Characterizing the role of endocytic recycling in the maintenance of polarity in the Caenorhabditis elegans embryo

In biology, the asymmetric localization of molecules or organelles within a single cell is crucial for the specification of cell types found in multicellular organisms. The mechanisms that polarize a cell are widely conserved. The PAR polarity complex and the small GTPase Cdc42 are regulate cellular polarity in embryos, neuronal cells and epithelial cells. A balance between endocytosis, recycling and lateral diffusion has been described as a key mechanism that stabilizes polarized protein localization. However, it remains poorly understood how exactly endocytosis and recycling could achieve this function in higher eukaryotic cells.;In this dissertation, I have characterized an endocytic mechanism that participates in the maintenance of PAR asymmetry in the C. elegans embryo. C. elegans dynamin, DYN-1, localizes to cortical puncta that become enriched at the anterior half of the embryo, where endocytosis occurs primarily during maintenance phase. Depletion of DYN-1 significantly reduces the enrichment of endocytosis , and destabilization PAR and CDC-42 asymmetry during polarity maintenance phase is observed. I found that the anterior polarity cue, PAR-6, localizes to cytoplasmic puncta that emerge from the cortex into the cell from dynamin-rich sites. Taken together, this work suggests that dynamin-dependent endocytosis of polarity cues plays a role in stabilizing polarity during maintenance phase.;In the subsequent study, I found that the Arp2/3 complex achieves both of these roles. Depletion of the ARX-2 specifically disrupts maintenance of actin and DYN-1 foci. Localizations of polarity cues are destabilized in a manner similar to that observed in dyn-1 RNAi-treated embryos. I do not observe disruptions in endocytosis, but do observe significantly larger early endosomes and accumulations of cytoplasmic PAR-6 in association with these endosomes. I suggest that I have uncovered an actin-dependent mechanism for the rapid recycling of internalized anterior polarity cues, and this contributes to the stable localization at the cortex.;Together, my work has characterized a novel mechanism in which dynamin participates in the internalization of polarity cues, and Arp2/3-mediated actin dynamics appears to participate in the regulation of their sorting and recycling at the early endosome. Efficient function of these events contributes to the stabilization of cortical asymmetry.