CNKSR2

Summary: Refine the Densin interaction to PDZ-domain binding.

Reason: The original human brain-library screen recovered human MAGUIN-1 fragments with the Densin C-terminal PDZ region; the exact positive MINT record preserves that source pair. Subsequent binding-site and ternary-complex experiments used rat MAGUIN-1 fragments, with loss of binding after deleting terminal THV. This supports the specific domain interaction while preserving human screening versus rat mechanistic scope. The terminal motif is absent from current human products Q8WXI2-2 and -4.

Proposed replacements: PDZ domain binding

Summary: Refine the positive cross-species SAM-fragment interaction to SAM domain binding.

Reason: Purified human CNK2 residues 5–84 bind the Drosophila HYP SAM domain (residues 21–98). Original pull-down controls, size exclusion, surface plasmon resonance, crystallography and interface mutations establish the SAM-domain contact directly. GO:0032093 identifies the experimentally recognized domain class; the observed discrete 1:1 heterodimer remains a property of this same assay rather than a second replacement. The original fly partner and human fragment scope are preserved. Cellular RAF-pathway rescue uses fly proteins and does not establish a native human HYP/KSR complex or human kinase activity.

Proposed replacements: SAM domain binding

Summary: Refine the EGFR interaction to epidermal growth factor receptor binding.

Reason: Actual human-library Methods and Results identify Cnk2 as a positive EGFR-WT interactor in the mammalian membrane two-hybrid assay and confirm the pair by tagged coimmunoprecipitation. Cnk2 has similar binding patterns with wild-type and tested oncogenic EGFR variants. The exact two MINT records retain the assay and construct tags. The detailed TACC3-dependent cancer mechanisms in the same paper are not assigned to CNKSR2.

Proposed replacements: epidermal growth factor receptor binding

Summary: Refine the GRID2IP fragment interaction to PDZ-domain binding.

Reason: The exact positive holdup records measure the biotinylated human CNKSR2 C-terminal peptide 1025–1034 against GRID2IP PDZ fragment(s) 1-87, 272-370. These domain–motif assays establish PDZ-domain binding, not a demonstrated endogenous full-length complex or the partner protein’s other functions. The tested tail is present in Q8WXI2-1 and -5 but absent from -2 and -4; retention in -5 is sequence context, not a full-protein assay.

Proposed replacements: PDZ domain binding

Summary: Refine the MPDZ fragment interaction to PDZ-domain binding.

Reason: The exact positive holdup records measure the biotinylated human CNKSR2 C-terminal peptide 1025–1034 against MPDZ PDZ fragment(s) 1858-1949, 1624-1716, 688-793. These domain–motif assays establish PDZ-domain binding, not a demonstrated endogenous full-length complex or the partner protein’s other functions. The tested tail is present in Q8WXI2-1 and -5 but absent from -2 and -4; retention in -5 is sequence context, not a full-protein assay.

Proposed replacements: PDZ domain binding

Summary: Refine the DLG4 fragment interaction to PDZ-domain binding.

Reason: The exact positive holdup records measure the biotinylated human CNKSR2 C-terminal peptide 1025–1034 against DLG4 PDZ fragment(s) 155-259, 62-152. These domain–motif assays establish PDZ-domain binding, not a demonstrated endogenous full-length complex or the partner protein’s other functions. The tested tail is present in Q8WXI2-1 and -5 but absent from -2 and -4; retention in -5 is sequence context, not a full-protein assay.

Proposed replacements: PDZ domain binding

Summary: Refine the APBA1 fragment interaction to PDZ-domain binding.

Reason: The exact positive holdup records measure the biotinylated human CNKSR2 C-terminal peptide 1025–1034 against APBA1 PDZ fragment(s) 655-741, 743-822. These domain–motif assays establish PDZ-domain binding, not a demonstrated endogenous full-length complex or the partner protein’s other functions. The tested tail is present in Q8WXI2-1 and -5 but absent from -2 and -4; retention in -5 is sequence context, not a full-protein assay. The APBA1 655–741 record also carries its explicit residue-655 mutation feature.

Proposed replacements: PDZ domain binding

Summary: Refine the TJP1 fragment interaction to PDZ-domain binding.

Reason: The exact positive holdup records measure the biotinylated human CNKSR2 C-terminal peptide 1025–1034 against TJP1 PDZ fragment(s) 184-264, 415-516. These domain–motif assays establish PDZ-domain binding, not a demonstrated endogenous full-length complex or the partner protein’s other functions. The tested tail is present in Q8WXI2-1 and -5 but absent from -2 and -4; retention in -5 is sequence context, not a full-protein assay.

Proposed replacements: PDZ domain binding

Summary: Refine the DLG1 fragment interaction to PDZ-domain binding.

Reason: The exact positive holdup records measure the biotinylated human CNKSR2 C-terminal peptide 1025–1034 against DLG1 PDZ fragment(s) 221-314, 457-570, 314-412. These domain–motif assays establish PDZ-domain binding, not a demonstrated endogenous full-length complex or the partner protein’s other functions. The tested tail is present in Q8WXI2-1 and -5 but absent from -2 and -4; retention in -5 is sequence context, not a full-protein assay.

Proposed replacements: PDZ domain binding

Summary: Refine the IL16 fragment interaction to PDZ-domain binding.

Reason: The exact positive holdup records measure the biotinylated human CNKSR2 C-terminal peptide 1025–1034 against IL16 PDZ fragment(s) 1106-1201, 1203-1332. These domain–motif assays establish PDZ-domain binding, not a demonstrated endogenous full-length complex or the partner protein’s other functions. The tested tail is present in Q8WXI2-1 and -5 but absent from -2 and -4; retention in -5 is sequence context, not a full-protein assay.

Proposed replacements: PDZ domain binding

Summary: Refine the SCRIB fragment interaction to PDZ-domain binding.

Reason: The exact positive holdup records measure the biotinylated human CNKSR2 C-terminal peptide 1025–1034 against SCRIB PDZ fragment(s) 858-949, 718-814, 998-1093. These domain–motif assays establish PDZ-domain binding, not a demonstrated endogenous full-length complex or the partner protein’s other functions. The tested tail is present in Q8WXI2-1 and -5 but absent from -2 and -4; retention in -5 is sequence context, not a full-protein assay.

Proposed replacements: PDZ domain binding

Summary: Refine the DLG2 fragment interaction to PDZ-domain binding.

Reason: The exact positive holdup records measure the biotinylated human CNKSR2 C-terminal peptide 1025–1034 against DLG2 PDZ fragment(s) 418-513, 189-283. These domain–motif assays establish PDZ-domain binding, not a demonstrated endogenous full-length complex or the partner protein’s other functions. The tested tail is present in Q8WXI2-1 and -5 but absent from -2 and -4; retention in -5 is sequence context, not a full-protein assay.

Proposed replacements: PDZ domain binding

Summary: Refine the PDZK1 fragment interaction to PDZ-domain binding.

Reason: The exact positive holdup records measure the biotinylated human CNKSR2 C-terminal peptide 1025–1034 against PDZK1 PDZ fragment(s) 375-459, 238-329, 2-107. These domain–motif assays establish PDZ-domain binding, not a demonstrated endogenous full-length complex or the partner protein’s other functions. The tested tail is present in Q8WXI2-1 and -5 but absent from -2 and -4; retention in -5 is sequence context, not a full-protein assay.

Proposed replacements: PDZ domain binding

Summary: Refine the FRMPD2 fragment interaction to PDZ-domain binding.

Reason: The exact positive holdup records measure the biotinylated human CNKSR2 C-terminal peptide 1025–1034 against FRMPD2 PDZ fragment(s) 770-861. These domain–motif assays establish PDZ-domain binding, not a demonstrated endogenous full-length complex or the partner protein’s other functions. The tested tail is present in Q8WXI2-1 and -5 but absent from -2 and -4; retention in -5 is sequence context, not a full-protein assay.

Proposed replacements: PDZ domain binding

Summary: Refine the MAGI2 fragment interaction to PDZ-domain binding.

Reason: The exact positive holdup records measure the biotinylated human CNKSR2 C-terminal peptide 1025–1034 against MAGI2 PDZ fragment(s) 772-865, 915-1017, 1141-1229, 600-682, 410-522. These domain–motif assays establish PDZ-domain binding, not a demonstrated endogenous full-length complex or the partner protein’s other functions. The tested tail is present in Q8WXI2-1 and -5 but absent from -2 and -4; retention in -5 is sequence context, not a full-protein assay.

Proposed replacements: PDZ domain binding

Summary: Refine the NHERF4 fragment interaction to PDZ-domain binding.

Reason: The exact positive holdup records measure the biotinylated human CNKSR2 C-terminal peptide 1025–1034 against NHERF4 PDZ fragment(s) 220-308, 464-550. These domain–motif assays establish PDZ-domain binding, not a demonstrated endogenous full-length complex or the partner protein’s other functions. The tested tail is present in Q8WXI2-1 and -5 but absent from -2 and -4; retention in -5 is sequence context, not a full-protein assay. A separate related Q86UT5-2 fragment record is retained in the evidence index and is not silently relabeled as the canonical partner.

Proposed replacements: PDZ domain binding

Summary: Refine the LNX2 fragment interaction to PDZ-domain binding.

Reason: The exact positive holdup records measure the biotinylated human CNKSR2 C-terminal peptide 1025–1034 against LNX2 PDZ fragment(s) 459-560, 334-427. These domain–motif assays establish PDZ-domain binding, not a demonstrated endogenous full-length complex or the partner protein’s other functions. The tested tail is present in Q8WXI2-1 and -5 but absent from -2 and -4; retention in -5 is sequence context, not a full-protein assay.

Proposed replacements: PDZ domain binding

Summary: Refine the PATJ fragment interaction to PDZ-domain binding.

Reason: The exact positive holdup records measure the biotinylated human CNKSR2 C-terminal peptide 1025–1034 against PATJ PDZ fragment(s) 113-229, 1673-1763, 547-651, 238-335, 1425-1530, 673-780, 1058-1167. These domain–motif assays establish PDZ-domain binding, not a demonstrated endogenous full-length complex or the partner protein’s other functions. The tested tail is present in Q8WXI2-1 and -5 but absent from -2 and -4; retention in -5 is sequence context, not a full-protein assay.

Proposed replacements: PDZ domain binding

Summary: Refine the LNX1 fragment interaction to PDZ-domain binding.

Reason: The exact positive holdup records measure the biotinylated human CNKSR2 C-terminal peptide 1025–1034 against LNX1 PDZ fragment(s) 371-470, 500-596, 261-371. These domain–motif assays establish PDZ-domain binding, not a demonstrated endogenous full-length complex or the partner protein’s other functions. The tested tail is present in Q8WXI2-1 and -5 but absent from -2 and -4; retention in -5 is sequence context, not a full-protein assay.

Proposed replacements: PDZ domain binding

Summary: Refine the DLG3 fragment interaction to PDZ-domain binding.

Reason: The exact positive holdup records measure the biotinylated human CNKSR2 C-terminal peptide 1025–1034 against DLG3 PDZ fragment(s) 222-314, 381-477, 124-219. These domain–motif assays establish PDZ-domain binding, not a demonstrated endogenous full-length complex or the partner protein’s other functions. The tested tail is present in Q8WXI2-1 and -5 but absent from -2 and -4; retention in -5 is sequence context, not a full-protein assay.

Proposed replacements: PDZ domain binding

Summary: Refine the LRRC7 fragment interaction to PDZ-domain binding.

Reason: The exact positive holdup records measure the biotinylated human CNKSR2 C-terminal peptide 1025–1034 against LRRC7 PDZ fragment(s) 1445-1536. These domain–motif assays establish PDZ-domain binding, not a demonstrated endogenous full-length complex or the partner protein’s other functions. The tested tail is present in Q8WXI2-1 and -5 but absent from -2 and -4; retention in -5 is sequence context, not a full-protein assay.

Proposed replacements: PDZ domain binding

Summary: Refine the APBA2 fragment interaction to PDZ-domain binding.

Reason: The exact positive holdup records measure the biotinylated human CNKSR2 C-terminal peptide 1025–1034 against APBA2 PDZ fragment(s) 655-735, 565-653. These domain–motif assays establish PDZ-domain binding, not a demonstrated endogenous full-length complex or the partner protein’s other functions. The tested tail is present in Q8WXI2-1 and -5 but absent from -2 and -4; retention in -5 is sequence context, not a full-protein assay.

Proposed replacements: PDZ domain binding

Summary: Refine the GRIP2 fragment interaction to PDZ-domain binding.

Reason: The exact positive holdup records measure the biotinylated human CNKSR2 C-terminal peptide 1025–1034 against GRIP2 PDZ fragment(s) 446-548, 544-642, 648-740, 238-339, 934-1023. These domain–motif assays establish PDZ-domain binding, not a demonstrated endogenous full-length complex or the partner protein’s other functions. The tested tail is present in Q8WXI2-1 and -5 but absent from -2 and -4; retention in -5 is sequence context, not a full-protein assay.

Proposed replacements: PDZ domain binding

Summary: Refine the WHRN fragment interaction to PDZ-domain binding.

Reason: The exact positive holdup records measure the biotinylated human CNKSR2 C-terminal peptide 1025–1034 against WHRN PDZ fragment(s) 813-904, 136-226. These domain–motif assays establish PDZ-domain binding, not a demonstrated endogenous full-length complex or the partner protein’s other functions. The tested tail is present in Q8WXI2-1 and -5 but absent from -2 and -4; retention in -5 is sequence context, not a full-protein assay.

Proposed replacements: PDZ domain binding

Summary: Refine the GRIP1 fragment interaction to PDZ-domain binding.

Reason: The exact positive holdup records measure the biotinylated human CNKSR2 C-terminal peptide 1025–1034 against GRIP1 PDZ fragment(s) 145-239, 464-567, 242-344, 563-659, 996-1086. These domain–motif assays establish PDZ-domain binding, not a demonstrated endogenous full-length complex or the partner protein’s other functions. The tested tail is present in Q8WXI2-1 and -5 but absent from -2 and -4; retention in -5 is sequence context, not a full-protein assay.

Proposed replacements: PDZ domain binding

Summary: Accept the cytoplasmic pool of CNKSR2.

Reason: The combined InterPro and UniProt-location mapping agrees with the human CNK2 study, which reports both membrane and cytoplasmic fractions. CNKSR2 is a multidomain peripheral scaffold rather than an integral membrane channel.

Summary: Accept conserved plasma-membrane association.

Reason: Direct human CNK2 evidence reports lateral plasma-membrane localization in MDCK cells. The genuine IBD record lists the target itself and rat Cnksr2 as descendant seeds at PTN002656117; actual PANTHER topology places all these leaves within the CNKSR2 clade. Target self-inclusion is legitimate experimental grounding. Current rat same-term records are propagated, but the separate IBD seed role is now verified rather than inferred from those records alone.

Summary: Accept directly reported human CNK2 membrane localization.

Reason: The complete original abstract explicitly reports human CNK2 in membrane and cytoplasmic fractions and full-length CNK2 at the lateral plasma membrane of MDCK cells. This positive localization is retained at that construct and heterologous-cell scope; unrecovered detailed Methods are not claimed read.

Summary: Retain inherited receptor-signaling context.

Reason: The genuine Bilateria-node IBD explicitly uses FlyBase cnk as its seed, and the actual tree places human CNKSR2 below that node. Fly CNK/HYP/KSR experiments establish coordinated RAF signaling; human CNK2 binds EGFR, Raf and Rlf, with complementary mammalian neuronal scaffold evidence. Retain the inherited broad receptor-signaling context without importing a particular fly developmental phenotype or intrinsic human catalytic activity.

Summary: Accept regulation of signal transduction through spatial organization.

Reason: The InterPro mapping is consistent with the experimentally defined CNK2 scaffold: disrupting its Vilse interaction alters localization and Rac-GTP balance in neuronal models without changing the isolated GAP activity of Vilse. CNKSR2 organizes the signaling machinery rather than supplying GAP or kinase chemistry.

Summary: Accept the conserved postsynaptic-density location.

Reason: The mouse ortholog Q80YA9 has a current IDA source for this location, although that phosphoproteomic target table was not recovered. Independent endogenous rat-neuron localization with PSD-95, and mouse postsynaptic functional studies, support the conserved location. The original Ensembl evidence and source identifiers remain intact; this is not direct human ultrastructural evidence.

Summary: Refine broad membrane localization to plasma membrane.

Reason: The InterPro/UniProt location mapping is supported more specifically by the human CNK2 lateral plasma-membrane result and by membrane-associated neuronal scaffold experiments. The protein is peripherally associated, and localization depends on the tested C-terminal region; a transmembrane topology is not implied. This source-specific refinement consolidates broad membrane coverage into an already supported plasma-membrane term; it is not a NEW annotation or additional activity.

Proposed replacements: plasma membrane

Summary: Refine Raf binding to protein serine/threonine kinase binding.

Reason: The original positive human CNK2 report maps binding to Raf regulatory and kinase regions. Official RAF1 identity establishes the Ser/Thr kinase class. This is a refinement of the binding partner class, not a claim that CNKSR2 is a kinase or that every Raf interaction activates signaling.

Proposed replacements: protein serine/threonine kinase binding

Summary: Represent the organizing activity as protein-complex scaffold activity.

Reason: The genuine IBD at the Bilateria node PTN000303668 is seeded by fly cnk and human CNKSR1. The actual PANTHER tree places CNKSR2 inside that broader family, on the distinct PTN002656117 branch; selected alignment inspection confirms the target sequence and shared SAM-domain region. Those data support the inherited organizing role and do not show that the original ancestral placement was erroneous. GO:0140378 is the more precise description of the independently characterized CNKSR2-lineage mechanism: human screening and rat MAGUIN-1 ternary-assembly experiments establish an integral organizing component holding Densin and PSD-95 together. The replacement applies to this target review, not a proposed wholesale relabeling of the Bilateria ancestor or every CNKSR1/fly activity. Current GO guidance explicitly distinguishes complex scaffolds from generic adaptors; neuronal recruitment studies complement this scaffold mechanism.

Proposed replacements: protein complex scaffold activity

Summary: Accept intracellular signaling through CNK2 partner organization.

Reason: The original human CNK2 study reports Raf, Ral and Rlf interactions and proposes integration of their signals; the detailed mechanistic read is abstract-limited. Independently, full neuronal studies connect CNK2 binding-site manipulations to partner localization and Rac-GTP readouts. The broad process fits this organizing role without assigning intrinsic kinase, GEF or GAP activity.

Summary: Accept conserved self-association without fixing a dimer stoichiometry.

Reason: The rat donor has a positive interaction annotation from the original MAGUIN study. Actual deletion and coimmunoprecipitation experiments map self-association to its C-terminal leucine-rich region, described as dimer or multimer. The corresponding long-form region is absent from short human products Q8WXI2-2 and -4. Retain identical-protein binding without asserting a resolved universal homodimer or all-product activity.

Summary: Retain neuronal cell-body localization as a secondary compartment.

Reason: The authentic rat MAGUIN abstract explicitly reports neuronal cell bodies and neurites; the donor IDA relationship is preserved. The principal organizing function is at neuronal postsynaptic sites. Cell-body detection is not evidence for a separate molecular activity.

Summary: Accept the conserved postsynaptic membrane association.

Reason: Rat MAGUIN localizes to neuronal membranes and coprecipitates with postsynaptic MAGUK proteins, and later endogenous rat hippocampal imaging places CNK2 with PSD-95. This supports the transferred postsynaptic membrane location at a peripheral scaffold scope, not a transmembrane or ion-channel role.

Summary: Retain urinary-exosome proteomic detection.

Reason: The original laboratory database explicitly lists CNKSR2/NP_055742 with two peptides under study reference 2, which is this publication, in urinary exosomes from healthy human volunteers. This target-specific detection supports the existing location at proteomic scope; it does not establish a secretion mechanism or an extracellular scaffold function.

Summary: Accept localization at glutamatergic synapses.

Reason: The doubled namespace in original MGI:MGI:2661175 is preserved and resolved to official mouse Cnksr2/Q80YA9; RGD:708454 resolves to rat Cnksr2/Q9Z1T4. Actual rodent neuronal studies show postsynaptic localization and functional association with excitatory synapses. Those positive descendant sources and the now-inspected IBD seed list and actual node-to-target tree path support transfer; no full historical curation-session reconstruction is claimed.

Summary: Accept the ortholog-supported glutamatergic synapse location.

Reason: Both exact supplied proteins are Cnksr2 orthologs. Rat endogenous CNK2 overlaps with PSD-95 in dendritic spines, while mouse experiments establish postsynaptic AMPA-receptor clustering and excitatory-response effects. Source species and tagged-versus-endogenous experiments remain distinct.

Summary: Accept conserved postsynaptic-specialization organization.

Reason: The mapped mouse and rat Cnksr2 studies directly connect the scaffold to PSD composition and organization: perturbations change PSD-95 or Homer content, and interaction-region manipulations change partner recruitment. Mouse loss reduces PSD-95 signal without reducing cluster density; rat results likewise distinguish PSD size from spine number. This supports the existing inherited process, not a claim that CNKSR2 catalyzes receptor signaling. Current IBD seeds and the actual CNKSR2-clade topology independently resolve the source provenance.

Summary: Accept the ortholog-based postsynaptic organization assertion.

Reason: The exact mouse and rat donor records have experimental grounding. Actual mouse knockout and rat knockdown/construct studies alter postsynaptic protein accumulation, with membrane-binding and partner-binding mechanisms independently examined. The broad organization role is supported; every measured phenotype is not promoted to a separate human process.

Summary: Accept peripheral association with the postsynaptic-density membrane.

Reason: The exact rat ortholog has direct neuronal localization evidence. Endogenous CNK2 overlaps with postsynaptic markers, and C-terminal deletion removes membrane association while preserving selected protein contacts. These data support an extrinsic scaffold associated with the PSD membrane, with no membrane-spanning segment asserted.

Summary: Accept the existing postsynapse-organization role.

Reason: In rat hippocampal neurons, endogenous CNK2 binds Vilse, and CNK2 depletion or selective loss of Vilse binding changes spine maturation. Wild-type rescue and interaction-disruption constructs connect the physical organizing work to the phenotype. Vilse supplies GAP catalysis; CNK2 localizes and combines signaling partners. This existing process is retained without adding redundant new spine-development annotations.