Optogenetic analysis of RhoA-mediated furrow formation during cytokinesis

Cytokinesis is the final stage of cell division in which an actomyosin-based contractile ring physically divides the cell into two genetically equivalent daughter cells. The cytokinetic cleavage furrow is controlled by the position of the anaphase spindle and this positioning is critical to ensure that cells divide properly to generate two genetically equivalent daughter cells. The anaphase spindle performs this function, in part by controlling the activity of the small GTPase RhoA, which is required for actomyosin-based contractile ring assembly and furrow ingression during cytokinesis. However, it remains unclear whether the spindle makes additional contributions to furrow assembly or whether a local zone of RhoA activity is sufficient to give rise to furrow formation. Likewise it is not known whether the entire cortex equally responsive to RhoA and whether the spindle modulates the response to active RhoA? Furthermore is contractile ring assembly subject to cell cycle regulation? Here, we have addressed these and other questions by using optogenetics to gain tight spatial and temporal control of RhoA activity in mammalian cells. Light-mediated recruitment of a RhoGEF domain to the plasma membrane leads to rapid induction of RhoA activity, as indicated by accumulation of a RhoA biosensor, myosin II, and F-actin. Light-mediated induction of RhoA activity, leads to assembly of cytokinetic furrows that partially ingress. Furthermore, furrow formation in response to RhoA activation is not temporally or spatially restricted. These results indicate RhoA activation is sufficient to induce assembly of functional contractile rings and furrow formation.