RAB-5 Controls the Cortical Organization and Dynamics of PAR Proteins to Maintain C. elegans Early Embryonic Polarity

Vincent Hyenne,Ramraj Velmurugan,Thierry Tremblay-Boudreault,Dinah Loerke,Jean-Claude Labbé,Barth D Grant,Sang-Chul Nam

doi:10.1371/journal.pone.0035286

Abstract

In all organisms, cell polarity is fundamental for most aspects of cell physiology. In many species and cell types, it is controlled by the evolutionarily conserved PAR-3, PAR-6 and aPKC proteins, which are asymmetrically localized at the cell cortex where they define specific domains. While PAR proteins define the antero-posterior axis of the early C. elegans embryo, the mechanism controlling their asymmetric localization is not fully understood. Here we studied the role of endocytic regulators in embryonic polarization and asymmetric division. We found that depleting the early endosome regulator RAB-5 results in polarity-related phenotypes in the early embryo. Using Total Internal Reflection Fluorescence (TIRF) microscopy, we observed that PAR-6 is localized at the cell cortex in highly dynamic puncta and depleting RAB-5 decreased PAR-6 cortical dynamics during the polarity maintenance phase. Depletion of RAB-5 also increased PAR-6 association with clathrin heavy chain (CHC-1) and this increase depended on the presence of the GTPase dynamin, an upstream regulator of endocytosis. Interestingly, further analysis indicated that loss of RAB-5 leads to a disorganization of the actin cytoskeleton and that this occurs independently of dynamin activity. Our results indicate that RAB-5 promotes C. elegans embryonic polarity in both dynamin-dependent and -independent manners, by controlling PAR-6 localization and cortical dynamics through the regulation of its association with the cell cortex and the organization of the actin cytoskeleton.

Highlights

Formation of polarized domains within a cell, through the asymmetric partition of proteins, lipids and RNAs, is required for a number of processes such as asymmetric cell division, cell migration, morphogenesis and maintenance of tissue architecture
These results indicate that depletion of RAB-5, RAB-7 and RAB-11 regulate polarity-dependent processes in the early C. elegans embryo
Our findings reveal that RAB-5 plays a significant role in polarization of the early C. elegans embryo

Summary

Introduction

Formation of polarized domains within a cell, through the asymmetric partition of proteins, lipids and RNAs, is required for a number of processes such as asymmetric cell division, cell migration, morphogenesis and maintenance of tissue architecture (reviewed in [1,2]). Most PAR proteins display a cortical and polarized localization that is crucial for their function. In the early C. elegans embryo, the anterior localization and the size of the domain occupied by the anterior PAR proteins are governed by actomyosin-dependent cortical flows, which initiate soon after fertilization and relocalize cortical components from the posterior to the anterior pole of the embryo [4]. Anterior and posterior PAR proteins mutually exclude one-another during the polarity maintenance phase, until the asymmetric division of the zygote. Some anterior PAR proteins have been shown to interact with the membrane or with the underlying actin cytoskeleton in other cell types [2], the mechanisms that promote their asymmetric enrichment and dynamically regulate their cortical anchoring in C. elegans are unclear. The genes responsible to determine the proper location and size of the domain occupied by these proteins are largely unknown

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