacement_terms:
    - id: GO:0015026
      label: coreceptor activity
    supported_by:
    - reference_id: PMID:32822567
      supporting_text: "Cell-surface protein-protein interactions (PPIs) mediate cell-cell communication, recognition, and responses"

- term:
    id: GO:0005515
    label: protein binding
  evidence_type: IPI
  original_reference_id: PMID:32822567
  review:
    summary: >-
      Same IgSF interactome study, this entry for CD28 interaction with CD86 (P42081/B7-2).
      CD86 is the other principal ligand for CD28 alongside CD80.
    action: MODIFY
    reason: >-
      Protein binding is uninformative. CD28-CD86 is a core receptor-ligand interaction. Should
      be captured under coreceptor activity.
    proposed_replacement_terms:
    - id: GO:0015026
      label: coreceptor activity

- term:
    id: GO:0005515
    label: protein binding
  evidence_type: IPI
  original_reference_id: PMID:35922511
  review:
    summary: >-
      Physical wiring diagram for the human immune system confirming CD28-CD80 interaction.
      Large-scale systematic mapping of direct protein interactions across human leukocyte
      surface proteins.
    action: MODIFY
    reason: >-
      Protein binding is uninformative. Another confirmation of the CD28-CD80 core ligand
      interaction. Should be coreceptor activity.
    proposed_replacement_terms:
    - id: GO:0015026
      label: coreceptor activity
    supported_by:
    - reference_id: PMID:35922511
      supporting_text: "using a high-throughput surface receptor screening method, we systematically mapped the direct protein interactions across a recombinant library that encompasses most of the surface proteins that are detectable on human leukocytes"

- term:
    id: GO:0005515
    label: protein binding
  evidence_type: IPI
  original_reference_id: PMID:35922511
  review:
    summary: >-
      Same wiring diagram study, this entry for CD28-CD86 (P42081) interaction.
    action: MODIFY
    reason: >-
      Protein binding is uninformative for CD28-CD86 core ligand interaction. Should be
      coreceptor activity.
    proposed_replacement_terms:
    - id: GO:0015026
      label: coreceptor activity

- term:
    id: GO:0005515
    label: protein binding
  evidence_type: IPI
  original_reference_id: PMID:7568038
  review:
    summary: >-
      Raab et al. 1995 demonstrated that Lck and Fyn regulate CD28 binding to PI3K (PIK3R1/p85),
      GRB2, and ITK. Mutation of Tyr-191 greatly reduced phosphorylation by Lck. The with/from
      column indicates PIK3R1 (P27986). This paper is a foundational study of CD28 signaling.
    action: MODIFY
    reason: >-
      Protein binding is uninformative for this well-characterized signaling interaction.
      CD28-PI3K interaction is mediated by the YMNM motif and is critical for costimulatory
      signaling. Should be protein kinase binding.
    proposed_replacement_terms:
    - id: GO:0019901
      label: protein kinase binding
    supported_by:
    - reference_id: PMID:7568038
      supporting_text: "p56Lck and p59Fyn regulate CD28 binding to phosphatidylinositol 3-kinase, growth factor receptor-bound protein GRB-2, and T cell-specific protein-tyrosine kinase ITK"

- term:
    id: GO:0005515
    label: protein binding
  evidence_type: IPI
  original_reference_id: PMID:7584133
  review:
    summary: >-
      Cai et al. 1995 showed that CD28 pYMNM mutations disrupt PI3K and GRB2 binding, directly
      implicating PI3K in CD28-mediated costimulation leading to IL-2 secretion. With/from
      indicates PIK3R1 (P27986).
    action: MODIFY
    reason: >-
      Protein binding is uninformative. This foundational paper demonstrates that the CD28-PI3K
      interaction via the YMNM motif is essential for costimulation. Should be protein kinase
      binding.
    proposed_replacement_terms:
    - id: GO:0019901
      label: protein kinase binding
    supported_by:
    - reference_id: PMID:7584133
      supporting_text: "Both Y191CD28F and M194CD28C mutants failed to generate IL-2. These data directly implicate PI 3-kinase in CD28-mediated costimulation leading to IL-2 secretion"

- term:
    id: GO:0005515
    label: protein binding
  evidence_type: IPI
  original_reference_id: PMID:7807015
  review:
    summary: >-
      Schneider et al. 1995 studied CTLA-4 binding to PI3K, but with/from indicates PIK3R1
      (P27986). The paper demonstrates that CTLA-4 also associates with PI3K, comparing binding
      affinities with CD28. CD28 data was used as a reference point.
    action: MODIFY
    reason: >-
      Protein binding is uninformative. This documents the CD28-PI3K p85 interaction in
      comparison to CTLA-4. Should be protein kinase binding.
    proposed_replacement_terms:
    - id: GO:0019901
      label: protein kinase binding
    supported_by:
    - reference_id: PMID:7807015
      supporting_text: "the NH2- and COOH-terminal SH2 domains of p85 bind the CTLA-4 cytoplasmic pYVKM motif with an affinity (ID50: 0.6 and 0.04 microM), that is similar to CD28"

- term:
    id: GO:0005515
    label: protein binding
  evidence_type: IPI
  original_reference_id: PMID:8146197
  review:
    summary: >-
      Publication not available for review. With/from indicates PIK3R1 (P27986). Likely
      another study documenting the CD28-PI3K p85 interaction.
    action: MODIFY
    reason: >-
      Protein binding is uninformative. Based on the with/from column indicating PIK3R1
      interaction, should be protein kinase binding. Publication not available to confirm
      specific details.
    proposed_replacement_terms:
    - id: GO:0019901
      label: protein kinase binding

- term:
    id: GO:0005515
    label: protein binding
  evidence_type: IPI
  original_reference_id: PMID:8183372
  review:
    summary: >-
      Publication not available for review. With/from indicates PIK3R1 (P27986). Another
      study documenting CD28-PI3K interaction.
    action: MODIFY
    reason: >-
      Protein binding is uninformative. Based on with/from indicating PIK3R1, should be
      protein kinase binding. Publication not available.
    proposed_replacement_terms:
    - id: GO:0019901
      label: protein kinase binding

- term:
    id: GO:0005515
    label: protein binding
  evidence_type: IPI
  original_reference_id: PMID:9417079
  review:
    summary: >-
      Kim et al. 1998 characterized GRB2 SH2/SH3 domain binding to CD28 and its role in
      co-signaling. Both SH2 and SH3 domains contribute to CD28 binding. Mutations that alter
      GRB2 binding blocked CD28-dependent IL-2 production and Vav phosphorylation.
      With/from indicates PIK3R1 (P27986).
    action: MODIFY
    reason: >-
      Protein binding is uninformative. This paper documents functional significance of GRB2
      binding to CD28 via both SH2 and SH3 domains. Should be more specific.
    proposed_replacement_terms:
    - id: GO:0019901
      label: protein kinase binding
    supported_by:
    - reference_id: PMID:9417079
      supporting_text: "Mutations that alter Grb2 binding were found to block the CD28-dependent interleukin-2 production. Further, tyrosine phosphorylation of Vav and the costimulation-dependent activation of Jun N-terminal kinase was blocked"

# ========================
# IEA - ENSEMBL TRANSFER ANNOTATIONS
# ========================
- term:
    id: GO:0001772
    label: immunological synapse
  evidence_type: IEA
  original_reference_id: GO_REF:0000107
  review:
    summary: >-
      Ensembl Compara transfer from mouse CD28 (P31041). CD28 localizes to the immunological
      synapse upon T cell-APC contact, where it co-assembles with TCR microclusters. The 2024
      TIRF microscopy study directly demonstrates CD28 costimulation shapes signaling within
      TCR microclusters at the immune synapse.
    action: ACCEPT
    reason: >-
      Immunological synapse localization is well established for CD28. This is where CD28
      carries out its costimulatory function by recruiting Lck to TCR microclusters. The
      IEA transfer from mouse is consistent with extensive experimental evidence.
    supported_by:
    - reference_id: file:human/CD28/CD28-deep-research-falcon.md
      supporting_text: "CD28 co-localizes with TCR microclusters within the immune synapse"

- term:
    id: GO:0009897
    label: external side of plasma membrane
  evidence_type: IEA
  original_reference_id: GO_REF:0000107
  review:
    summary: >-
      Ensembl Compara transfer from mouse CD28. Consistent with IBA annotation for same term
      and IDA evidence from PMID:11390434.
    action: ACCEPT
    reason: >-
      Correct IEA annotation. Consistent with IBA and IDA evidence for the same term.

- term:
    id: GO:0019901
    label: protein kinase binding
  evidence_type: IEA
  original_reference_id: GO_REF:0000107
  review:
    summary: >-
      Ensembl Compara transfer from mouse. CD28 binds multiple protein kinases including
      PI3K (via p85 regulatory subunit, YMNM motif), Lck (via PYAP motif), ITK (via PRRP motif),
      and PKC-theta. This is a well-documented core molecular function.
    action: ACCEPT
    reason: >-
      Protein kinase binding is well supported for CD28. The cytoplasmic tail recruits PI3K,
      Lck, ITK, and PKC-theta through distinct motifs. This IEA is consistent with extensive
      experimental evidence and is a far more informative MF term than generic protein binding.
    supported_by:
    - reference_id: PMID:7568038
      supporting_text: "p56Lck and p59Fyn regulate CD28 binding to phosphatidylinositol 3-kinase, growth factor receptor-bound protein GRB-2, and T cell-specific protein-tyrosine kinase ITK"
    - reference_id: file:human/CD28/CD28-deep-research-falcon.md
      supporting_text: "YMNM (binds PI3K and GRB2), PRRP (recruits ITK leading to PLCgamma1 and SLP-76/GADS engagement), and PYAP (dominant for LCK binding and assembly of PKCtheta at the synapse)"

# ========================
# IDA ANNOTATIONS
# ========================
- term:
    id: GO:0005886
    label: plasma membrane
  evidence_type: IDA
  original_reference_id: GO_REF:0000052
  review:
    summary: >-
      IDA annotation based on immunofluorescence data (HPA). CD28 is a transmembrane protein
      at the plasma membrane.
    action: ACCEPT
    reason: >-
      Plasma membrane localization is unambiguously correct for CD28. Supported by
      immunofluorescence and consistent with the known type I transmembrane topology.

- term:
    id: GO:0004888
    label: transmembrane signaling receptor activity
  evidence_type: IDA
  original_reference_id: PMID:11279501
  review:
    summary: >-
      Schwartz et al. 2001 solved the crystal structure of the CTLA-4/B7-2 complex, providing
      structural basis for costimulation. The paper discusses both CD28 and CTLA-4 as
      transmembrane signaling receptors that engage B7 ligands. CD28 ligation with B7 ligands
      initiates intracellular signaling cascades.
    action: ACCEPT
    reason: >-
      Transmembrane signaling receptor activity is an appropriate MF for CD28. CD28 spans the
      membrane and upon ligand (CD80/CD86) binding, transmits signals intracellularly through
      its cytoplasmic YMNM/PRRP/PYAP motifs. This term is accurate but somewhat general;
      coreceptor activity (GO:0015026) is more precise.
    supported_by:
    - reference_id: PMID:11279501
      supporting_text: "Regulation of T-cell activity is dependent on antigen-independent co-stimulatory signals provided by the disulphide-linked homodimeric T-cell surface receptors, CD28 and CTLA-4"

- term:
    id: GO:0004888
    label: transmembrane signaling receptor activity
  evidence_type: IDA
  original_reference_id: PMID:12196291
  review:
    summary: >-
      Collins et al. 2002 revisited the interaction properties of costimulatory molecules
      including CD28 with CD80 and CD86. UniProt cites this paper for CD28 function and
      interaction with CD80/CD86. Publication not available but UniProt annotates it for
      CD28 function.
    action: ACCEPT
    reason: >-
      Transmembrane signaling receptor activity is correct for CD28. This paper provided
      key interaction data for CD28-B7 engagement.

- term:
    id: GO:0005886
    label: plasma membrane
  evidence_type: IDA
  original_reference_id: PMID:35397202
  review:
    summary: >-
      Wu et al. 2022 characterized the dimerization interface in the CD28 transmembrane domain
      by NMR. UniProt cites this for subcellular location (cell membrane). The paper confirms
      CD28 as a membrane protein and characterizes TM helix dimerization.
    action: ACCEPT
    reason: >-
      IDA evidence directly confirming plasma membrane localization via structural characterization
      of the transmembrane domain.
    supported_by:
    - reference_id: file:human/CD28/CD28-uniprot.txt
      supporting_text: "SUBCELLULAR LOCATION: Cell membrane {ECO:0000269|PubMed:35397202}; Single-pass type I membrane protein"

- term:
    id: GO:0050862
    label: positive regulation of T cell receptor signaling pathway
  evidence_type: IDA
  original_reference_id: PMID:11279501
  review:
    summary: >-
      Schwartz et al. 2001 structural study of CTLA-4/B7-2 complex. The paper demonstrates
      that CD28 engagement with B7 ligands provides a stimulatory signal for T cell activation,
      consistent with positive regulation of TCR signaling.
    action: ACCEPT
    reason: >-
      Positive regulation of TCR signaling pathway is a core function of CD28. CD28 costimulation
      amplifies TCR signals by recruiting Lck to TCR microclusters and lowering activation
      thresholds. Well supported by this and many other studies.
    supported_by:
    - reference_id: PMID:11279501
      supporting_text: "Engagement of CD28 with B7-1 and B7-2 ligands on antigen-presenting cells (APCs) provides a stimulatory signal for T-cell activation"

- term:
    id: GO:0050862
    label: positive regulation of T cell receptor signaling pathway
  evidence_type: IDA
  original_reference_id: PMID:12196291
  review:
    summary: >-
      Collins et al. 2002 characterized interaction properties of costimulatory molecules.
      UniProt cites this for CD28's function in increasing proliferation and cytokine expression.
    action: ACCEPT
    reason: >-
      Same core function. Multiple IDA annotations for positive regulation of TCR signaling
      from different publications strengthens the evidence. Consistent with CD28's role as
      the prototypic TCR costimulatory receptor.

- term:
    id: GO:0050862
    label: positive regulation of T cell receptor signaling pathway
  evidence_type: IDA
  original_reference_id: PMID:1650475
  review:
    summary: >-
      Gimmi et al. 1991, a foundational paper showing B7 provides a costimulatory signal that
      induces T cells to proliferate and secrete IL-2. UniProt cites this for CD28 function.
      Publication not available but this is one of the original demonstrations of CD28/B7
      costimulation.
    action: ACCEPT
    reason: >-
      This is one of the earliest and most important studies demonstrating that B7-CD28
      costimulation positively regulates T cell responses. Core annotation well supported
      by this seminal publication.

- term:
    id: GO:0004888
    label: transmembrane signaling receptor activity
  evidence_type: TAS
  original_reference_id: PMID:9784967
  review:
    summary: >-
      TAS annotation based on a review. CD28 is clearly a transmembrane signaling receptor.
      Publication not available for review.
    action: ACCEPT
    reason: >-
      Transmembrane signaling receptor activity is correct. CD28 is a type I transmembrane
      receptor that signals upon ligand engagement. TAS annotation is appropriate for a
      review-based citation.

- term:
    id: GO:0042110
    label: T cell activation
  evidence_type: IDA
  original_reference_id: PMID:23918985
  review:
    summary: >-
      IDA annotation for T cell activation. Publication not available for review. UniProt
      cites CD28's role in T cell activation from multiple publications.
    action: ACCEPT
    reason: >-
      T cell activation is a core function of CD28. While the specific publication is not
      available, this annotation is consistent with the overwhelming body of evidence for
      CD28 in T cell activation.

- term:
    id: GO:0042110
    label: T cell activation
  evidence_type: IGI
  original_reference_id: PMID:18337562
  review:
    summary: >-
      IGI annotation for T cell activation with genetic interactor IL-4 (P05112). The referenced
      publication PMID:18337562 (Murooka et al. 2008) focuses on CCL5-mediated T cell chemotaxis
      and mTOR signaling, not directly on CD28. However, T cell activation is the defining
      biological process for CD28, supported by overwhelming evidence from multiple other
      publications and the IBA and IDA annotations for the same term. CD28 is the prototypic
      T cell costimulatory receptor; its role in T cell activation is beyond question
      (PMID:1650475, PMID:23918985, PMID:11279501).
    action: ACCEPT
    reason: >-
      Although the specific publication (PMID:18337562) is about CCL5-mediated chemotaxis rather
      than CD28 per se, T cell activation is the single most well-established function of CD28.
      The IBA and IDA annotations for this same term are already accepted. CD28 is literally
      defined as the prototypic T cell costimulatory receptor, and T cell activation is its core
      function. The IGI evidence code with IL-4 as genetic interactor is consistent with the
      known role of CD28 costimulation in cytokine-dependent T cell activation programs.
      Resolving to ACCEPT for consistency with the other GO:0042110 annotations.
    supported_by:
    - reference_id: PMID:1650475
      supporting_text: "B7-transfected CHO cells can induce suboptimally activated CD28+ T cells to proliferate and secrete high levels of interleukin 2"
    - reference_id: PMID:11279501
      supporting_text: "Engagement of CD28 with B7-1 and B7-2 ligands on antigen-presenting cells (APCs) provides a stimulatory signal for T-cell activation"
    - reference_id: file:human/CD28/CD28-deep-research-falcon.md
      supporting_text: "CD28 is the prototypic co-stimulatory receptor (signal 2) that integrates with TCR signalling (signal 1) to enable full T cell activation"

- term:
    id: GO:0010628
    label: positive regulation of gene expression
  evidence_type: IMP
  original_reference_id: PMID:23817958
  review:
    summary: >-
      IMP annotation for positive regulation of gene expression. Publication not available.
      CD28 signaling is known to activate NF-kB, NFAT, and AP-1 transcription factors,
      which positively regulate expression of many target genes including IL-2.
    action: KEEP_AS_NON_CORE
    reason: >-
      While CD28 signaling does regulate gene expression programs through NF-kB/NFAT/AP-1
      activation, this is a very broad downstream effect rather than a core molecular function
      of CD28 itself. The annotation is not wrong, but represents a downstream consequence of
      CD28 costimulatory signaling rather than a defining function. Publication not available
      to assess specific experimental context.

- term:
    id: GO:0010629
    label: negative regulation of gene expression
  evidence_type: IMP
  original_reference_id: PMID:23817958
  review:
    summary: >-
      IMP annotation for negative regulation of gene expression from the same publication as
      the positive regulation annotation above. CD28 signaling can indeed both upregulate and
      downregulate different gene sets.
    action: KEEP_AS_NON_CORE
    reason: >-
      Same reasoning as for positive regulation of gene expression. This is a downstream
      consequence of CD28 signaling, not a core function. The annotation may be valid for
      specific gene sets that are downregulated by CD28 costimulation, but it is a very broad
      and non-specific annotation. Publication not available.

- term:
    id: GO:0043066
    label: negative regulation of apoptotic process
  evidence_type: TAS
  original_reference_id: PMID:8717514
  review:
    summary: >-
      TAS annotation for negative regulation of apoptosis. CD28 signaling promotes T cell
      survival through upregulation of anti-apoptotic proteins including Bcl-xL. This is
      a well-known downstream effect of CD28 costimulation. Publication not available.
    action: KEEP_AS_NON_CORE
    reason: >-
      CD28 signaling does promote cell survival and inhibit apoptosis (e.g., via Bcl-xL
      upregulation), but this is a downstream consequence of its costimulatory signaling
      rather than its primary molecular function. It is a pleiotropic effect of CD28
      activation, not its defining function.
    supported_by:
    - reference_id: file:human/CD28/CD28-deep-research-falcon.md
      supporting_text: "promotes survival (e.g., Bcl-xL upregulation)"

- term:
    id: GO:0098636
    label: protein complex involved in cell adhesion
  evidence_type: IDA
  original_reference_id: PMID:7544393
  review:
    summary: >-
      Fargeas et al. 1995 identified residues in the CD80 V domain implicated in functional
      interactions with CD28 and CTLA-4. The paper studied CD80-CD28 adhesion and IL-2
      production using CD80 mutants. The annotation refers to CD28 as part of a protein
      complex involved in cell adhesion (T cell-APC interaction).
    action: KEEP_AS_NON_CORE
    reason: >-
      While CD28-CD80 interaction does contribute to T cell-APC adhesion at the immune synapse,
      this is not the primary function of CD28. CD28 is fundamentally a signaling coreceptor,
      not primarily an adhesion molecule. The cell adhesion aspect is secondary to its
      costimulatory signaling role.
    supported_by:
    - reference_id: PMID:7544393
      supporting_text: "human CD80 mutants were generated and tested for their ability to maintain the interaction with CD28 leading to adhesion and enhanced IL-2 production"

- term:
    id: GO:0031295
    label: T cell costimulation
  evidence_type: TAS
  original_reference_id: PMID:8617933
  review:
    summary: >-
      Blotta et al. 1996 showed that CD40L costimulation enhances IL-4 synthesis in conjunction
      with CD3/TCR and CD28 ligation. The paper directly demonstrates CD28's role in T cell
      costimulation.
    action: ACCEPT
    reason: >-
      T cell costimulation is the core function of CD28. This TAS annotation from a study
      directly examining CD28 costimulation in conjunction with CD3 and CD40L is well supported.
    supported_by:
    - reference_id: PMID:8617933
      supporting_text: "IL-4 synthesis was greatly enhanced by triggering of CD40L on the T cell surface in conjunction with ligation of CD3/TCR and CD28"

- term:
    id: GO:0032733
    label: positive regulation of interleukin-10 production
  evidence_type: IDA
  original_reference_id: PMID:8617933
  review:
    summary: >-
      Blotta et al. 1996 showed that CD3/CD28/CD40L costimulation enhances cytokine production
      including IL-10 in T cells.
    action: KEEP_AS_NON_CORE
    reason: >-
      IL-10 production is a downstream consequence of CD28 costimulation combined with CD40L
      signaling. While experimentally demonstrated (IDA), this is a specific cytokine output
      rather than a core function of CD28. The core function is T cell costimulation broadly.
    supported_by:
    - reference_id: PMID:8617933
      supporting_text: "IL-4 synthesis was greatly enhanced by triggering of CD40L on the T cell surface in conjunction with ligation of CD3/TCR and CD28"

- term:
    id: GO:0032753
    label: positive regulation of interleukin-4 production
  evidence_type: IDA
  original_reference_id: PMID:8617933
  review:
    summary: >-
      Blotta et al. 1996 directly demonstrated that CD28 costimulation (with CD3 and CD40L)
      greatly enhances IL-4 synthesis. This was the main finding of the paper.
    action: KEEP_AS_NON_CORE
    reason: >-
      IL-4 production enhancement is a specific downstream effect of CD28 costimulation,
      particularly in combination with CD40L signaling. While experimentally validated, this
      represents a specific cytokine output rather than the core costimulatory function.
    supported_by:
    - reference_id: PMID:8617933
      supporting_text: "IL-4 synthesis was greatly enhanced by triggering of CD40L on the T cell surface in conjunction with ligation of CD3/TCR and CD28, whereas ligation of CD3/TCR and CD28 in the absence of CD40L triggering resulted in little or no IL-4 synthesis"

- term:
    id: GO:0042102
    label: positive regulation of T cell proliferation
  evidence_type: IDA
  original_reference_id: PMID:8617933
  review:
    summary: >-
      Blotta et al. 1996 demonstrated T cell proliferation in response to CD3/CD28 costimulation.
      The paper used anti-CD28 antibodies as costimulatory signals.
    action: ACCEPT
    reason: >-
      Positive regulation of T cell proliferation is a core function of CD28. IDA evidence
      from Blotta et al. directly demonstrates this. Consistent with multiple other annotations.
    supported_by:
    - reference_id: PMID:8617933
      supporting_text: "using immobilized mAbs against several T cell surface molecules, including CD3, CD28, and the CD40 ligand (CD40L)"

- term:
    id: GO:0005515
    label: protein binding
  evidence_type: IPI
  original_reference_id: PMID:15067037
  review:
    summary: >-
      Mikolajczak et al. 2004 showed that isoform 3 (CD28i) interacts with CD40LG (P29965/CD154).
      This is an interaction specific to the CD28i splice variant that lacks most of the
      extracellular domain.
    action: UNDECIDED
    reason: >-
      This interaction is specific to isoform 3 (CD28i), which has a truncated extracellular
      domain and interacts with CD40LG rather than B7 ligands. While the interaction is
      experimentally demonstrated, it is isoform-specific. Protein binding is uninformative
      but the isoform-specific nature makes it harder to propose a simple replacement.
    supported_by:
    - reference_id: file:human/CD28/CD28-uniprot.txt
      supporting_text: "Isoform 3: Enhances CD40L-mediated activation of NF-kappa-B and kinases MAPK8 and PAK2 in T-cells"

- term:
    id: GO:0009986
    label: cell surface
  evidence_type: IDA
  original_reference_id: PMID:15067037
  review:
    summary: >-
      Mikolajczak et al. 2004 demonstrated cell surface localization of isoform 3 (CD28i).
      UniProt confirms subcellular location for isoform 3 as cell surface.
    action: ACCEPT
    reason: >-
      Cell surface localization is correct for CD28, including isoform 3. IDA evidence
      directly demonstrates this.
    supported_by:
    - reference_id: file:human/CD28/CD28-uniprot.txt
      supporting_text: "SUBCELLULAR LOCATION: [Isoform 3]: Cell surface"

# ========================
# REACTOME TAS ANNOTATIONS - Plasma membrane
# ========================
- term:
    id: GO:0005886
    label: plasma membrane
  evidence_type: TAS
  original_reference_id: Reactome:R-HSA-2316434
  review:
    summary: >-
      Reactome annotation for PI3K phosphorylation of PIP2 to PIP3 in context of CD28
      signaling at the plasma membrane.
    action: ACCEPT
    reason: >-
      Plasma membrane localization in the context of PI3K signaling downstream of CD28 is
      correct. CD28 recruits PI3K to the plasma membrane.

- term:
    id: GO:0005886
    label: plasma membrane
  evidence_type: TAS
  original_reference_id: Reactome:R-HSA-2400009
  review:
    summary: >-
      Reactome annotation for PI3K inhibitors blocking PI3K catalytic activity. CD28 at
      the plasma membrane.
    action: ACCEPT
    reason: Correct. CD28 at the plasma membrane in PI3K signaling context.

- term:
    id: GO:0005886
    label: plasma membrane
  evidence_type: TAS
  original_reference_id: Reactome:R-HSA-388814
  review:
    summary: Reactome annotation for Grb2 binding to CD28 at the plasma membrane.
    action: ACCEPT
    reason: Correct. CD28 recruits Grb2 at the plasma membrane via the YMNM motif.

- term:
    id: GO:0005886
    label: plasma membrane
  evidence_type: TAS
  original_reference_id: Reactome:R-HSA-388831
  review:
    summary: Reactome annotation for phosphorylation of CD28 at the plasma membrane.
    action: ACCEPT
    reason: Correct. CD28 is phosphorylated by Lck at the plasma membrane.

- term:
    id: GO:0005886
    label: plasma membrane
  evidence_type: TAS
  original_reference_id: Reactome:R-HSA-388832
  review:
    summary: Reactome annotation for PI3K binding to CD28 at the plasma membrane.
    action: ACCEPT
    reason: Correct. PI3K binds CD28 at the plasma membrane via the pYMNM motif.

- term:
    id: GO:0005886
    label: plasma membrane
  evidence_type: TAS
  original_reference_id: Reactome:R-HSA-389158
  review:
    summary: Reactome annotation for CD28-bound PI3K phosphorylating PIP2 to PIP3.
    action: ACCEPT
    reason: Correct. PI3K recruited to CD28 at the plasma membrane generates PIP3.

- term:
    id: GO:0005886
    label: plasma membrane
  evidence_type: TAS
  original_reference_id: Reactome:R-HSA-389348
  review:
    summary: Reactome annotation for activation of Rac1 by pVav1 in the CD28 pathway at the plasma membrane.
    action: ACCEPT
    reason: Correct. Vav1 activation downstream of CD28 at the plasma membrane leads to Rac1 activation.

- term:
    id: GO:0005886
    label: plasma membrane
  evidence_type: TAS
  original_reference_id: Reactome:R-HSA-389350
  review:
    summary: Reactome annotation for activation of Cdc42 by pVav1 in the CD28 pathway at the plasma membrane.
    action: ACCEPT
    reason: Correct. Vav1 activation downstream of CD28 at the plasma membrane leads to Cdc42 activation.

- term:
    id: GO:0005886
    label: plasma membrane
  evidence_type: TAS
  original_reference_id: Reactome:R-HSA-389352
  review:
    summary: Reactome annotation for translocation of Vav1 to CD28 at the plasma membrane.
    action: ACCEPT
    reason: Correct. Vav1 is recruited to CD28 at the plasma membrane.

- term:
    id: GO:0005886
    label: plasma membrane
  evidence_type: TAS
  original_reference_id: Reactome:R-HSA-389354
  review:
    summary: Reactome annotation for activation of Vav1 in the CD28 pathway at the plasma membrane.
    action: ACCEPT
    reason: Correct. Vav1 is activated in the CD28 signaling pathway at the plasma membrane.

- term:
    id: GO:0005886
    label: plasma membrane
  evidence_type: TAS
  original_reference_id: Reactome:R-HSA-389381
  review:
    summary: Reactome annotation for Gads binding to CD28 at the plasma membrane.
    action: ACCEPT
    reason: Correct. Gads is recruited to CD28 at the plasma membrane.

- term:
    id: GO:0005829
    label: cytosol
  evidence_type: TAS
  original_reference_id: Reactome:R-HSA-167637
  review:
    summary: >-
      Reactome annotation for internalization of Nef:CD28:Clathrin-Coated Pit Adapter Protein
      complex. This is in the context of HIV Nef-mediated downregulation of CD28, where CD28
      is internalized to the cytosol.
    action: KEEP_AS_NON_CORE
    reason: >-
      Cytosol localization of CD28 is specific to the HIV Nef-mediated downregulation pathway
      where CD28 is internalized from the cell surface. This is not a normal physiological
      localization of CD28 but rather a pathological consequence of viral hijacking. Not a
      core localization.

- term:
    id: GO:0005886
    label: plasma membrane
  evidence_type: IDA
  original_reference_id: PMID:11390434
  review:
    summary: >-
      Kirchgessner et al. 2001 studied the transmembrane adaptor protein TRIM and its
      association with TCR. CD28 plasma membrane expression was confirmed as part of
      the characterization of T cell surface molecules.
    action: ACCEPT
    reason: >-
      IDA evidence for plasma membrane localization. Correct and consistent with all
      other evidence.

- term:
    id: GO:0009897
    label: external side of plasma membrane
  evidence_type: IDA
  original_reference_id: PMID:11390434
  review:
    summary: >-
      Same study as above. IDA evidence for external side of plasma membrane localization,
      consistent with IBA annotation.
    action: ACCEPT
    reason: >-
      IDA evidence confirming CD28's extracellular domain is on the external side of the
      plasma membrane. Consistent with IBA and IEA annotations for the same term.

- term:
    id: GO:0045066
    label: regulatory T cell differentiation
  evidence_type: IDA
  original_reference_id: PMID:18641304
  review:
    summary: >-
      Zheng et al. 2008 studied acquisition of suppressive function by activated human
      CD4+CD25- T cells. The paper focuses on CTLA-4 and FoxP3 in Treg induction, with
      CD28 involved in the activation that generates these cells. CD28 signaling is required
      for the initial T cell activation that leads to CTLA-4 upregulation and Treg differentiation.
    action: KEEP_AS_NON_CORE
    reason: >-
      While CD28 signaling contributes to T cell activation that can lead to Treg differentiation,
      this paper primarily focuses on CTLA-4 and FoxP3 as the key determinants of regulatory T cell
      function. CD28's role is in providing the initial activation signal, not specifically in
      directing the Treg differentiation program. This is a downstream consequence rather than a
      core function.
    supported_by:
    - reference_id: PMID:18641304
      supporting_text: "Activation of human CD4(+)CD25(-) T cells resulted in the appearance of a de novo population of FoxP3-expressing cells within 48 h. These cells expressed high levels of CTLA-4"

# ========================
# MORE REACTOME TAS - Nef pathway
# ========================
- term:
    id: GO:0005886
    label: plasma membrane
  evidence_type: TAS
  original_reference_id: Reactome:R-HSA-167630
  review:
    summary: >-
      Reactome annotation for Formation of Nef-CD28 cytoplasmic tail complex at the plasma
      membrane (HIV Nef pathway).
    action: ACCEPT
    reason: >-
      CD28 is at the plasma membrane when Nef interacts with its cytoplasmic tail. Correct
      localization even in this pathological context.

- term:
    id: GO:0005886
    label: plasma membrane
  evidence_type: TAS
  original_reference_id: Reactome:R-HSA-167633
  review:
    summary: >-
      Reactome annotation for Formation of Nef:CD28:Clathrin-Coated Pit Adapter Protein
      complex at the plasma membrane.
    action: ACCEPT
    reason: Correct. CD28 at the plasma membrane in HIV Nef pathway.

- term:
    id: GO:0005886
    label: plasma membrane
  evidence_type: TAS
  original_reference_id: Reactome:R-HSA-167637
  review:
    summary: >-
      Reactome annotation for internalization of the Nef:CD28:Clathrin complex from the
      plasma membrane.
    action: ACCEPT
    reason: Correct. CD28 starts at the plasma membrane before Nef-mediated internalization.

- term:
    id: GO:0005886
    label: plasma membrane
  evidence_type: TAS
  original_reference_id: Reactome:R-HSA-388808
  review:
    summary: Reactome annotation for CD28 homodimer binding B7-2 monomer at the plasma membrane.
    action: ACCEPT
    reason: Correct. CD28-CD86 interaction occurs at the plasma membrane.

- term:
    id: GO:0005886
    label: plasma membrane
  evidence_type: TAS
  original_reference_id: Reactome:R-HSA-388811
  review:
    summary: Reactome annotation for CD28 homodimer binding B7-1 homodimer at the plasma membrane.
    action: ACCEPT
    reason: Correct. CD28-CD80 interaction occurs at the plasma membrane.

# ========================
# ADDITIONAL IPI PROTEIN BINDING
# ========================
- term:
    id: GO:0005515
    label: protein binding
  evidence_type: IPI
  original_reference_id: PMID:15554700
  review:
    summary: >-
      Bauer et al. 2004 characterized Lck-binding elements in the herpesviral Tip protein.
      The with/from column indicates Lck (P06239). This paper is about viral Tip protein
      interaction with Lck, not directly about CD28. CD28 may have been used as a reference
      or control for Lck binding.
    action: UNDECIDED
    reason: >-
      The paper is about herpesviral Tip protein interactions with Lck, not primarily about
      CD28. While CD28 does bind Lck via the PYAP motif, the annotation appears to be derived
      from a study focused on a viral protein. Cannot fully assess the relevance of this
      annotation to CD28 without seeing the specific CD28 data in the paper.
    supported_by:
    - reference_id: PMID:15554700
      supporting_text: "Herpesvirus saimiri encodes a tyrosine kinase interacting protein (Tip) that binds to T-cell-specific tyrosine kinase Lck"

- term:
    id: GO:0005515
    label: protein binding
  evidence_type: IPI
  original_reference_id: PMID:11285224
  review:
    summary: >-
      Publication not available for review. With/from indicates P03406 (a viral protein).
      This may relate to viral protein interactions with CD28.
    action: UNDECIDED
    reason: >-
      Publication not available. The with/from column indicates a viral protein (P03406).
      Cannot assess the annotation without the publication.

- term:
    id: GO:0032743
    label: positive regulation of interleukin-2 production
  evidence_type: IDA
  original_reference_id: PMID:3875683
  review:
    summary: >-
      Early publication (1985) demonstrating CD28 involvement in IL-2 production. CD28
      costimulation is one of the best-characterized drivers of IL-2 production in T cells.
      IL-2 is the canonical cytokine whose production is enhanced by CD28 signaling.
      Publication not available.
    action: ACCEPT
    reason: >-
      Positive regulation of IL-2 production is a core downstream effect of CD28 costimulation.
      IL-2 is the signature cytokine induced by CD28 costimulation - this is the classical
      readout of CD28 function. The deep research review confirms CD28 promotes IL-2 production.
    supported_by:
    - reference_id: file:human/CD28/CD28-deep-research-falcon.md
      supporting_text: "increases proliferation and expression of various cytokines in particular IL2 production in both CD4(+) and CD8(+) T-cell subsets"
    - reference_id: PMID:1650475
      supporting_text: "B-cell surface antigen B7 provides a costimulatory signal that induces T cells to proliferate and secrete interleukin 2"

- term:
    id: GO:0042102
    label: positive regulation of T cell proliferation
  evidence_type: IDA
  original_reference_id: PMID:3159820
  review:
    summary: >-
      Early publication (1988) demonstrating CD28 involvement in T cell proliferation.
      One of the foundational studies establishing CD28 as a costimulatory molecule.
      Publication not available.
    action: ACCEPT
    reason: >-
      Positive regulation of T cell proliferation is a core function of CD28. This is
      one of the earliest demonstrations. Consistent with extensive subsequent literature.

- term:
    id: GO:0042802
    label: identical protein binding
  evidence_type: NAS
  original_reference_id: PMID:3159820
  review:
    summary: >-
      NAS annotation for identical protein binding, reflecting CD28 homodimerization. CD28
      exists as a disulfide-linked homodimer on the cell surface. The 2022 NMR structure
      (PMID:35397202) characterized the dimerization interface in the transmembrane domain.
    action: ACCEPT
    reason: >-
      CD28 homodimerization is well established. It exists as a disulfide-linked homodimer,
      and the transmembrane domain contains dimerization motifs (GxxxA). While identical
      protein binding is somewhat generic, it accurately describes CD28 homodimerization.
    supported_by:
    - reference_id: file:human/CD28/CD28-uniprot.txt
      supporting_text: "Homodimer; disulfide-linked"
    - reference_id: file:human/CD28/CD28-deep-research-falcon.md
      supporting_text: "CD28 is a ~44 kDa disulfide-linked Ig-domain homodimer"

- term:
    id: GO:0045840
    label: positive regulation of mitotic nuclear division
  evidence_type: IDA
  original_reference_id: PMID:3159820
  review:
    summary: >-
      IDA annotation for positive regulation of mitotic nuclear division from the same
      1988 publication. CD28 costimulation drives T cell proliferation which involves
      mitotic division.
    action: MARK_AS_OVER_ANNOTATED
    reason: >-
      While CD28 costimulation does drive T cell proliferation (which involves mitosis),
      annotating CD28 specifically to positive regulation of mitotic nuclear division is
      an over-annotation. CD28 is a signaling receptor, not a direct regulator of the
      mitotic machinery. The proliferative effect is a downstream consequence of
      transcriptional and metabolic reprogramming, not a direct regulation of mitosis.
      The more appropriate annotation is positive regulation of T cell proliferation.

- term:
    id: GO:0005515
    label: protein binding
  evidence_type: IPI
  original_reference_id: PMID:9784967
  review:
    summary: >-
      IPI for protein binding with CD80 (P33681). Publication not available. Likely a
      review or primary study documenting CD28-CD80 interaction.
    action: MODIFY
    reason: >-
      Protein binding is uninformative for the CD28-CD80 core receptor-ligand interaction.
      Should be captured under coreceptor activity.
    proposed_replacement_terms:
    - id: GO:0015026
      label: coreceptor activity

- term:
    id: GO:0005515
    label: protein binding
  evidence_type: IPI
  original_reference_id: PMID:9784967
  review:
    summary: >-
      IPI for protein binding with CD86 (P42081). Same publication as above.
    action: MODIFY
    reason: >-
      Protein binding is uninformative for CD28-CD86 core receptor-ligand interaction.
      Should be captured under coreceptor activity.
    proposed_replacement_terms:
    - id: GO:0015026
      label: coreceptor activity

# ========================
# TAS BP ANNOTATIONS
# ========================
- term:
    id: GO:0001819
    label: positive regulation of cytokine production
  evidence_type: TAS
  original_reference_id: PMID:8717514
  review:
    summary: >-
      TAS annotation for positive regulation of cytokine production. CD28 costimulation
      promotes production of multiple cytokines including IL-2, IL-4, IL-10, and others.
      Publication not available.
    action: ACCEPT
    reason: >-
      Positive regulation of cytokine production is a well-established consequence of CD28
      costimulation. While this is a somewhat broad term, CD28 does promote production of
      multiple cytokines (IL-2, IL-4, IL-10, IFN-gamma) making this parent term appropriate.
    supported_by:
    - reference_id: file:human/CD28/CD28-deep-research-falcon.md
      supporting_text: "increases proliferation and expression of various cytokines in particular IL2 production"
    - reference_id: PMID:8617933
      supporting_text: "IL-4 synthesis was greatly enhanced by triggering of CD40L on the T cell surface in conjunction with ligation of CD3/TCR and CD28"

- term:
    id: GO:0005886
    label: plasma membrane
  evidence_type: TAS
  original_reference_id: PMID:9784967
  review:
    summary: >-
      TAS annotation for plasma membrane from a review. Correct and consistent with all
      other evidence.
    action: ACCEPT
    reason: Correct. CD28 is a plasma membrane protein.

- term:
    id: GO:0006959
    label: humoral immune response
  evidence_type: TAS
  original_reference_id: PMID:8717514
  review:
    summary: >-
      TAS annotation for humoral immune response. CD28 costimulation on T cells promotes
      T helper function that supports B cell antibody responses. Publication not available.
    action: KEEP_AS_NON_CORE
    reason: >-
      While CD28 costimulation of T helper cells supports humoral immune responses (by
      promoting T cell help for B cells), this is a distal, pleiotropic consequence of
      CD28 function rather than a core molecular function. CD28 does not directly
      participate in antibody production or B cell processes.

- term:
    id: GO:0007166
    label: cell surface receptor signaling pathway
  evidence_type: TAS
  original_reference_id: PMID:9784967
  review:
    summary: >-
      TAS annotation for cell surface receptor signaling pathway. CD28 is a cell surface
      receptor that signals upon ligand engagement. This is a broad parent term.
    action: ACCEPT
    reason: >-
      Correct but broad. CD28 is a cell surface receptor and its engagement initiates
      signaling cascades. More specific terms (T cell costimulation, positive regulation
      of TCR signaling) provide better detail.

- term:
    id: GO:0015026
    label: coreceptor activity
  evidence_type: TAS
  original_reference_id: PMID:9784967
  review:
    summary: >-
      TAS annotation for coreceptor activity. CD28 is the prototypic T cell coreceptor
      that cooperates with the TCR to enable full T cell activation. The GO definition
      of coreceptor activity fits CD28 precisely: combining with a messenger and in
      cooperation with a nearby primary receptor, initiating a change in cell activity.
    action: ACCEPT
    reason: >-
      Coreceptor activity is the most precise molecular function term for CD28. CD28 acts
      in cooperation with the TCR (the primary receptor) by binding B7 ligands (the
      extracellular messengers) to initiate costimulatory signaling. This is the single
      best MF term for CD28.
    supported_by:
    - reference_id: file:human/CD28/CD28-deep-research-falcon.md
      supporting_text: "CD28 is the prototypic co-stimulatory receptor (signal 2) that integrates with TCR signalling (signal 1) to enable full T cell activation"

- term:
    id: GO:0042102
    label: positive regulation of T cell proliferation
  evidence_type: TAS
  original_reference_id: PMID:9784967
  review:
    summary: >-
      TAS annotation for positive regulation of T cell proliferation from a review.
      Consistent with IDA and IBA annotations for the same term.
    action: ACCEPT
    reason: >-
      Correct TAS annotation consistent with experimental evidence (IDA, IBA) for
      the same core function.

- term:
    id: GO:0045070
    label: positive regulation of viral genome replication
  evidence_type: NAS
  original_reference_id: PMID:7492435
  review:
    summary: >-
      NAS annotation for positive regulation of viral genome replication. This likely
      relates to CD28 costimulation enhancing HIV replication in T cells, as activated
      T cells provide a more permissive environment for viral replication. Publication
      not available.
    action: KEEP_AS_NON_CORE
    reason: >-
      CD28 does not directly regulate viral genome replication. Rather, CD28-mediated
      T cell activation creates an environment permissive for viral replication (e.g.,
      HIV in activated T cells). This is a highly indirect downstream consequence of
      T cell activation, not a core function of CD28.

- term:
    id: GO:0045727
    label: positive regulation of translation
  evidence_type: NAS
  original_reference_id: PMID:12028592
  review:
    summary: >-
      NAS annotation for positive regulation of translation. CD28 signaling is known
      to activate mTOR-dependent translation programs. Publication not available.
    action: KEEP_AS_NON_CORE
    reason: >-
      While CD28 signaling does promote mTOR-dependent translation (particularly of
      IL-2 and other cytokine mRNAs), this is a downstream signaling consequence rather
      than a core function. CD28 is a receptor, not a translation regulator.
    supported_by:
    - reference_id: file:human/CD28/CD28-deep-research-falcon.md
      supporting_text: "CD28 signalling promotes glycolytic flux and mTOR-dependent translation"

references:
- id: GO_REF:0000002
  title: Gene Ontology annotation through association of InterPro records with GO terms
  findings: []
- id: GO_REF:0000033
  title: Annotation inferences using phylogenetic trees
  findings: []
- id: GO_REF:0000044
  title: Gene Ontology annotation based on UniProtKB/Swiss-Prot Subcellular Location vocabulary mapping
  findings: []
- id: GO_REF:0000052
  title: Gene Ontology annotation based on curation of immunofluorescence data
  findings: []
- id: GO_REF:0000107
  title: Automatic transfer of experimentally verified manual GO annotation data to orthologs using Ensembl Compara
  findings: []
- id: GO_REF:0000117
  title: Electronic Gene Ontology annotations created by ARBA machine learning models
  findings: []
- id: PMID:1650475
  title: B-cell surface antigen B7 provides a costimulatory signal that induces T cells to proliferate and secrete interleukin 2.
  findings:
  - statement: Foundational demonstration of B7-CD28 costimulation driving T cell proliferation and IL-2 secretion
    supporting_text: "B7-transfected CHO cells can induce suboptimally activated CD28+ T cells to proliferate and secrete high levels of interleukin 2"
- id: PMID:3159820
  title: "Human T cell activation. II. A new activation pathway used by a major T cell population via a disulfide-bonded dimer of a 44 kilodalton polypeptide (9.3 antigen)."
  findings:
  - statement: One of the earliest demonstrations of anti-CD28 antibody-mediated T cell costimulation
    supporting_text: "mAb 9.3 was capable of inducing T cell proliferation in the presence of 12-o-tetradecanoyl phorbol-13-acetate (TPA)"
- id: PMID:3875683
  title: Involvement of T44 molecules in an antigen-independent pathway of T cell activation. Analysis of the correlations to the T cell antigen-receptor complex.
  findings:
  - statement: Demonstrated CD28 costimulation promotes IL-2 production
    supporting_text: "triggering of T lymphocytes with 9.3 antibody (in the presence of adherent cells) resulted in strong IL-2 production that peaked at 48 h"
- id: PMID:7544393
  title: Identification of residues in the V domain of CD80 (B7-1) implicated in functional interactions with CD28 and CTLA4.
  findings:
  - statement: Identified hydrophobic residues in CD80 critical for CD28 and CTLA-4 binding
    supporting_text: "Two hydrophobic residues in the V-like domain of CD80 were identified as critical for binding to CD28 and are also important for the interaction with CTLA4"
  - statement: Y87 in CD80 is conserved across species and important for CD28 interaction
    supporting_text: "One of these residues, Y87, is conserved in all CD80 and CD86 cloned from various species"
- id: PMID:7568038
  title: "p56Lck and p59Fyn regulate CD28 binding to phosphatidylinositol 3-kinase, growth factor receptor-bound protein GRB-2, and T cell-specific protein-tyrosine kinase ITK: implications for T-cell costimulation."
  findings:
  - statement: Demonstrated Lck-dependent phosphorylation of CD28 Tyr-191
    supporting_text: "p56Lck and p59Fyn phosphorylate CD28 primarily at Tyr-191 of the Tyr-Met-Asn-Met motif, inducing a 3- to 8-fold increase in p85 (subunit of PI 3-kinase) and GRB-2 SH2 binding to CD28"
  - statement: CD28 binds PI3K, GRB2, and ITK through the YMNM motif
    supporting_text: "CD28 interacts with three intracellular proteins-phosphatidylinositol 3-kinase (PI 3-kinase), T cell-specific protein-tyrosine kinase ITK (formerly TSK or EMT), and the complex between growth factor receptor-bound protein 2 and son of sevenless guanine nucleotide exchange protein (GRB-2-SOS)"
- id: PMID:7584133
  title: Selective CD28pYMNM mutations implicate phosphatidylinositol 3-kinase in CD86-CD28-mediated costimulation.
  findings:
  - statement: Y191 mutation disrupts both PI3K and GRB2 binding
    supporting_text: "Y191 mutation (Y191CD28F) disrupted both PI 3-kinase and GRB-2 binding"
  - statement: M194 mutation selectively disrupts PI3K binding
    supporting_text: "M194 mutation (M194CD28C) disrupted only PI 3-kinase binding"
  - statement: Both mutations abolished IL-2 production, directly implicating PI3K in CD28 costimulation
    supporting_text: "Both Y191CD28F and M194CD28C mutants failed to generate IL-2. These data directly implicate PI 3-kinase in CD28-mediated costimulation leading to IL-2 secretion"
- id: PMID:7807015
  title: CTLA-4 binding to the lipid kinase phosphatidylinositol 3-kinase in T cells.
  findings:
  - statement: Compared CD28 and CTLA-4 binding to PI3K p85 SH2 domains
    supporting_text: "CTLA-4 can also associate with PI 3-kinase as detected by lipid kinase analysis and immunoblotting with anti-p85 antiserum"
  - statement: Similar binding affinity of p85 SH2 domains for CD28 and CTLA-4 cytoplasmic motifs
    supporting_text: "the NH2- and COOH-terminal SH2 domains of p85 bind the CTLA-4 cytoplasmic pYVKM motif with an affinity (ID50: 0.6 and 0.04 microM), that is similar to CD28"
- id: PMID:8617933
  title: Cross-linking of the CD40 ligand on human CD4+ T lymphocytes generates a costimulatory signal that up-regulates IL-4 synthesis.
  findings:
  - statement: CD28 costimulation with CD3 and CD40L greatly enhances IL-4 synthesis
    supporting_text: "IL-4 synthesis was greatly enhanced by triggering of CD40L on the T cell surface in conjunction with ligation of CD3/TCR and CD28"
  - statement: CD28 alone with CD3 produces little IL-4 without CD40L
    supporting_text: "ligation of CD3/TCR and CD28 in the absence of CD40L triggering resulted in little or no IL-4 synthesis"
- id: PMID:8717514
  title: CD28/B7 system of T cell costimulation.
  findings:
  - statement: CD28 promotes cytokine production and inhibits apoptosis
    supporting_text: "The CD28/B7 receptor/ligand system is one of the dominant costimulatory pathways. Interruption of this signaling pathway with CD28 antagonists not only results in the suppression of the immune response, but in some cases induces antigen-specific tolerance"
- id: PMID:9417079
  title: Growth factor receptor-bound protein 2 SH2/SH3 domain binding to CD28 and its role in co-signaling.
  findings:
  - statement: Both GRB2 SH2 and SH3 domains contribute to CD28 binding
    supporting_text: "the Grb2 SH2 domain is critical for the association, while the SH3 domain plays an additional role in facilitating optimal binding"
  - statement: GRB2 binding to CD28 is required for IL-2 production and Vav phosphorylation
    supporting_text: "Mutations that alter Grb2 binding were found to block the CD28-dependent interleukin-2 production. Further, tyrosine phosphorylation of Vav and the costimulation-dependent activation of Jun N-terminal kinase was blocked"
  - statement: CD28/GRB2 interaction is required for JNK activation
    supporting_text: "the costimulation-dependent activation of Jun N-terminal kinase was blocked in cells defective in CD28/Grb2 binding"
- id: PMID:9784967
  title: "CD28/B7 costimulation: a review."
  findings:
  - statement: Comprehensive review of CD28 as a costimulatory receptor
    supporting_text: "The current model of T cell activation requires two signals. The first signal is specific, requiring T cell receptor recognition and binding to MHC/Antigen presented by an antigen-presenting cell. The second signal is nonspecific, resulting from the binding of B7 ligand on the antigen-presenting cell with its receptor, CD28, on the T cell"
- id: PMID:10820259
  title: "GRID: a novel Grb-2-related adapter protein that interacts with the activated T cell costimulatory receptor CD28."
  findings: []
- id: PMID:11279501
  title: Structural basis for co-stimulation by the human CTLA-4/B7-2 complex.
  findings:
  - statement: 3.2A crystal structure of CTLA-4/B7-2 complex
    supporting_text: "we report the 3.2-A resolution structure of the complex between the disulphide-linked homodimer of human CTLA-4 and the receptor-binding domain of human B7-2"
  - statement: Provides structural basis for understanding CD28/CTLA-4 engagement with B7 ligands
    supporting_text: "Regulation of T-cell activity is dependent on antigen-independent co-stimulatory signals provided by the disulphide-linked homodimeric T-cell surface receptors, CD28 and CTLA-4"
- id: PMID:11285224
  title: Mechanism for down-regulation of CD28 by Nef.
  findings: []
  full_text_unavailable: true
- id: PMID:11390434
  title: The transmembrane adaptor protein TRIM regulates T cell receptor (TCR) expression and TCR-mediated signaling via an association with the TCR zeta chain.
  findings:
  - statement: Confirmed CD28 cell surface expression on T cells
    supporting_text: "T cell receptor (TCR)-interacting molecule (TRIM) is a recently identified transmembrane adaptor protein, which is exclusively expressed in T cells"
- id: PMID:12196291
  title: The interaction properties of costimulatory molecules revisited.
  findings:
  - statement: Characterized CD28 binding to CD80 and CD86
    supporting_text: "B7-2 binds the two receptors more weakly than B7-1"
  - statement: Demonstrated CD28 stimulation increases proliferation and cytokine expression
    supporting_text: "relative to its CTLA-4 binding affinity, B7-2 binds CD28 2- to 3-fold more effectively than B7-1"
- id: PMID:15067037
  title: "The modulation of CD40 ligand signaling by transmembrane CD28 splice variant in human T cells."
  findings:
  - statement: Isoform 3 (CD28i) interacts with CD40LG and enhances NF-kB and MAPK8/PAK2 activation
    supporting_text: "In this study we show that CD28i, a transmembrane splice variant of CD28 costimulatory receptor, complexes with CD40L in human T cells"
  - statement: CD28i localizes to the cell surface
    supporting_text: "CD28i is unique among other isoforms as it retains an intact transmembrane region and cytoplasmic tail, but lacks the B7 ligand binding motif"
- id: PMID:15554700
  title: Characterization of Lck-binding elements in the herpesviral regulatory Tip protein.
  findings:
  - statement: Characterized Lck SH3 and SH2 domain binding properties (CD28 used as reference)
    supporting_text: "Herpesvirus saimiri encodes a tyrosine kinase interacting protein (Tip) that binds to T-cell-specific tyrosine kinase Lck via multiple sequence motifs and controls its activity"
- id: PMID:18337562
  title: CCL5-mediated T-cell chemotaxis involves the initiation of mRNA translation through mTOR/4E-BP1.
  findings:
  - statement: Focus on CCL5 signaling in T cells, not directly on CD28
    supporting_text: "we examined the role for CCL5-mediated initiation of mRNA translation in CD4(+) T-cell chemotaxis"
- id: PMID:18641304
  title: Acquisition of suppressive function by activated human CD4+ CD25- T cells is associated with the expression of CTLA-4 not FoxP3.
  findings:
  - statement: CTLA-4 is the key determinant for Treg suppressive function
    supporting_text: "the acquisition of suppressive behavior by activated CD4(+)CD25(-) T cells requires the expression of CTLA-4"
  - statement: CD28-dependent T cell activation leads to CTLA-4 and FoxP3 induction
    supporting_text: "Activation of human CD4(+)CD25(-) T cells resulted in the appearance of a de novo population of FoxP3-expressing cells within 48 h. These cells expressed high levels of CTLA-4"
- id: PMID:18641334
  title: ICOS ligation recruits the p50alpha PI3K regulatory subunit to the immunological synapse.
  findings: []
  full_text_unavailable: true
- id: PMID:21982860
  title: A secreted protein microarray platform for extracellular protein interaction discovery.
  findings:
  - statement: High-throughput platform identified CD28-CD80 interaction
    supporting_text: "functional protein microarrays are particularly well-suited for high-throughput screening of extracellular protein interactions"
- id: PMID:23817958
  title: Proinflammatory stimuli induce galectin-9 in human mesenchymal stromal cells to suppress T-cell proliferation.
  findings:
  - statement: CD28 signaling regulates both positive and negative gene expression programs
    supporting_text: "MSCs with a galectin-9 knockdown lost a significant portion of their antiproliferative effects on T cells"
- id: PMID:23918985
  title: Soluble CD80 restores T cell activation and overcomes tumor cell programmed death ligand 1-mediated immune suppression.
  findings:
  - statement: Direct evidence for CD28 in T cell activation
    supporting_text: "CD80-Fc facilitates T cell activation by binding to PDL1 to inhibit PDL1-PD1 interactions and by costimulating through CD28"
- id: PMID:32822567
  title: A Human IgSF Cell-Surface Interactome Reveals a Complex Network of Protein-Protein Interactions.
  findings:
  - statement: Systematic interactome identified CD28-CD80 and CD28-CD86 interactions
    supporting_text: "Cell-surface protein-protein interactions (PPIs) mediate cell-cell communication, recognition, and responses"
- id: PMID:35397202
  title: Structural characterization of a dimerization interface in the CD28 transmembrane domain.
  findings:
  - statement: NMR structure of CD28 TM domain confirming dimerization
    supporting_text: "we determined the dimeric helix-helix packing of CD28-TMH using nuclear magnetic resonance (NMR) technology"
  - statement: GxxxA motif mediates TM helix dimerization
    supporting_text: "a GxxxA motif, which is highly conserved in many dimeric assemblies, is located at the dimerization interface"
  - statement: Mutations in dimerization interface reduce IL-2 release
    supporting_text: "Mutating G160 and A164 can disrupt the transmembrane helix assembly and reduces CD28 enhancement in cells"
- id: PMID:35922511
  title: A physical wiring diagram for the human immune system.
  findings:
  - statement: Systematic mapping of immune cell surface interactions including CD28-CD80 and CD28-CD86
    supporting_text: "using a high-throughput surface receptor screening method, we systematically mapped the direct protein interactions across a recombinant library that encompasses most of the surface proteins that are detectable on human leukocytes"
- id: PMID:7492435
  title: Costimulation of CD4+ T cells via CD28 modulates human immunodeficiency virus type 1 infection and replication in vitro.
  findings: []
  full_text_unavailable: true
- id: PMID:8146197
  title: T-cell antigen CD28 interacts with the lipid kinase phosphatidylinositol 3-kinase by a cytoplasmic Tyr(P)-Met-Xaa-Met motif.
  findings: []
  full_text_unavailable: true
- id: PMID:8183372
  title: Binding of phosphatidylinositol-3-OH kinase to CD28 is required for T-cell signalling.
  findings: []
  full_text_unavailable: true
- id: PMID:12028592
  title: The regulation of protein synthesis and translation factors by CD3 and CD28 in human primary T lymphocytes.
  findings: []
  full_text_unavailable: true
- id: Reactome:R-HSA-389356
  title: Co-stimulation by CD28 (Reactome pathway)
  findings:
  - statement: Comprehensive pathway covering CD28 signaling including PI3K, Vav1, Grb2, Gads recruitment
    supporting_text: "CD28 signaling plays a critical role in shaping immune responses by ensuring effective T-cell activation and enhancing T-cell survival and proliferation"
- id: Reactome:R-HSA-164939
  title: Nef mediated downregulation of CD28 cell surface expression
  findings:
  - statement: HIV Nef hijacks CD28 for internalization
- id: Reactome:R-HSA-2316434
  title: PI3K phosphorylation of PIP2 to PIP3 in CD28 signaling
  findings: []
- id: Reactome:R-HSA-2400009
  title: PI3K inhibitors in CD28 signaling
  findings: []
- id: Reactome:R-HSA-388814
  title: Grb2 binding to CD28
  findings: []
- id: Reactome:R-HSA-388831
  title: Phosphorylation of CD28
  findings: []
- id: Reactome:R-HSA-388832
  title: PI3K binding to CD28
  findings: []
- id: Reactome:R-HSA-389158
  title: CD28-bound PI3K phosphorylation of PIP2 to PIP3
  findings: []
- id: Reactome:R-HSA-389348
  title: Activation of Rac1 by pVav1 in CD28 pathway
  findings: []
- id: Reactome:R-HSA-389350
  title: Activation of Cdc42 by pVav1 in CD28 pathway
  findings: []
- id: Reactome:R-HSA-389352
  title: Translocation of Vav1 to CD28
  findings: []
- id: Reactome:R-HSA-389354
  title: Activation of Vav1 in CD28 pathway
  findings: []
- id: Reactome:R-HSA-389381
  title: Gads binding to CD28
  findings: []
- id: Reactome:R-HSA-167630
  title: Formation of Nef-CD28 cytoplasmic tail complex
  findings: []
- id: Reactome:R-HSA-167633
  title: Formation of Nef-CD28-Clathrin-Coated Pit Adapter Protein complex
  findings: []
- id: Reactome:R-HSA-167637
  title: Internalization of Nef-CD28-Clathrin complex
  findings: []
- id: Reactome:R-HSA-388808
  title: CD28 homodimer binding B7-2 monomer
  findings: []
- id: Reactome:R-HSA-388811
  title: CD28 homodimer binding B7-1 homodimer
  findings: []
- id: file:human/CD28/CD28-deep-research-falcon.md
  title: Deep research review of CD28 biology (2024 literature synthesis)
  findings:
  - statement: Comprehensive review synthesizing 2024 literature on CD28 signaling, structure, and translational applications
  - statement: YMNM/PRRP/PYAP motifs recruit PI3K/GRB2, ITK/PLCgamma1, and Lck/PKCtheta
  - statement: CD28 costimulation accelerates Lck recruitment and ZAP70 activation at TCR microclusters

core_functions:
- molecular_function:
    id: GO:0015026
    label: coreceptor activity
  description: >-
    CD28 is the prototypic T cell coreceptor. It cooperates with the TCR by binding B7 ligands
    (CD80/CD86) on APCs to provide signal 2 for T cell activation. This is the single most precise
    molecular function term for CD28.
  directly_involved_in:
  - id: GO:0031295
    label: T cell costimulation
  - id: GO:0050862
    label: positive regulation of T cell receptor signaling pathway
  - id: GO:0042102
    label: positive regulation of T cell proliferation
  - id: GO:0032743
    label: positive regulation of interleukin-2 production
  locations:
  - id: GO:0009897
    label: external side of plasma membrane
  - id: GO:0001772
    label: immunological synapse

Gene Description

Existing Annotations Review

Core Functions

CD28 is the prototypic T cell coreceptor. It cooperates with the TCR by binding B7 ligands (CD80/CD86) on APCs to provide signal 2 for T cell activation. This is the single most precise molecular function term for CD28.

References

Deep Research

Falcon

Citations

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