Abstract 15746: Cardiac PI3Kγ Regulates GSK-3 Independent Of Akt By Scaffolding PP2A
G-protein coupled receptor activation of phosphoinositide 3-kinase γ (PI3Kγ) regulates glycogen synthase kinase-3 (GSK-3) via protein kinase B (Akt) mediating cardiac hypertrophy. We show here that PI3Kγ regulates cardiac GSK-3 phosphorylation downstream of growth factor activated PI3Kα-Akt axis. Insulin stimulation results in rapid dephosphorylation of GSK-3 in PI3Kγ knock out mice (PI3Kγ KO) compared to littermate controls despite sustained Akt activation. Confocal microscopy and immunoblotting show marked reduction in GSK-3 phosphorylation in PI3Kγ KO. Analysis of GSK-3 dephosphorylating protein phosphatase 2A (PP2A) activity showed significant elevation in the PI3Kγ KO suggesting a novel regulation of PP2A by PI3Kγ. Importantly, GSK-3 associated PP2A and phosphatase activity are significantly higher in PI3Kγ KO mice accounting for the loss of GSK-3 phosphorylation. PI3Kγ KO mice were characterized by significant PP2A methylation consistent with the idea that methylation of PP2A enhances phosphatase activity. Furthermore, enhanced association of PP2A methyl transferase-1 (PPMT-1) with PP2A is observed in PI3Kγ KO compared to controls. Consistent with this observation, GSK-3 in PI3Kγ KO is associated with higher levels of methylated PP2A leading to elevated PP2A activity resulting in GSK-3 dephosphorylation. Interestingly, cardiac specific expression of inactive PI3Kγ transgene (deletion in catalytic domain) in the PI3Kγ KO genetic background reversed GSK-3 phosphorylation, associated phosphatase and PPMT activity suggesting PI3Kγ inhibits GSK-3 associated PP2A in a kinase independent manner. Mechanistically, PI3Kγ alters PP2A activity by regulating PP2A methylation via inhibition of PP2A methyl transferase-1 (PPMT-1) as PI3Kγ KO mice have elevated PPMT activity. Normalization of GSK-3 phosphorylation following cardiac expression of inactive PI3Kγ transgene on a PI3Kγ KO background demonstrates that PI3Kγ acts as a scaffold inhibiting PP2A methylation by PPMT leading to reduced PP2A activity. NFATc3 the downstream target of GSK-3 is significantly phosphorylated and exported out of the nucleus in PI3Kγ KO mice compared to controls implicating a role of scaffolding function of PI3Kγ in cardiac physiology/pathology.
- © 2011 by American Heart Association, Inc.