PHOSPHOINOSITIDE-3-OH KINASE-DEPENDENT REGULATION OF GLYCOGEN-SYNTHASE KINASE-3 AND PROTEIN-KINASE B AKT BY THE INTEGRIN-LINKED KINASE/

Integrin-linked kinase (ILK) is an ankyrin-repeat containing serine-th reonine protein kinase capable of interacting with the cytoplasmic dom ains of integrin beta 1, beta 2, and beta 3 subunits, Overexpression o f ILK in epithelial cells disrupts cell-extracellular matrix as well a s cell-cell interactions, suppresses suspension-induced apoptosis (als o called Anoikis), and stimulates anchorage-independent cell cycle pro gression. In addition, ILK induces nuclear translocation of beta-caten in, where the latter associates with a T cell factor/lymphocyte enhanc er-binding factor 1 (TCF/LEF-1) to form an activated transcription fac tor. We now demonstrate that ILK activity is rapidly, but transiently, stimulated upon attachment of cells to fibronectin, as well as by ins ulin, in a phosphoinositide-3-OH kinase [Pi(3)K]-dependent manner. Fur thermore, phosphatidylinositol(3,4,5)trisphosphate specifically stimul ates the activity of ILK irt vitro, and in addition, membrane targette d constitutively active Pi(3)K activates ILK in vivo. We also demonstr ate here that ILK is an upstream effector of the Pi(3)K-dependent regu lation of both protein kinase B (PKB/AKT) and glycogen synthase kinase 3 (GSK-3), Specifically, ILK can directly phosphorylate GSK-3 in vitr o and when stably, or transiently, overexpressed in cells can inhibit GSK-3 activity, whereas the overexpression of kinase-deficient ILK enh ances GSK-3 activity. In addition, kinase-active ILK can phosphorylate PKB/AKT on serine-473, whereas kinase-deficient ILK severely inhibits endogenous phosphorylation of PKB/AKT on serine-473, demonstrating th at ILK is involved in agonist stimulated, Pi(3)K-dependent, PKB/AKT ac tivation. ILK is thus a receptor-proximal effector for the Pi(3)K-depe ndent, extracellular matrix and growth factor mediated, activation of PKB/AKT, and inhibition of GSK-3.