PROTEIN-KINASE-C AND MAMMARY CELL-DIFFERENTIATION - INVOLVEMENT OF PROTEIN-KINASE-C-ALPHA IN THE INDUCTION OF BETA-CASEIN EXPRESSION

Treatment of HC11 mouse mammary epithelial cells with the lactogenic h ormones dexamethasone, insulin, and prolactin (DIP) leads to cellular differentiation and production of the milk protein beta-casein. The fo llowing experimental evidence suggests the involvement of protein kina se C (PKC) in DIP induced signal transduction. Down-regulation of PKC by 12-O-tetradecanoylphorbol-13-acetate or addition of CGP 41251, a se lective inhibitor of PKC, inhibited beta-casein protein expression ind uced by DIP in HC11 cells. This inhibition occurs at the level of tran scription, since the DIP mediated activation of a beta-casein promoter -luciferase reporter construct or of mammary gland specific factor (MC F), an essential transcription factor for beta-casein promoter activit y, was also inhibited by CCP 41251. Inhibition or down-regulation of P KC reduced the activation of MCF by prolactin as well. PKC-alpha, the only conventional PKC isoform expressed in HC11 cells, is most likely involved in the DIP induced beta-casein expression. (a) Only PKC-alpha and PKC-epsilon are down-regulated by 12-O-tetradecanoylphorbol-13-ac etate whereas PKC-delta and PKC-zeta are not. (b) Of the PKC isoforms expressed in HC11 cells, CCP 41251 inhibits PKC-alpha more potently th an PKC-delta, PKC-epsilon, and PKC-zeta. The IC50 for the inhibition o f beta-casein synthesis, MGF activation, and beta-casein promoter acti vity by CCP 41251 correlated well with the IC50 of PKC-alpha inhibitio n. (c) Finally, only PKC-alpha translocated to membrane fractions afte r DIP or prolactin treatment. Taken together, these data indicate that PKC-alpha plays an important role in the signaling pathway activated by prolactin during beta-casein induction.