Possible role of Ca2+-independent protein kinase C isozyme, nPKCɛ, in thyrotropin- releasing hormone-stimulated signal transduction: Differential down-regulation of nPKCɛ in GH4C1 cells

Abstract Protein kinase C (PKC) molecular species of GH 4 C 1 cells were analyzed after separation by hydroxyapatite column chromatography. A novel Ca 2+ -independent PKC, nPKC ɛ , was identified together with two conventional Ca 2+ -dependent PKCs, PKC α and β II by analysis of kinase and phorbol ester-binding activities, immunoblotting using isozyme-specific antibodies, and Northern blotting. These PKCs are down-regulated differently when cells are stimulated by outer stimuli; phorbol esters deplete PKC β II and nPKC ɛ from the cells more rapidly than PKC α , whereas thyrotropin-releasing hormone (TRH) at 200 nM depletes nPKC ɛ exclusively with a time course similar to that induced by phorbol esters. However, translocation of PKC α and β II to the membranes is elicited by both TRH and phorbol esters. These results suggest that TRH and phorbol ester activate PKC α and β II differently but that nPKC ɛ is stimulated similarly by both stimuli. Thus, in GH 4 C 1 cells, Ca 2+ -independent nPKC ɛ may play a crucial role distinct from that mediated by Ca 2+ -dependent PKC α and β II in a cellular response elicited by both TRH and phorbol esters.