Supplementary Figure 5: Cdk1 inhibition in the presence of mitotic spindle triggers anisotropic localization of cortical F-actin and myosin II.

(a) Confocal images of H2B-mCherry (red) and tubulin-GFP (green) expressing mitotic HeLa cells. The cells were either untreated or treated with the indicated inhibitor to induce mitotic arrest. Protease inhibitor, MG132, treated cells exhibited bipolar morphology similar to untreated cells. STC, an Eg5 inhibitor, treated cells had mono-polar spindle morphology. Microtubule depolymerizer, nocodazole (Noc), treated cells were devoid of mitotic spindle. Scale bar of 10 μm applies to all images. (b) Confocal images of MYH9-GFP (green) and Lifeact-mCherry (red) expressing mitotic HeLa cells. Images were acquired every 4 minutes. Cells were untreated, or preincubated for 30 min with MG132, STC or nocodazole (Noc) at concentrations indicated. Nine minutes after starting the experiment 3 μM Cdk1/2 inhibitor III was introduced. Scale bar of 10 μm applies to all images. (c) Scheme of the mitotic spindle organizing the cortical enrichment of actomyosin when Cdk1 is inhibited. In Cdk1 inhibited cells with a bi (untreated or MG132 arrest) or mono (STC arrest) polar spindle, cortical actomyosin is enriched in regions furthest from the spindle poles. Mitotic cells with no spindle (nocodazole arrest) were devoid of cortical myosin II enrichment upon Cdk1 inhibition.