Centrosome-induced membrane infolding linked to Rac pathway and Arp2/3 network recruitment during actin cap formation in the Drosophila embryo
Authors: Rebecca Tam; Nicolas Vergara Ruiz; Tony Harris
Affiliation: University of Toronto
Keywords: a. cytoskeleton; q. cellular remodeling
The cell cortex is a dynamic network of proteins that allows cells to change shape during processes like migration and division. Local activation of the Rac-GEF pathway leads to the assembly of Arp2/3 networks which change the cell cortex. Moreover, centrosomes influence the cortex during cell migration and division. How centrosomes activate the assembly of actin networks is not well understood. During early Drosophila embryogenesis, an actin cap forms above the nucleus and the growth of the cap creates a dome-like structure that houses the mitotic apparatus. The actin cap forms below the plasma membrane and requires centrosomes and the Rac-GEF, Sponge. What remains unclear is how centrosomes promote the local recruitment of the Rac-GEF pathway. My current research shows that the plasma membrane forms apical folds and sub-apical tubules above the centrosomes. Moreover, Spg, Rac-GTP, Scar, and Arp2/3 localized to these membrane structures above the centrosomes where nascent actin caps form. To closely investigate the relationship between centrosomes, the plasma membrane, and the Rac-GEF pathway, we inhibited cap expansion by depleting Arp3 using RNAi. Membrane folds accumulated excessively above expected centrosomes and sub-apical membrane tubules persisted. In addition, the Rac-GEF pathway proteins showed higher levels of buildup above expected centrosomes compared to the control. Notably, the plasma membrane and the Rac-GEF pathway proteins had a more focused relationship with the centrosomes in Arp3 RNAi, suggesting Arp2/3 network growth normally spreads the components away from the sites of initial cap induction. To see how the centrosomes influence the plasma membrane and the Rac-GEF pathway localization, centrosomin (Cnn) RNAi was used. In Cnn RNAi, the membrane was disorganized, with membrane folds lacking specific association with the centrosomes and sub-apical tubules being less numerous. Furthermore, the accumulation of the Rac-GEF pathway was diminished in Cnn RNAi. Together, these results suggest that the centrosome promotes the localization of the Rac-GEF pathway through a mechanism linked to local plasma membrane infolding.