Protein kinase Cµ regulation of the JNK pathway is triggered via phosphoinositide-dependent kinase 1 and protein kinase C-epsilon

The protein kinase C (PKC)-related enzyme PKCmu/PKD (protein kinase D) is activated by activation loop phosphorylation through PKCeta. Here we demonstrate that PKCmu is activated by the direct phosphorylation of PKC,E. PKCmu colocalizes with PKCepsilon in HEK293 and MCF7 cells as shown by confocal immunofluorescence analyses. PDK1, known as the upstream kinase for several PKC isozymes, associates intracellularly with PKCepsilon and PKCeta. PKCeta is phosphorylated by PDK1 in vitro, leading to kinase activation as similarly reported for PKCepsilon activation by PDK1. Coexpression of PDK1, PKCe and PKCmu in HEK293 cells results in PKCmu activation. In contrast, the coexpression of PDK1 and PKCeta with PKCmu does not activate PKCeta or consequently PKCmu. PDK1/PKCepsilon-triggered activation of PKCmu inhibits JNE a downstream effector of PKCmu, whereas upon transient expression of PDK1, PKCeta, and PKCmu, JNK is not affected. These data implicate PKCepsilon as the biologically important upstream kinase for PKCmu in HEK293 cells, regulating downstream effectors. Our results further indicate a PDK1/PKCeta/PKCmu controlled negative regulation of PKCeta kinase activity. In this study, we show that differentially activated kinase cascades involving PDK1 and novel PKC isotypes are responsible for the regulation of PKCmu activity and consequently inhibit the JNK pathway.