TBC1D10A

TBC1D10A (EPI64) is a TBC-domain RabGAP that coordinates membrane trafficking, exocytosis–endocytosis coupling, and autophagy by inactivating multiple small GTPases. It functions as a GAP for Rab27A to control secretory granule exocytosis in melanocytes and parotid acinar cells, for Rab8a to regulate tubular endosome recycling via the effector JFC1, and for Rab35 to modulate Weibel-Palade body exocytosis, cilium length, and NDP52-dependent xenophagy/mitophagy (PMID:16923811, PMID:21832089, PMID:22219378, PMID:28566286, PMID:28848034, PMID:31432619). Its TBC domain additionally binds and stabilizes Arf6-GTP, linking RabGAP activity to Arf6-dependent actin remodeling and microvillar structure, while TBC1D10A also exhibits RasGAP activity at the cell periphery (PMID:17145964, PMID:23248241). In pancreatic β-cells, the Arf6 GEF ARNO recruits TBC1D10A to the plasma membrane downstream of PIP3/PI3K signaling, where it couples post-exocytic clathrin-mediated endocytosis to insulin granule secretion through coordinated Rab27A inactivation (PMID:26683831, PMID:31353211). TBC1D10A is scaffolded to apical microvilli through direct PDZ-mediated binding to EBP50 via its C-terminal DTYL motif, and this interaction is required for its proper localization and microvillar maintenance (PMID:11285285).

1. 2001 High

   Identification of TBC1D10A as a microvillar TBC/RabGAP protein anchored by direct PDZ-mediated binding to EBP50 established that RabGAP enzymes are specifically scaffolded at the apical membrane.

   Evidence Affinity chromatography from placental microvilli, direct binding assays, site-directed mutagenesis of PDZ-binding motif, immunolocalization

   PMID:11285285

   Open questions at the time

   • Target Rab substrate not identified
   • Functional consequence of microvillar localization unknown
2. 2006 High

   Demonstration that TBC1D10A is a specific Rab27A GAP and simultaneously binds Arf6-GTP to regulate microvillar structure resolved its dual enzymatic targets and linked its catalytic activity to both secretory and cytoskeletal processes.

   Evidence In vitro Rab27A-GAP assay with catalytic mutagenesis, melanosome aggregation in melanocytes, direct TBC domain–Arf6-GTP binding, Arf6-GTP loading assays, microvillar imaging

   PMID:16923811 PMID:17145964

   Open questions at the time

   • Structural basis of dual Rab27A-GAP and Arf6-binding activities unresolved
   • Whether Rab27A and Arf6 regulation are coordinated in the same cell type unknown
3. 2007 Medium

   A functional screen placed TBC1D10A in the Shiga toxin retrograde trafficking pathway (cell surface to Golgi), extending its role beyond secretory exocytosis to endosome-to-Golgi transport.

   Evidence RabGAP overexpression screen with Shiga toxin and EGF uptake assays

   PMID:17562788

   Open questions at the time

   • The specific Rab substrate inactivated by TBC1D10A in this pathway was not identified
   • Endogenous contribution not tested by loss-of-function
4. 2011 High

   Antibody inhibition and antisense knockdown in parotid acinar cells demonstrated that endogenous TBC1D10A functions as a Rab27 GAP to control regulated amylase secretion, validating its physiological role in exocrine exocytosis.

   Evidence Anti-TBC domain antibody inhibition in permeabilized cells, antisense knockdown, amylase secretion and GTP-Rab27 measurement in rat parotid cells

   PMID:21832089

   Open questions at the time

   • Whether TBC1D10A also regulates post-exocytic membrane retrieval in acinar cells not addressed
   • Redundancy with other Rab27 GAPs not tested
5. 2012 High

   Discovery of Rab8a as an additional GAP substrate and of the effector JFC1 as a direct binding partner revealed a recruitment mechanism whereby TBC1D10A engages Rab8a-GTP through its effector to drive Rab8a inactivation and Arf6-dependent membrane remodeling; separately, RasGAP activity was demonstrated in vivo, broadening TBC1D10A's catalytic scope beyond Rab GTPases.

   Evidence Rab8a-GTP pulldown, direct JFC1–EPI64 binding, GAP-dead mutagenesis epistasis for Rab8a; FRET-based Ras activity sensors, Ras-GTP pulldown, confocal FRET imaging for Ras

   PMID:22219378 PMID:23248241

   Open questions at the time

   • Whether RasGAP and RabGAP activities are independently regulated unknown
   • Structural basis for JFC1-mediated Rab8a recruitment not determined
6. 2015 High

   In β-cells, the discovery that ARNO recruits TBC1D10A to the plasma membrane downstream of glucose-induced PIP3 signaling established a molecular mechanism coupling insulin granule exocytosis to subsequent clathrin-mediated endocytosis through coordinated Arf6 and Rab27A regulation.

   Evidence Co-immunoprecipitation of ARNO–EPI64, live-cell imaging, dominant-negative/knockdown epistasis, transferrin uptake in β-cells

   PMID:26683831

   Open questions at the time

   • Whether TBC1D10A's Rab8a-GAP or Ras-GAP activities contribute in β-cells not tested
   • Upstream receptor identity not fully characterized
7. 2017 High

   Identification of Rab35 as a key TBC1D10A substrate connected its GAP activity to Weibel-Palade body exocytosis in endothelial cells and, independently, to suppression of NDP52-mediated xenophagy and mitophagy, revealing a TBC1D10A–Rab35 axis that bridges secretion and selective autophagy.

   Evidence Genome-wide RabGAP screen, co-IP, epistasis with GAP-insensitive Rab35(Q67A) for WPB exocytosis; overexpression in xenophagy/mitophagy assays, NDP52 localization, TBK1 epistasis

   PMID:28566286 PMID:28848034

   Open questions at the time

   • How TBK1 mechanistically counteracts TBC1D10A activity not resolved
   • Whether TBC1D10A is the sole Rab35 GAP in these contexts unknown
8. 2019 Medium

   Localization of TBC1D10A and Rab35 to primary cilia, with functional effects on cilium length, extended the TBC1D10A–Rab35 axis to ciliogenesis; identification of IRR as an upstream interactor in β-cells further defined the signaling cascade governing TBC1D10A membrane recruitment.

   Evidence GFP-fusion localization and siRNA/morpholino knockdown in mammalian cells and zebrafish for cilia; Co-IP of IRR–EPI64, siRNA knockdown with transferrin uptake and PIP3 measurement for β-cells

   PMID:31353211 PMID:31432619

   Open questions at the time

   • Mechanistic role of TBC1D10A in ciliary Rab35 regulation not biochemically dissected
   • IRR–EPI64 interaction based on single Co-IP without reciprocal validation
   • Whether TBC1D10A's ciliary function is conserved in mammals beyond zebrafish not established

• No structural model of TBC1D10A exists, and how a single TBC domain achieves substrate selectivity among Rab27A, Rab8a, Rab35, and Ras, and how these activities are spatiotemporally coordinated in a given cell type, remain unresolved.
• No crystal or cryo-EM structure available
• In vivo substrate hierarchy and selectivity determinants unknown
• Post-translational regulation of TBC1D10A activity not characterized