CED-10/Rac1 Regulates Endocytic Recycling through the RAB-5 GAP TBC-2

Lin Sun,Zheng Zhou,Farhad Karbassi,Ou Liu,Jigar Desai,Marc-André Sylvain,Barth D Grant,Anbing Shi,Christian E Rocheleau,Julie G Donaldson

doi:10.1371/journal.pgen.1002785

Abstract

Rac1 is a founding member of the Rho-GTPase family and a key regulator of membrane remodeling. In the context of apoptotic cell corpse engulfment, CED-10/Rac1 acts with its bipartite guanine nucleotide exchange factor, CED-5/Dock180-CED-12/ELMO, in an evolutionarily conserved pathway to promote phagocytosis. Here we show that in the context of the Caenorhabditis elegans intestinal epithelium CED-10/Rac1, CED-5/Dock180, and CED-12/ELMO promote basolateral recycling. Furthermore, we show that CED-10 binds to the RAB-5 GTPase activating protein TBC-2, that CED-10 contributes to recruitment of TBC-2 to endosomes, and that recycling cargo is trapped in recycling endosomes in ced-12, ced-10, and tbc-2 mutants. Expression of GTPase defective RAB-5(Q78L) also traps recycling cargo. Our results indicate that down-regulation of early endosome regulator RAB-5/Rab5 by a CED-5, CED-12, CED-10, TBC-2 cascade is an important step in the transport of cargo through the basolateral recycling endosome for delivery to the plasma membrane.

Highlights

The C. elegans intestine has proven to be a powerful model system for the study of epithelial cell membrane trafficking mechanisms
Our work shows that RAB-5 deactivation is important for cargo recycling, and it provides some of the first mechanistic insight into how changes in Rabs can be controlled during endocytic recycling
Loss of CED-10/Rac1, CED-12/Elmo, or CED-5/Dock180 leads to intracellular accumulation of recycling cargo

Summary

Introduction

The C. elegans intestine has proven to be a powerful model system for the study of epithelial cell membrane trafficking mechanisms. The worm intestine is a simple epithelial tube consisting of 20 enterocyte cells that form nine ‘‘donut-like’’ intestinal rings [1]. Each of these 20 cells is terminally differentiated, and each intestinal cell is maintained for the life of the animal without replacement [1]. Like mammalian intestinal epithelial cells, C. elegans enterocytes display apicobasal polarity with defined apical junctions separating the apical and basolateral domains [1]. The apical enterocyte membranes, which form the intestinal lumen, display a prominent microvillar brush border, with an overlying glycocalyx and underlying subapical terminal web rich in actin and intermediate filaments [1]. The basolateral membrane is in contact with the pseudocoelom (body cavity) and is responsible for the exchange of molecules between the intestine and other tissues of the body

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