Uncovering the membrane mechanism of cytokinesis using live cell imaging and subcellular optogenetic tools.

Cytokinesis is the physical separation of two cells that occurs after the completion of mitosis. The mechanism underlying it is very complex. Until recently, the processes that lead to cleavage furrow formation and the extensive membrane remodelling that culminates in cytokinesis were unknown. This was due to the inexistence of methods to rapidly induce cytokinetic processes at will while simultaneously imaging membrane dynamics with high spatiotemporal precision.

 

In this webinar, we will present how we used a combination of optical techniques such as subcellular optogenetics, FRAP, TIRF, confocal imaging (Dragonfly) and SRRF-stream imaging to uncover the membrane dynamics during the final steps of cytokinesis. By optically activating the small GTPase RhoA in the middle of a cell, we were able to induce cleavage furrow formation, and continued activation resulting in elongation of the furrow to form a structure resembling the intercellular bridge. Real-time confocal imaging showed that furrow formation is mediated by actomyosin contractility, plasma membrane flow to the furrow, localized decrease in membrane tension, and increased endocytosis in the furrow. Furthermore, FRAP experiments showed exocytosis in the intercellular bridge-like structure. TIRF combined with SRRF-stream imaging revealed a second exocytosis mechanism involving incorporation of membrane lipids from vesicles into the plasma membrane at the front edge of the nascent daughter cell. These results allowed us to propose a model in which spatially separated but coordinated plasma membrane depletion and addition mediate membrane remodelling during cytokinetic processes.

 

In this webinar, the following questions will be addressed:

 

Learning Objectives: