The protein kinase inhibitor 6-dimethylaminopurine does not inhibit micronuclear mitosis, but impairs the rearrangement of cytoplasmic MTOCs and execution of cytokinesis in the ciliate Paramecium during transition to interphase

Abstract

Summary Divisional morphogenesis in Paramecium involves sequential activation of several distinct intra- and extranuclear MTOCs, duplication of microtubular organelles, and extensive reorganization of other cytoskeletal structures. This system is therefore useful in the investigation of the role of putative protein kinases at several stages of these interdependent processes. The experiments reported here were performed by treating cells prior to division with 6-DMAP, an inhibitor of activity of some protein kinases, with nocodazole, an inhibitor of microtubule assembly, and by microinjection of the monoclonal antibody MPM-2, specific for phosphoproteins. Simultaneously, the distribution of MTOCs was monitored using the CTR 210 anti-centrosomal antibody, and the phosphorylation state of cytoskeletal organelles was examined using the MPM-2 and MPM-3 antibodies, directed against mitotic-phase phosphorylated epitopes. We show that 6-DMAP and microinjection of MPM-2 antibody do not inhibit completion of micronuclear mitosis or duplication of oral and basal body MTOCs, and have only minor effects on microtubule assembly in dividing Paramecium . However, 6-DMAP does impair execution of cytokinesis and interferes with two classes of MTOCs: the oral filamentous network, which becomes disorganized, and the cortical MTOCs which become impaired in their interactions with various cytoskeletal elements (epiplasm, parasomal sacs, kinetodesmal fibers, infraciliary lattice) during transition to interphase. We assume that activation of some 6-DMAP sensitive protein kinases coordinates cytokinesis and final processing of the interphase cytoskeleton in Paramecium .