MICROTUBULE-ENTRAINED KINASE-ACTIVITIES ASSOCIATED WITH THE CORTICAL CYTOSKELETON DURING CYTOKINESIS

Research over the past few years has demonstrated the central role of protein phosphorylation in regulating mitosis and the cell cycle, Howe ver, little is known about how the mechanisms regulating the entry int o mitosis contribute to the positional and temporal regulation of the actomyosin-based contractile ring formed during cytokinesis, Recent st udies implicate p34(cdc2) a negative regulator of myosin II activity, suggesting a link between the mitotic cycle and cytokinesis. In an eff ort to study the relationship between protein phosphorylation and cyto kinesis, we examined the in vivo and in vitro phosphorylation of actin -associated cortical cytoskeletal (CSK) proteins in an isolated model of the sea urchin egg cortex, Examination of cortices derived from egg s or zygotes labeled with P-32-orthophosphate reveals a number of cort ex-associated phosphorylated proteins, including polypeptides of 20, 4 3 and 66 kDa, These three major phosphoproteins are also detected when isolated cortices are incubated with [P-32]ATP in vitro, suggesting t hat the kinases that phosphorylate these substrates are also specifica lly associated with the cortex, The kinase activities in vivo and in v itro are stimulated by fertilization and display cell cycle-dependent activities, Gel autophosphorylation assays, kinase assays and immunobl ot analysis reveal the presence of p34(cdc2) as well as members of the mitogen-activated protein kinase family, whose activities in the CSK peak at cell division, Nocodazole, which inhibits microtubule formatio n and thus blocks cytokinesis, significantly delays the time of peak c ortical protein phosphorylation as well as the peak in whole-cell hist one HI kinase activity, These results suggest that a key element regul ating cortical contraction during cytokinesis is the timing of protein kinase activities associated with the cortical cytoskeleton that is i n turn regulated by the mitotic apparatus.