Signalling Pathways Regulating the Dephosphorylation of Ser729in the Hydrophobic Domain of Protein Kinase Cε upon Cell Passage*

We have recently demonstrated that in quiescent fibroblasts protein kinase C (PKC) ε95is phosphorylated at Ser729, Ser703, and Thr566and that upon passage of quiescent cells phosphorylation at Ser729is lost, giving rise to PKCε87. Ser729may be rephosphorylated later, suggesting cycling between PKCε87and PKCε95. Here we show that the dephosphorylation at Ser729is insensitive to okadaic acid, calyculin, ascomycin C, and cyclosporin A, suggesting that dephosphorylation at this site is not mediated through protein phosphatases 1, 2A or 2B. We demonstrate that this dephosphorylation at Ser729requires serum and cell readhesion and is sensitive to rapamycin, PD98059, chelerythrine, and Ro-31–8220. These results suggest that the phosphorylation status of Ser729in the hydrophobic domain at Ser729is regulated independently of the phosphorylation status of other sites in PKCε, by a mTOR-sensitive phosphatase. The mitogen-activated protein kinase pathway and PKC are also implicated in regulating the dephosphorylation at Ser729.