Regulation of the activity of protein kinases depending on their functional domains

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RAC-protein kinase (RAC-PK) and protein kinase C (PKC) have a catalytic domain highly homologous each other in the carboxyl-terminal regions, and the regulatory domain at their amino-terminal regions. RAC-PK and PKC have a PH domain and a Cl region, respectively, in the regulatory domain, and PKC has been shown to associate with PKC through the PH domain. We examined the regulation of these protein kinases through the regulatory domain in this study. In the initial study, the screening of the cDNA library was carried out and a novel clone of RAC-PK gamma was isolated in addition to the previously known RAC-PK alpha and beta. The PH domain of three types of RAC-PK associated with PKC subspecies in vitro. Analysis using deletion mutants revealed that the specific region for the binding with the PH domain was hardly identified in the PKC molecule, suggesting that the whole structure of PKC is important for the interaction with the PH domain. During the analysis of the measurement of RAC-PK activity expressed in cultured cell lines, it was revealed that RAC-PK is activated by cellular stress such as heat shock and hyperosmotic treatment. On the other hand, RAC-PK has been reported to be a downstream target of phosphatidylinositol 3-kinase, that is activated by the growth factor-stimulation. Activation of RAC-PK by cellular stress was, however, not suppressed by wortmannin, a potent inhibitor of phosphatidylinositol 3-kinase. It is plausible that RAC-PK is activated through alternative pathways. Furthermore, the association of RAC-PK and PKC was enhanced by stress treatment. The mutual interaction between RAC-PK and PKC,probably through the protein-protein interaction, may have an important role in the intracellular signal transduction.