A spatiotemporally coordinated cascade of protein kinase C activation controls isoform-selective translocation

In pituitary GH3B6 cells, signaling involving the protein kinase C (PKC) multigene family can self-organize into a spatiotemporally coordinated cascade of isoform activation. Indeed, thyrotropin-releasing hormone (TRH) receptor activation sequentially activated green fluorescent protein (GFP)-tagged or endogenous PKC beta 1, PKC alpha, PKC epsilon, and PKC delta, resulting in their accumulation at the entire plasma membrane (PKC beta and -delta) or selectively at the cell-cell contacts (PKC alpha and -epsilon). The duration of activation ranged from 20 s for PKC alpha to 20 min for PKC epsilon. PKC epsilon and -E selective localization was lost in the presence of Go6976, suggesting that accumulation at cell-cell contacts is dependent on the activity of a conventional PKC. Constitutively active, dominant-negative PKCs and small interfering RNAs showed that PKC alpha localization is controlled by PKC beta 1 activity and is calcium independent, while PKC epsilon localization is dependent on PKCa activity. PKC delta was independent of the cascade linking PKC beta 1, -alpha, and -epsilon. Furthermore, PKCa., but not PKC epsilon, is involved in the TRH-induced beta-catenin relocation at cell-cell contacts, suggesting that PKC epsilon is not the unique functional effector of the cascade. Thus, TRH receptor activation results in PKC beta 1 activation, which in turn initiates a calcium-independent but PKC beta 1 activity-dependent sequential translocation of PKC(x and -E. These results challenge the current understanding of PKC signaling and raise the question of a functional dependence between isoforms.