id: Q96AY4 gene_symbol: TTC28 product_type: PROTEIN taxon: id: NCBITaxon:9606 label: Homo sapiens description: >- Tetratricopeptide repeat protein 28 (TTC28/TPRBK), a very large (~271 kDa, ~2365-2481 AA) protein built from ~25-28 tetratricopeptide repeat (TPR) motifs. TPR domains are helical repeat motifs that mediate protein-protein interactions, and TTC28 acts as a scaffold/adaptor rather than an enzyme. Mechanistic work (Zhang et al. 2024, PNAS) establishes that TTC28 is a substrate of HSPA8/HSC70 chaperone-mediated autophagy (CMA)/microautophagy: its TPR domains bind the C-terminal PTIEEVD motif of HSPA8, and it carries multiple KFERQ-like motifs, leading to LAMP2A-dependent lysosomal turnover. Functionally, TTC28 is required for the maintenance of chromosomal stability, acting through regulation of mitosis and cytokinesis. It is mainly cytoplasmic with perinuclear enrichment and localizes to mitotic structures including the midbody and spindle apparatus, where it colocalizes with beta-tubulin (TUBB) and partially overlaps Aurora B (AURKB). Loss of TTC28 increases micronuclei frequency and DNA-damage markers (gamma-H2AX, comet assay), and TTC28 is frequently mutated/down-regulated in cancers where its loss may contribute to chromosomal instability (CIN). existing_annotations: - term: id: GO:0000922 label: spindle pole evidence_type: IEA original_reference_id: GO_REF:0000044 review: summary: Phylogenetic inference for spindle pole localization. TTC28 concentrates at spindle poles during mitosis. action: ACCEPT reason: Core localization confirmed by IDA (PMID:23036704). supported_by: - reference_id: file:human/TTC28/TTC28-deep-research-falcon.md supporting_text: >- Detectable in mitotic structures including the **midbody**, consistent with involvement in cytokinesis. - term: id: GO:0005813 label: centrosome evidence_type: IEA original_reference_id: GO_REF:0000044 review: summary: Subcellular location annotation for centrosome. TTC28 localizes to centrosomes throughout cell cycle. action: ACCEPT reason: Well-established centrosomal localization. - term: id: GO:0005819 label: spindle evidence_type: IEA original_reference_id: GO_REF:0000044 review: summary: Subcellular location annotation for spindle. TTC28 localizes to mitotic spindle structures, consistent with falcon deep research reporting perinuclear/midbody localization and TUBB colocalization. action: ACCEPT reason: Consistent with spindle pole localization and microtubule association. supported_by: - reference_id: file:human/TTC28/TTC28-deep-research-falcon.md supporting_text: >- Proteomics identified **TUBB (β-tubulin)** as a TTC28-binding candidate, and confocal microscopy shows **TTC28/TUBB colocalization** in the perinuclear cytoplasm and **midbody**. - term: id: GO:0005856 label: cytoskeleton evidence_type: IEA original_reference_id: GO_REF:0000044 review: summary: Broad cytoskeleton term. TTC28 associates with microtubule cytoskeleton structures and perturbs tubulin gene expression on loss. action: KEEP_AS_NON_CORE reason: Too general. More specific spindle/centrosome/midbody terms preferred, but microtubule association is supported. supported_by: - reference_id: file:human/TTC28/TTC28-deep-research-falcon.md supporting_text: >- TTC28 knockout perturbs expression of tubulin-related genes (e.g., **TUBB6, TUBA1A, TTL**), reinforcing a functional link to microtubule dynamics. - term: id: GO:0030496 label: midbody evidence_type: IEA original_reference_id: GO_REF:0000044 review: summary: Duplicate midbody annotation with IEA evidence. action: ACCEPT reason: Consistent midbody localization. - term: id: GO:0051301 label: cell division evidence_type: IEA original_reference_id: GO_REF:0000043 review: summary: Cell division process. TTC28 functions in mitosis/cytokinesis. action: ACCEPT reason: Core biological process. supported_by: - reference_id: file:human/TTC28/TTC28-deep-research-falcon.md supporting_text: >- supports **high-fidelity mitosis/cytokinesis** and thereby reduces **chromosomal instability** - term: id: GO:0030496 label: midbody evidence_type: IDA original_reference_id: PMID:23036704 review: summary: Direct assay evidence for midbody localization from key paper. TTC28 concentrates at midbody during cytokinesis. Independently confirmed by falcon deep research (Zhang 2024 confocal imaging). action: ACCEPT reason: Experimental evidence from PMID:23036704, core localization. supported_by: - reference_id: PMID:23036704 supporting_text: Oct 1. A novel big protein TPRBK possessing 25 units of TPR motif is essential for the progress of mitosis and cytokinesis. - reference_id: file:human/TTC28/TTC28-deep-research-falcon.md supporting_text: >- Detectable in mitotic structures including the **midbody**, consistent with involvement in cytokinesis. - term: id: GO:0007346 label: regulation of mitotic cell cycle evidence_type: IMP original_reference_id: PMID:23036704 review: summary: Mutant phenotype evidence for regulation of mitotic cell cycle. TTC28 depletion disrupts cell division progression. Falcon deep research (Zhang 2024) reinforces a mitosis/cytokinesis regulatory role required for chromosomal stability. action: ACCEPT reason: Experimental evidence from PMID:23036704, core biological process. supported_by: - reference_id: PMID:23036704 supporting_text: Oct 1. A novel big protein TPRBK possessing 25 units of TPR motif is essential for the progress of mitosis and cytokinesis. - reference_id: file:human/TTC28/TTC28-deep-research-falcon.md supporting_text: >- a large TPR scaffold/adaptor that couples chaperone/autophagy machinery to the fidelity of mitotic and cytokinetic processes, likely through protein interaction networks rather than enzymatic catalysis - term: id: GO:0019900 label: kinase binding evidence_type: IPI original_reference_id: PMID:23036704 review: summary: >- Kinase binding from protein interaction study. Falcon deep research (Zhang 2024) provides specific support: TTC28 partially overlaps and is linked to Aurora B kinase (AURKB) at mitotic structures, consistent with a TPR scaffold engaging a mitotic kinase. Note the best-supported molecular interaction is with the HSPA8/HSC70 chaperone (via its C-terminal PTIEEVD motif), which is an ATPase but is more precisely a chaperone-binding (CMA substrate) relationship than generic kinase binding. action: KEEP_AS_NON_CORE reason: >- The kinase-binding annotation is plausible (AURKB association reported), but falcon evidence indicates the dominant, mechanistically defining interaction is HSPA8 chaperone binding driving CMA turnover, not a catalytic-kinase scaffolding function. Retained as non-core pending identification of the specific kinase(s) bound. supported_by: - reference_id: file:human/TTC28/TTC28-deep-research-falcon.md supporting_text: >- TTC28 shows **partial overlap** with **AURKB (Aurora B kinase)** - term: id: GO:0007049 label: cell cycle evidence_type: IMP original_reference_id: PMID:39630868 negated: false review: summary: >- NEW annotation grounded in Zhang et al. 2024 (PNAS): TTC28 is required for high-fidelity mitosis and cytokinesis, and its loss increases micronuclei frequency and DNA-damage markers, placing its activity within the cell cycle. action: NEW reason: >- Zhang et al. 2024 demonstrate by mutant/knockout phenotype (IMP) that loss of TTC28 increases micronuclei frequency ~3-fold and that TTC28 regulates mitosis and cytokinesis to maintain genome integrity. This supports the broader cell-cycle context; the more specific regulation of mitotic cell cycle term remains the core BP. supported_by: - reference_id: PMID:39630868 supporting_text: >- the baseline frequency of micronuclei (FMN) in human cancer cells with TTC28 knockout cells was three times greater than that in cells with wild-type TTC28 (7.7% vs. 2.3%, P = 4.86E-09). - reference_id: file:human/TTC28/TTC28-deep-research-falcon.md supporting_text: >- It reports cell-cycle regulation of TTC28 abundance and multiple genome instability readouts (micronuclei, γH2AX, comet assays). - term: id: GO:0030544 label: Hsp70 protein binding evidence_type: IPI original_reference_id: PMID:39630868 review: summary: >- NEW annotation from falcon deep research: TTC28 directly binds the HSPA8 (HSC70, an Hsp70-family chaperone) via its TPR domains engaging the HSPA8 C-terminal PTIEEVD motif. This is the best-supported molecular interaction of TTC28 and underlies its turnover by chaperone-mediated autophagy. action: NEW reason: >- Direct interaction with HSPA8 (an Hsp70-family member) demonstrated by Zhang et al. 2024 (PNAS) via CoIP/BiFC/mutant analysis; a TPR-PTIEEVD chaperone-binding mode. The specific Hsp70 protein binding term is more informative than the generic heat shock protein binding parent. supported_by: - reference_id: file:human/TTC28/TTC28-deep-research-falcon.md supporting_text: >- TTC28 **directly binds HSPA8** through the **HSPA8 C-terminal PTIEEVD motif** - reference_id: PMID:39630868 supporting_text: >- The tetratricopeptide repeat domains of TTC28 bind to the C-terminal motif (PTIEEVD) in HSPA8, resulting in the subsequent degradation of TTC28 via CMA/microautophagy. - term: id: GO:0061684 label: chaperone-mediated autophagy evidence_type: IDA original_reference_id: PMID:39630868 review: summary: >- NEW annotation from falcon deep research: TTC28 is a substrate of HSPA8/LAMP2A-dependent chaperone-mediated autophagy (CMA)/microautophagy and carries multiple KFERQ-like motifs. CMA control of TTC28 abundance is itself required for maintenance of genome stability. action: NEW reason: >- Established as a CMA substrate by Zhang et al. 2024 (PNAS); CMA-mediated TTC28 degradation is a master regulator of TTC28's genome-stability function. supported_by: - reference_id: file:human/TTC28/TTC28-deep-research-falcon.md supporting_text: >- TTC28 also contains **16 KFERQ-like motifs**, consistent with CMA targeting logic. - reference_id: PMID:39630868 supporting_text: >- the subsequent degradation of TTC28 via CMA/microautophagy. - term: id: GO:0005829 label: cytosol evidence_type: IDA original_reference_id: PMID:39630868 review: summary: >- NEW annotation from falcon deep research: TTC28 is mainly cytoplasmic with perinuclear enrichment, the compartment where it engages cytosolic chaperones (HSPA8) and mitotic/cytoskeletal machinery. action: NEW reason: >- Imaging/fractionation in human cancer cell lines shows predominantly cytoplasmic localization (Zhang et al. 2024, PNAS); consistent with cytosolic chaperone engagement. supported_by: - reference_id: file:human/TTC28/TTC28-deep-research-falcon.md supporting_text: >- Mainly cytoplasmic**, with **perinuclear enrichment** references: - id: GO_REF:0000043 title: Gene Ontology annotation based on UniProtKB/Swiss-Prot keyword mapping findings: [] - id: GO_REF:0000044 title: Gene Ontology annotation based on UniProtKB/Swiss-Prot Subcellular Location vocabulary mapping, accompanied by conservative changes to GO terms applied by UniProt. findings: [] - id: PMID:23036704 title: A novel big protein TPRBK possessing 25 units of TPR motif is essential for the progress of mitosis and cytokinesis. findings: [] - id: PMID:39630868 title: The essential role of TTC28 in maintaining chromosomal stability via HSPA8 chaperone-mediated autophagy. findings: - statement: >- The tetratricopeptide repeat domains of TTC28 bind the C-terminal PTIEEVD motif of HSPA8, targeting TTC28 for degradation via CMA/microautophagy; TTC28 is thus an HSPA8-mediated CMA substrate. supporting_text: >- The tetratricopeptide repeat domains of TTC28 bind to the C-terminal motif (PTIEEVD) in HSPA8, resulting in the subsequent degradation of TTC28 via CMA/microautophagy. reference_section_type: ABSTRACT - statement: >- Loss of TTC28 increases micronuclei frequency ~3-fold, and TTC28 overexpression rescues this, demonstrating TTC28 is required for chromosomal stability. supporting_text: >- the baseline frequency of micronuclei (FMN) in human cancer cells with TTC28 knockout cells was three times greater than that in cells with wild-type TTC28 (7.7% vs. 2.3%, P = 4.86E-09). reference_section_type: ABSTRACT - statement: >- Mechanistically, TTC28 maintains genome integrity by regulating mitosis and cytokinesis, downstream of CMA. supporting_text: >- Mechanistically, TTC28 regulates mitosis and cytokinesis, which are involved in the maintenance of genome integrity by CMA. reference_section_type: ABSTRACT - statement: >- TTC28 is a conserved vertebrate gene that is frequently mutated and down-regulated in human cancers, where comprehensive loss may drive chromosomal instability. supporting_text: >- TTC28, a conserved gene in vertebrates, is frequently mutated and down-regulated in numerous human cancers. reference_section_type: ABSTRACT - id: file:human/TTC28/TTC28-deep-research-falcon.md title: Falcon (Edison Scientific Literature) deep research report on human TTC28 (Q96AY4) function and disease relevance. findings: - statement: >- TTC28 is a very large (~271 kDa) TPR-containing scaffold protein with ~28 TPR domains and multiple KFERQ-like motifs consistent with recognition by chaperone-mediated autophagy. supporting_text: >- TTC28 as containing **28 TPR domains** and multiple KFERQ-like motifs consistent with recognition by **chaperone-mediated autophagy (CMA)** reference_section_type: RESULTS - statement: >- TTC28 is mainly cytoplasmic with perinuclear enrichment, consistent with a TPR scaffold acting where chaperone pathways intersect mitotic machinery. supporting_text: >- Mainly cytoplasmic**, with **perinuclear enrichment** reference_section_type: RESULTS - statement: >- The strongest functional phenotype of TTC28 is chromosomal/genome stability, assayed by micronuclei frequency and DNA damage markers. supporting_text: >- The strongest functional phenotype linked to TTC28 is **chromosomal/genome stability** as assayed by micronuclei frequency and DNA damage markers. reference_section_type: RESULTS - statement: >- TTC28 is best interpreted as a TPR scaffold/adaptor (not an enzyme) coupling chaperone/autophagy machinery to mitotic and cytokinetic fidelity. supporting_text: >- a large TPR scaffold/adaptor that couples chaperone/autophagy machinery to the fidelity of mitotic and cytokinetic processes, likely through protein interaction networks rather than enzymatic catalysis reference_section_type: DISCUSSION - statement: >- TTC28 is not established as an enzyme with defined substrate specificity; the evidence is interaction/scaffold/quality-control centric. supporting_text: >- TTC28 is **not** established here as an enzyme with defined substrate specificity; the evidence is interaction/scaffold/quality-control centric. reference_section_type: DISCUSSION aliases: - KIAA1043 - TPRBK - TPR repeat protein 28 core_functions: - description: >- Scaffolds protein-protein interactions via TPR repeats to organize and support high-fidelity mitosis and cytokinesis (at centrosomes, spindle poles, and the midbody), thereby maintaining chromosomal stability. The defining molecular interaction is Hsp70 (HSPA8/HSC70) binding through the TPR domains engaging the HSPA8 C-terminal PTIEEVD motif (Zhang et al. 2024). TTC28 colocalizes with beta-tubulin and partially overlaps Aurora B at mitotic structures; loss increases micronuclei and DNA damage. molecular_function: id: GO:0030544 label: Hsp70 protein binding directly_involved_in: - id: GO:0051301 label: cell division - id: GO:0007346 label: regulation of mitotic cell cycle locations: - id: GO:0005813 label: centrosome - id: GO:0000922 label: spindle pole - id: GO:0030496 label: midbody supported_by: - reference_id: file:human/TTC28/TTC28-uniprot.txt supporting_text: TPR repeat protein localizing to centrosome, spindle pole, and midbody during cell division - reference_id: file:human/TTC28/TTC28-deep-research-falcon.md supporting_text: >- supports **high-fidelity mitosis/cytokinesis** and thereby reduces **chromosomal instability** - description: >- Acts as a substrate of HSPA8/HSC70 chaperone-mediated autophagy: its TPR domains bind the HSPA8 C-terminal PTIEEVD motif and, with multiple KFERQ-like motifs, target TTC28 for LAMP2A-dependent lysosomal turnover. This CMA control of TTC28 abundance is itself important for genome stability. molecular_function: id: GO:0030544 label: Hsp70 protein binding directly_involved_in: - id: GO:0061684 label: chaperone-mediated autophagy locations: - id: GO:0005829 label: cytosol supported_by: - reference_id: file:human/TTC28/TTC28-deep-research-falcon.md supporting_text: >- TTC28 **directly binds HSPA8** through the **HSPA8 C-terminal PTIEEVD motif** - reference_id: PMID:39630868 supporting_text: >- The tetratricopeptide repeat domains of TTC28 bind to the C-terminal motif (PTIEEVD) in HSPA8, resulting in the subsequent degradation of TTC28 via CMA/microautophagy. proposed_new_terms: [] suggested_questions: - question: Which specific kinases bind TTC28 and how does this regulate mitosis? experts: - Cell cycle researchers - question: Is TTC28's role in chromosomal stability mediated mainly through its own scaffolding activity at the midbody/spindle, or indirectly through CMA-controlled turnover of its abundance? experts: - Autophagy researchers - Cell cycle researchers suggested_experiments: - description: Co-IP and mass spec to identify TTC28-binding kinases experiment_type: proteomics hypothesis: TTC28 scaffolds mitotic kinases - description: >- Separation-of-function mutants disrupting the TPR-PTIEEVD (HSPA8) interface vs. midbody localization, assayed for micronuclei frequency, to dissect whether CMA targeting or mitotic scaffolding is the primary driver of chromosomal stability. experiment_type: mutational_analysis hypothesis: >- Mitotic scaffolding by TTC28 maintains chromosomal stability independently of its CMA-mediated turnover. status: COMPLETE