The GTPase Rab5 and phosphatidylinositol-3 phosphate [PI(3)P] coordinately regulate endosome trafficking. Rab5 recruits Vps34, the class III phosphoinositide 3-kinase (PI3K), to generate PI(3)P and recruit PI(3)P-binding proteins. Loss of Rab5 and loss of Vps34 have opposite effects on endosome size, suggesting that our understanding of how Rab5 and PI(3)P cooperate is incomplete. Here, we report a novel regulatory loop whereby Caenorhabditis elegans VPS-34 inactivates RAB-5 via recruitment of the TBC-2 Rab GTPase-activating protein. We found that loss of VPS-34 caused a phenotype with large late endosomes, as with loss of TBC-2, and that Rab5 activity (mice have two Rab5 isoforms, Rab5a and Rab5b) is increased in Vps34-knockout mouse embryonic fibroblasts (Vps34 is also known as PIK3C3 in mammals). We found that VPS-34 is required for TBC-2 endosome localization and that the pleckstrin homology (PH) domain of TBC-2 bound PI(3)P. Deletion of the PH domain enhanced TBC-2 localization to endosomes in a VPS-34-dependent manner. Thus, PI(3)P binding of the PH domain might be permissive for another PI(3)P-regulated interaction that recruits TBC-2 to endosomes. Therefore, VPS-34 recruits TBC-2 to endosomes to inactivate RAB-5 to ensure the directionality of endosome maturation.
Rab5 and Rab7 GTPases are key regulators of endosome maturation and lysosome fusion. They activate the class III phosphoinositide 3-kinase (PI3K) Vps34 to generate pools of phosphatidylinositol-3 phosphate [PI(3)P] on endosomes. Together PI(3)P and the GTP-bound Rabs coordinate the recruitment of endosomal regulators to drive early to late endosome maturation and ultimately lysosome fusion. Counterintuitively, loss of Vps34 results in enlarged endosomes, like those seen from expressing activated Rab GTPases. Two recent papers in the Journal of Cell Science, Jaber et al., 2016 and Law, Seo et al., 2017, demonstrate that a function of Vps34 is to inactive the Rab5 and Rab7 GTPases via recruitment of the TBC1D2 family of Rab GTPase Activating Proteins (GAPs).
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