TBC1D14 is a TBC (Tre-2/Bub2/Cdc16) domain protein of the Rab GTPase-activating protein family. It is a peripheral membrane protein found on Rab11- and transferrin-receptor-positive recycling endosomes and on the Golgi apparatus, and it shifts towards the Golgi on amino-acid starvation. It keeps the TBC catalytic arginine and glutamine, but recombinant TBC1D14 has no detectable GAP activity towards 47 Rabs, including Rab11, which it binds in the GTP-bound form. It acts instead as a Rab11 effector and adaptor: active Rab11 recruits it to recycling endosomes, and an N-terminal TRAPP-binding region (residues ~120-223) binds the TRAPPIII complex through TRAPPC8. This bridge supports TRAPP-dependent RAB1 activation and a constitutive transport step from peripheral recycling endosomes to the early Golgi that maintains the cycling pool of ATG9. Through this route TBC1D14 regulates starvation-induced autophagosome formation; its depletion increases starvation-induced LC3 lipidation, and its overexpression tubulates recycling endosomes and blocks autophagosome formation. It also binds the autophagy kinase ULK1, but its effect on ATG9 traffic is ULK1-independent.
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0005096 GTPase activator activity | IBA GO_REF:0000033 | MARK AS OVER ANNOTATED | Summary: IBA GTPase activator activity propagated from the family-root PAINT node PTN000537561 (PTHR47219), which is seeded by catalytically active TBC RabGAPs from yeast to human; TBC1D14 is not among the seeds. Reason: TBC1D14 sits inside PTHR47219 and keeps the TBC arginine finger (R472) and glutamine finger (Q508), so the root-node inference is phylogenetically reasonable. However, there is target-specific negative evidence: recombinant TBC1D14 showed no GAP activity against a panel of 47 Rabs, including its binding partner Rab11, and the Golgi/Shiga-toxin and autophagy effects of overexpression do not depend on the catalytic arginine (the R472A mutant still inhibits LC3 lipidation; the inactive point mutant blocks Shiga toxin transport). The TBC1D14-specific PAINT nodes (PTN000538677, PTN008586039), which are seeded by Q9P2M4 itself, carry location and autophagosome-assembly terms but no GAP term. Current evidence supports TBC1D14 acting as a Rab11 effector/TRAPP adaptor, not as a GAP. The data do not rule out GAP activity on an untested substrate, so this is flagged as over-annotation rather than removed. Propagation Review Root cause: PROPAGATION BAD Failure modes: FUNCTIONAL DIVERGENCE Sources checked: PANTHER:PTN000537561 SUPPORTS SOURCE BUT NOT TARGET Family-root IBD for GTPase activator activity, seeded by catalytically active TBC RabGAPs from several kingdoms. TBC1D14 descends from it but is not a seed. PANTHER:PTN000538677 NOT RELEVANT TBC1D14-clade node seeded by Q9P2M4; PAINT placed only location terms here, no GAP term. UniProtKB:Q9P2M4 SUPPORTS SOURCE BUT NOT TARGET Target. Recombinant protein lacked detectable GAP activity on 47 Rabs; catalytic residues R472/Q508 are retained, so loss is functional rather than a residue substitution. Supporting Evidence: PMID:22613832 We then performed a GAP assay screen using a panel of 47 Rabs loaded with [32P]GTP but were unable to detect GAP activity with either recombinant GST-TBC1D14 PMID:22613832 supporting the data that TBC1D14 has no GAP activity Reactome:R-HSA-8854759 TBC1D14 does not have the GAP activity and may function as Rab11 effector. PMID:17562788 However, these effects were not caused by Rab inactivation because the inactive point mutant gave a similar phenotype |
| GO:0005515 protein binding | IPI PMID:22613832 TBC1D14 regulates autophagosome formation via Rab11- and ULK... | MODIFY | Summary: IPI with ULK1 (Longatti 2012; yeast two-hybrid and co-IP mapped to the TBC domain). Reason: The interaction is real and reproducible, but bare protein binding is uninformative. ULK1 is a protein kinase, and the same paper already supports GO:0019901 protein kinase binding, which the gene carries. Proposed replacements: protein kinase binding Supporting Evidence: PMID:22613832 A yeast two-hybrid assay revealed an interaction between TBC1D14 and ULK1 PMID:22613832 In HEK293A cells, we could detect an interaction between the TBC domain of TBC1D14 and HA-ULK1 by coimmunoprecipitation |
| GO:0005515 protein binding | IPI PMID:22613832 TBC1D14 regulates autophagosome formation via Rab11- and ULK... | MODIFY | Summary: IPI with RAB11B (Longatti 2012). TBC1D14 binds activated (GTP-bound) Rab11, which recruits it to recycling endosomes. Reason: Bare protein binding is uninformative. The partner is a Rab small GTPase and the binding is to the active form, so GO:0031267 small GTPase binding is the informative MF. Binding is not accompanied by GAP activity. Proposed replacements: small GTPase binding Supporting Evidence: PMID:22613832 One of these, TBC1D14, binds ULK1 and Rab11 and disrupts recycling endosome (RE) traffic. PMID:22613832 However, the tubules formed by GFP-TBC1D14 were dependent on Rab11 PMID:22613832 TBC1D14 binds activated Rab11 but is not a |
| GO:0005515 protein binding | IPI PMID:32296183 A reference map of the human binary protein interactome. | REMOVE | Summary: IPI with NXF1 from the HuRI binary yeast two-hybrid reference map (Luck 2020). Reason: Bare protein binding is uninformative, and a single high-throughput Y2H pair with the mRNA export receptor NXF1 has no functional follow-up and no plausible link to TBC1D14 biology. Removal does not assert that the interaction is false. |
| GO:0005768 endosome | IEA GO_REF:0000117 | MODIFY | Summary: ARBA machine-learning prediction of endosome localization. Reason: Correct but general. Experimental data place TBC1D14 specifically on Rab11/transferrin-receptor-positive recycling endosomes; the gene already carries GO:0055037 recycling endosome by IDA. Proposed replacements: recycling endosome Supporting Evidence: PMID:22613832 One of these, TBC1D14, binds ULK1 and Rab11 and disrupts recycling endosome (RE) traffic. PMID:17562788 TBC1D14NDRecyclingendosomesRecyclingendosomes |
| GO:0005776 autophagosome | IBA GO_REF:0000033 | KEEP AS NON CORE | Summary: IBA from TBC1D14-clade node PTN000538677, seeded only by Q9P2M4's own IDA. Reason: The colocalization with LC3 is partial and minor; the predominant locations are recycling endosomes and, on starvation, the Golgi. TBC1D14 regulates autophagosome formation upstream (ATG9/RE membrane supply) rather than acting as a resident autophagosome component. The location is not wrong, so it is kept, but as peripheral. The IBA row merely re-expresses the same human IDA at the vertebrate node and gets the same action. Supporting Evidence: PMID:22613832 Endogenous TBC1D14 also shows partial colocalization with LC3 PMID:22613832 Amino acid starvation causes TBC1D14 to relocalize from REs to the |
| GO:0005776 autophagosome | IDA PMID:22613832 TBC1D14 regulates autophagosome formation via Rab11- and ULK... | KEEP AS NON CORE | Summary: IDA (Longatti 2012): partial colocalization of endogenous TBC1D14 with LC3 and low LC3 on a few overexpression-induced tubules. Reason: The colocalization with LC3 is partial and minor; the predominant locations are recycling endosomes and, on starvation, the Golgi. TBC1D14 regulates autophagosome formation upstream (ATG9/RE membrane supply) rather than acting as a resident autophagosome component. The location is not wrong, so it is kept, but as peripheral. The IBA row merely re-expresses the same human IDA at the vertebrate node and gets the same action. Supporting Evidence: PMID:22613832 Endogenous TBC1D14 also shows partial colocalization with LC3 PMID:22613832 Amino acid starvation causes TBC1D14 to relocalize from REs to the |
| GO:0005794 Golgi apparatus | IDA GO_REF:0000052 | ACCEPT | Summary: HPA immunofluorescence IDA, Golgi apparatus. Reason: Endogenous TBC1D14 is on the Golgi, and its Golgi pool increases on amino-acid starvation. Sub-Golgi assignments differ between sources (Golgi stack, cis-Golgi network, trans-Golgi network), so Golgi apparatus is the most precise term the combined evidence supports consistently. Supporting Evidence: PMID:26711178 We have previously demonstrated that endogenous TBC1D14 is localised to the Golgi stack file:human/TBC1D14/TBC1D14-uniprot.txt Golgi apparatus, cis-Golgi network PMID:26711178 Upon starvation and autophagy activation, TBC1D14 levels increase in the Golgi |
| GO:0005794 Golgi apparatus | IEA GO_REF:0000044 | ACCEPT | Summary: UniProt SubCell mapping to Golgi apparatus (UniProt cites cis- and trans-Golgi network from PMID:22613832/26711178). Reason: Endogenous TBC1D14 is on the Golgi, and its Golgi pool increases on amino-acid starvation. Sub-Golgi assignments differ between sources (Golgi stack, cis-Golgi network, trans-Golgi network), so Golgi apparatus is the most precise term the combined evidence supports consistently. Supporting Evidence: PMID:26711178 We have previously demonstrated that endogenous TBC1D14 is localised to the Golgi stack file:human/TBC1D14/TBC1D14-uniprot.txt Golgi apparatus, cis-Golgi network PMID:26711178 Upon starvation and autophagy activation, TBC1D14 levels increase in the Golgi |
| GO:0005829 cytosol | IEA GO_REF:0000108 | KEEP AS NON CORE | Summary: IEA inferred by inter-ontology link from the recycling endosome to Golgi transport annotation. Reason: TBC1D14 is a peripheral membrane protein recruited by Rab11-GTP; when Rab11 is depleted it becomes diffuse, consistent with a soluble cytosolic pool. The term is plausible but carries little information about function. Supporting Evidence: PMID:22613832 However, the tubules formed by GFP-TBC1D14 were dependent on Rab11 |
| GO:0010507 negative regulation of autophagy | IMP PMID:22613832 TBC1D14 regulates autophagosome formation via Rab11- and ULK... | ACCEPT | Summary: IMP (Longatti 2012): siRNA depletion of TBC1D14 increases starvation-induced LC3 lipidation and turnover. Reason: Loss-of-function with two siRNAs increases autophagic flux under starvation but not in fed cells, and overexpression inhibits autophagosome formation. This supports a negative regulatory role at the level of autophagy flux. The more mechanistic, sign-neutral autophagosome-assembly term is also carried (GO:2000785). Supporting Evidence: PMID:22613832 In cells in which TBC1D14 levels are substantially reduced, LC3 lipidation and turnover during starvation increase significantly. PMID:22613832 TBC1D14-depleted cells showed little increase in LC3 lipidation in fed conditions PMID:22613832 TBC1D14WT significantly inhibited formation, but not maturation, of yellow (mRFP-GFP) LC3-positive autophagosomes |
| GO:0019901 protein kinase binding | IPI PMID:22613832 TBC1D14 regulates autophagosome formation via Rab11- and ULK... | KEEP AS NON CORE | Summary: IPI with ULK1 (Longatti 2012). Reason: Binding to ULK1 is well supported (Y2H, co-IP, mapped to residues 224-330). Lamb 2016 showed that TBC1D14 and TRAPPIII regulate ATG9 traffic independently of ULK1, so the ULK1 interaction is real but not the core mechanism. Supporting Evidence: PMID:22613832 A yeast two-hybrid assay revealed an interaction between TBC1D14 and ULK1 PMID:22613832 In HEK293A cells, we could detect an interaction between the TBC domain of TBC1D14 and HA-ULK1 by coimmunoprecipitation PMID:26711178 These data suggest that TBC1D14 and TRAPP may act upstream of ULK1 in regulating ATG9 traffic, perhaps by maintaining normal traffic into the Golgi. |
| GO:0055037 recycling endosome | IBA GO_REF:0000033 | ACCEPT | Summary: IBA from TBC1D14-clade node PTN000538677, seeded by Q9P2M4's own experimental annotation. Reason: Recycling endosomes are a core site of TBC1D14 action: Rab11-GTP recruits it to the endocytic recycling compartment, where it in turn recruits TRAPP. Shown in two independent laboratories' datasets (Fuchs 2007; Longatti 2012; Lamb 2016). The IBA restates this at the vertebrate node. Supporting Evidence: PMID:22613832 One of these, TBC1D14, binds ULK1 and Rab11 and disrupts recycling endosome (RE) traffic. PMID:17562788 TBC1D14NDRecyclingendosomesRecyclingendosomes PMID:22613832 However, the tubules formed by GFP-TBC1D14 were dependent on Rab11 PMID:26711178 TBC1D14 in turn recruits TRAPP |
| GO:0055037 recycling endosome | IDA PMID:17562788 Specific Rab GTPase-activating proteins define the Shiga tox... | ACCEPT | Summary: IDA (Fuchs 2007): TBC1D14 localizes to recycling endosomes. Reason: Recycling endosomes are a core site of TBC1D14 action: Rab11-GTP recruits it to the endocytic recycling compartment, where it in turn recruits TRAPP. Shown in two independent laboratories' datasets (Fuchs 2007; Longatti 2012; Lamb 2016). The IBA restates this at the vertebrate node. Supporting Evidence: PMID:22613832 One of these, TBC1D14, binds ULK1 and Rab11 and disrupts recycling endosome (RE) traffic. PMID:17562788 TBC1D14NDRecyclingendosomesRecyclingendosomes PMID:22613832 However, the tubules formed by GFP-TBC1D14 were dependent on Rab11 PMID:26711178 TBC1D14 in turn recruits TRAPP |
| GO:0055037 recycling endosome | TAS Reactome:R-HSA-8854759 | ACCEPT | Summary: Reactome TAS: TBC1D14 binds RAB11 and ULK1 on recycling endosomes. Reason: Recycling endosomes are a core site of TBC1D14 action: Rab11-GTP recruits it to the endocytic recycling compartment, where it in turn recruits TRAPP. Shown in two independent laboratories' datasets (Fuchs 2007; Longatti 2012; Lamb 2016). The IBA restates this at the vertebrate node. Supporting Evidence: PMID:22613832 One of these, TBC1D14, binds ULK1 and Rab11 and disrupts recycling endosome (RE) traffic. PMID:17562788 TBC1D14NDRecyclingendosomesRecyclingendosomes PMID:22613832 However, the tubules formed by GFP-TBC1D14 were dependent on Rab11 PMID:26711178 TBC1D14 in turn recruits TRAPP |
| GO:0071955 recycling endosome to Golgi transport | IMP PMID:17562788 Specific Rab GTPase-activating proteins define the Shiga tox... | ACCEPT | Summary: IMP (Fuchs 2007): overexpressed TBC1D14 blocks Shiga toxin transport to the Golgi without affecting EGF uptake. Reason: The original evidence is an overexpression phenotype independent of the catalytic residue, which on its own would be weak. Lamb 2016 independently established the mechanism: TBC1D14 bridges Rab11-positive recycling endosomes to TRAPPIII, its depletion reduces RAB1 activation, and TBC1D14/TRAPPIII regulate a constitutive recycling-endosome-to-early-Golgi step that maintains the cycling pool of ATG9. This is the best-supported process for TBC1D14. Supporting Evidence: PMID:17562788 In the case of TBC1D14, although the Golgi was fragmented, Shiga toxin did not localize to these Golgi fragments, indicating there was a block in transport. PMID:17562788 However, these effects were not caused by Rab inactivation because the inactive point mutant gave a similar phenotype PMID:26711178 These data support a model where TBC1D14 acts as a bridge between a TRAPP complex and activated RAB11. PMID:26711178 However, TBC1D14 depletion resulted in a significant reduction in RAB1B bound to GFP PMID:26711178 These data suggest that TBC1D14 and TRAPP may act upstream of ULK1 in regulating ATG9 traffic, perhaps by maintaining normal traffic into the Golgi. |
| GO:1902017 regulation of cilium assembly | IMP NOT PMID:17646400 Functional dissection of Rab GTPases involved in primary cil... | ACCEPT | Summary: NOT IMP: in the Yoshimura 2007 overexpression screen of TBC RabGAPs, TBC1D14 did not affect primary cilium formation. Reason: TBC1D14 is not named in the cached main text; the per-GAP results are in the screen figure/supplement, which the curator read. The main text states that GAPs other than the hits had no effect on primary cilia, and nothing else links TBC1D14 to cilia. The negative annotation is accepted, deferring to the curator. Supporting Evidence: PMID:17646400 Other GAPs either had no effect on primary cilia |
| GO:2000785 regulation of autophagosome assembly | IBA GO_REF:0000033 | ACCEPT | Summary: IBA from TBC1D14-clade node PTN008586039 (Euteleostomi), seeded by Q9P2M4's own IMP. Reason: TBC1D14 regulates autophagosome formation by controlling membrane and ATG9 supply from recycling endosomes. The sign-neutral regulation term is appropriate: depletion increases starvation-induced autophagy (Longatti 2012), while the Lamb 2016 model places TBC1D14/TRAPPIII in maintaining the ATG9 cycling pool needed for initiation. PAINT placed this term on a TBC1D14-specific node, which supports transfer to TBC1D14 orthologs; the cod tbc1d14 models fall in this clade. Supporting Evidence: PMID:22613832 TBC1D14WT significantly inhibited formation, but not maturation, of yellow (mRFP-GFP) LC3-positive autophagosomes PMID:22613832 In cells in which TBC1D14 levels are substantially reduced, LC3 lipidation and turnover during starvation increase significantly. PMID:26711178 These data suggest that TBC1D14 and TRAPP may act upstream of ULK1 in regulating ATG9 traffic, perhaps by maintaining normal traffic into the Golgi. |
| GO:2000785 regulation of autophagosome assembly | IMP PMID:22613832 TBC1D14 regulates autophagosome formation via Rab11- and ULK... | ACCEPT | Summary: IMP (Longatti 2012): TBC1D14 overexpression inhibits formation, not maturation, of autophagosomes; depletion increases starvation-induced LC3 lipidation. Reason: TBC1D14 regulates autophagosome formation by controlling membrane and ATG9 supply from recycling endosomes. The sign-neutral regulation term is appropriate: depletion increases starvation-induced autophagy (Longatti 2012), while the Lamb 2016 model places TBC1D14/TRAPPIII in maintaining the ATG9 cycling pool needed for initiation. PAINT placed this term on a TBC1D14-specific node, which supports transfer to TBC1D14 orthologs; the cod tbc1d14 models fall in this clade. Supporting Evidence: PMID:22613832 TBC1D14WT significantly inhibited formation, but not maturation, of yellow (mRFP-GFP) LC3-positive autophagosomes PMID:22613832 In cells in which TBC1D14 levels are substantially reduced, LC3 lipidation and turnover during starvation increase significantly. PMID:26711178 These data suggest that TBC1D14 and TRAPP may act upstream of ULK1 in regulating ATG9 traffic, perhaps by maintaining normal traffic into the Golgi. |
| GO:0060090 molecular adaptor activity | IDA PMID:26711178 TBC1D14 regulates autophagy via the TRAPP complex and ATG9 t... | NEW | Summary: Proposed: TBC1D14 bridges GTP-bound Rab11 on recycling endosomes to the TRAPPIII complex (via TRAPPC8). Reason: Lamb 2016 showed binding of TBC1D14 to TRAPP through its N-terminal TRAPP-binding region and TRAPPC8, recruitment of TRAPP and RAB1 onto Rab11-positive tubules generated by TBC1D14, and reduced RAB1 activation on TBC1D14 depletion. TBC1D14 itself does the bridging work, so this is participation, not mere necessity. No more specific GO term for TRAPP complex binding exists. Supporting Evidence: PMID:26711178 These data support a model where TBC1D14 acts as a bridge between a TRAPP complex and activated RAB11. PMID:26711178 TRAPPC8 is required for TBC1D14 to bind the TRAPP complex PMID:26711178 However, TBC1D14 depletion resulted in a significant reduction in RAB1B bound to GFP |
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Download this section (compressed HTML)Q: TBC1D14 retains the TBC arginine and glutamine fingers yet shows no GAP activity on 47 Rabs. Is it a GAP for an untested Rab, or has its TBC domain been co-opted as a Rab11-binding effector module?
Q: Should the family-root GTPase activator activity IBA (PTN000537561) be blocked at the TBC1D14 clade node, given the human negative biochemical data?
Q: Is 'molecular adaptor activity' the best MF for the Rab11-TRAPPIII bridge, or should a GO term for TRAPP complex binding be created?
Experiment: Purify TBC1D14 TBC domain and test GAP activity on Rab1, Rab11 and the remaining Rabs with intrinsic hydrolysis controls; compare with an active TBC GAP carrying the same R/Q residues.
Type: in vitro biochemistry
Experiment: Reconstitute Rab11-GTP-dependent recruitment of TRAPPIII to liposomes with and without TBC1D14 or its TRAPP-binding region, and measure RAB1 GEF activity.
Type: in vitro reconstitution
Experiment: Knock out TBC1D14 in a zebrafish or cell model of a teleost and test ATG9 distribution and starvation-induced autophagy, to check conservation of the human mechanism for transfer to fish orthologs.
Type: genetic loss of function
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