TBC1D9B acts as a Rab11a-GTPase-Activating Protein

GALLO LI; YONG L; YIN XM; APODACA G

Boston

Conferencia; Gordon Conference on Lysosomes and Endocytosis; 2014

GRC Organization

Rab small GTPases regulate critical steps in membrane trafficking by cycling between an inactive GDP-bound state and an active GTP-bound state. The Rab GTPase cycle is regulated by activating enzymes or guanine nucleotide-exchange factors (GEFs), and inactivating enzymes or GTPase-activating proteins (GAPs). Thus the identification of GEFs and GAPs is crucial to understand the spatio-temporal regulation of Rab-mediated membrane trafficking events. Rab11a is a key modulator of vesicular trafficking processes, but there is limited information about the GEFs and GAPs that regulate its GTP-GDP cycle. Here we identified the TBC family member TBC1D9B as a GAP for Rab11a. We observed that TBC1D9B stimulated Rab11a GTPase activity in vitro, but not that of several other Rabs including Rab11b and Rab25. In polarized MDCK cells, TBC1D9B colocalized with Rab11a and overexpression of TBC1D9B, but not an inactive mutant, decreasedthe rate of basolateral-to-apical IgA transcytosis, a Rab11a-dependent pathway, while shRNA-mediated depletion of TBC1D9B increased the rate of this process. Interestingly, use of GST-pull-down assays shows that overexpressed TBC1D9B disrupts the interaction between Rab11a and its effector Sec15A. When a catalytic inactive mutant of TBC1D9Bis overexpressed, Rab11a interaction with Sec15A is not affected. We therefore conclude that expression of TBC1D9B decreases the amount of active Rab11a in the cell, acting as a Rab11a-GAP and regulating the Rab11a-dependent basolateral-to-apical transcytotic pathway.