Determining the role of individual ESCRT-III proteins in cytokinesis abscission

The ESCRT machinery participates in membrane constriction and fission in a variety of processes in cells. Cytosolic ESCRT-III proteins and the AAA ATPase VPS4, the driving force for membrane fission, assemble into cortical filaments to induce membrane constriction and severing. One of the ESCRT mediated processes is abscission of the intercellular bridge at the end of cytokinesis, which constitutes the last step of cell division.

Over 10 different ESCRT-III proteins were identified in the mammalian genome, but the specific role of ESCRT-III subunits in constriction and fission have not been defined. In vitro studies suggest that the ESCRT-III components CHMP2A was designated as one of the components required for the final fission event. To test the effect of CHMP2A on ESCRT mediated membrane remodeling in cells, we generated CHMP2A knock-out HeLa cells. Live-cell imaging experiments revealed that abscission is severely delayed in the absence of CHMP2A. Surprisingly, in contrast to other essential abscission components, all the cells eventually completed abscission, suggesting that CHMP2A is not essential for the final fission event. SIM imaging of the core ESCRT-III components CHMP4B and IST1 revealed defected in the spatiotemporal organization of the filament during the process. Remarkably, labeling both proteins in the same cell revealed that while these proteins co-localize in normal cells, their organization pattern is spatially separated in CHMP2A KO cells, suggesting that the polymerization process itself is affected by CHMP2A depletion. Together, these results suggest a role for CHMP2A in filament polymerization rather in the final fission process itself.