DYNAMIC CHANGES IN NUCLEAR ARCHITECTURE DURING MITOSIS - ON THE ROLE OF PROTEIN-PHOSPHORYLATION IN SPINDLE ASSEMBLY AND CHROMOSOME SEGREGATION

During mitosis, the vertebrate cell nucleus undergoes profound changes in architecture. At the onset of mitosis, the nuclear envelope breaks down, the nuclear lamina is depolymerized, and interphase chromatin i s condensed to chromosomes. Concomitantly, cytoplasmic microtubules ar e reorganized into a mitotic spindle apparatus, a highly dynamic struc ture required for the segregation of sister chromatids. Many of the ab ove events are controlled by reversible phosphorylation. Hence, our la boratory is interested in characterizing the kinases involved in promo ting progression through mitosis and in identifying their relevant sub strates. Prominent among the kinases responsible for regulating entry into mitosis is the Cdc2 kinase, the first member of the cyclin depend ent kinase (Cdk) family. Recently, we found that Cdc2 phosphorylates H sEg5, a human kinesin-related motor protein associated with centrosome s and the spindle apparatus. Our results indicate that phosphorylation regulates the association of HsEg5 with the mitotic spindle and that the function of this plus-end directed motor is essential for centroso me separation and bipolar spindle formation. Another kinase implicated in regulating progression through mitosis is Plk1 (polo-like kinase 1 ), the human homologue of the Drosophila gene product ''polo.'' By ant ibody microinjection we have found that Plk1 is required for the funct ional maturation of centrosomes and hence for entry into mitosis. Furt hermore, we found that microinjected anti-Plk1 antibodies caused a mor e severe block to cell cycle progression in diploid fibroblasts than i n immortalized tumor cells. This observation hints at the existence of a checkpoint linking Cdc2 activation to the presence of functional ce ntrosomes. (C) 1996 Academic Press, Inc.