Inhibition of Insulin-induced Activation of Akt by a Kinase-deficient Mutant of the ε Isozyme of Protein Kinase C*

Abstract

Akt, also known as protein kinase B, is a protein-serine/threonine kinase that is activated by growth factors in a phosphoinositide (PI) 3-kinase-dependent manner. Although Akt mediates a variety of biological activities, the mechanisms by which its activity is regulated remain unclear. The potential role of the ε isozyme of protein kinase C (PKC) in the activation of Akt induced by insulin has now been examined. Expression of a kinase-deficient mutant of PKCε (εKD), but not that of wild-type PKCε or of kinase-deficient mutants of PKCα or PKCλ, with the use of adenovirus-mediated gene transfer inhibited the phosphorylation and activation of Akt induced by insulin in Chinese hamster ovary cells or L6 myotubes. Whereas the εKD mutant did not affect insulin stimulation of PI 3-kinase activity, the phosphorylation and activation of Akt induced by a constitutively active mutant of PI 3-kinase were inhibited by εKD, suggesting that εKD affects insulin signaling downstream of PI 3-kinase. PDK1 (3′-phosphoinositide-dependent kinase 1) is thought to participate in Akt activation. Overexpression of PDK1 with the use of an adenovirus vector induced the phosphorylation and activation of Akt; εKD inhibited, whereas wild-type PKCε had no effect on, these actions of PDK1. These results suggest that εKD inhibits the insulin-induced phosphorylation and activation of Akt by interfering with the ability of PDK1 to phosphorylate Akt.