The final step in cell division is cytokinesis, where the cytoplasm divides into two after nuclear division. Cytokinesis involves multiple steps that are spatiotemporally organized for successful cell separation. During cytokinesis, the cell ensures that the actomyosin ring constricts only after complete chromosome segregation to help maintain genomic integrity in the progeny. In fission yeast, ring constriction is accompanied by septum ingression. Septum formation involves synthesis of a primary septum, which is then flanked by the secondary septum. Finally, the cells separate when digestive enzymes are delivered to break down the primary septum. The secondary septum that remains form the cell wall for the new ends generated as a result of cell division. These multiple steps need to occur in the right spatiotemporal order to ensure successful cell division and prevent cell death. How does the cell organize multi-step cytokinetic processes to successfully separate cells after division? Our lab has discovered a novel spatiotemporal activation pattern of the small GTPase Cdc42 that promotes distinct events during cytokinesis (Wei et al., 2016). While Cdc42 was implicated in cytokinesis, its precise role in cytokinesis was unclear, likely due to the complexity of its regulation. Our data indicate that the spatiotemporal activation pattern of Cdc42 at the division site helps to organize different cytokinetic events. Current projects in the lab are focused on the following questions.
a. How is Cdc42 spatiotemporal activation pattern established at the division site?
We have reported that during cytokinesis the Cdc42 activator Gef1 localizes to the actomyosin ring and activates Cdc42 to promote timely onset of ring constriction and septum ingression (Wei et al., 2016). The activator Scd1 localizes to the membrane barrier behind the ring and facilitates proper septum formation. Scd1 promotes primary septum formation while restricting excessive secondary septum formation (Wei et al., 2016). Finally, Cdc42 is inactivated at the division site, promoting successful cell abscission. Thus a gradient of active Cdc42 is established at the division site (Fig.1) and we posit that this gradient organizes different cytokinetic events. We are currently investigating how the different regulators of Cdc42 establish its spatiotemporal activation pattern over time at the division site.
b. How does the Cdc42 activation pattern organize cytokinetic events?
Studies show that Cdc42 and Rho GTPases are antagonistic to each other. The physiological significance for this crosstalk in fission yeast is not clear. In fission yeast, the RHO GTPases Rho1-Rho5 are required for different cytokinetic events. Our data suggest that Cdc42 engages in crosstalk with other Rho GTPases to organize different cytokinetic events (Fig.2). We are investigating how crosstalk between Cdc42 and Rho GTPases ensures temporal organization of cytokinetic events.
c. How does the actomyosin ring act as a landmark to coordinate membrane furrowing and septum ingression?
Reports indicate that in fission yeast, actomyosin ring constriction does not generate sufficient force to overcome the internal turgor pressure for furrow formation. Instead, septum ingression concurrent with constriction provides the force necessary to overcome turgor pressure. While the actomyosin ring is necessary, it is not sufficient for furrow formation in fission yeast. This suggests that the actomyosin ring has a distinct role in cytokinesis in addition to providing a mechanical structure. We posit that the actomyosin ring acts as a landmark for the recruitment of septum-synthesizing enzymes, thus ensuring that ring constriction is spatiotemporally concurrent with septum ingression. Our data indicate that the Cdc42 activator Gef1 localizes to the actomyosin ring and promotes even recruitment of proteins along along the ring. This ensures concentric septum ingression and furrow formation (Fig.3). We are investigating the molecular details of how proteins are recruited evenly along the ring and the role of Gef1 in this process.