Immunofluorescent localization of cyclic nucleotide-dependent protein kinases on the mitotic apparatus of cultured cells.

Abstract

Cyclic nucleotides and cyclic nucleotide-dependent protein kinases were implicated in the regulation of cell motility and division, processes that depend on the cell cytoskeleton. To determine whether cyclic nucleotides or their kinases are physically associated with the cytoskeleton during cell division, fluorescently labeled antibodies directed against cAMP, cGMP and the cyclic nucleotide-dependent protein kinases were used to localize these molecules in mitotic PtK1 cells. The cGMP-dependent protein kinase and the type II regulatory subunit of the cAMP-dependent protein kinase were localized on the mitotic spindle. Throughout mitosis, their distribution closely resembled that of tubulin. Antibodies to cAMP, cGMP and the type I regulatory and catalytic subunits of the cAMP-dependent protein kinase did not label the mitotic apparatus. The association between specific components of the cyclic nucleotide system and the mitotic spindle suggests that cyclic nucleotide-dependent phosphorylation of spindle proteins, such as those of microtubules, may play a fundamental role in the regulation of spindle assembly and chromosome motion.