Title : Hyperglycemia inhibits
endothelial nitric oxide synthase activity by posttranslational modification at the
Akt site
Abstract :
- Endothelial nitric oxide synthase ( eNOS ) is activated by phosphorylation of serine 1177 by the protein kinase Akt/PKB
- Since hyperglycemia-induced mitochondrial superoxide overproduction increases O-linked N-acetylglucosamine modification and decreases O-linked phosphorylation of the transcription factor Sp1 , the effect of hyperglycemia and the hexosamine pathway on eNOS was evaluated
- In bovine aortic endothelial cells, hyperglycemia inhibited eNOS activity 67%, and treatment with glucosamine had a similar effect
- Hyperglycemia-associated inhibition of eNOS was accompanied by a twofold increase in O-linked N-acetylglucosamine modification of eNOS and a reciprocal decrease in O-linked serine phosphorylation at residue 1177
- Both the inhibition of eNOS and the changes in its post-translational modifications were reversed by antisense inhibition of glutamine glutamine:fructose-6-phosphate amidotransferase , the rate-limiting enzyme of the hexosamine pathway, or by blocking mitochondrial superoxide overproduction with uncoupling protein-1 (UCP-1 ) or manganese superoxide dismutase ( MnSOD )
- Immunoblot analysis of cells expressing myc-tagged wild-type human eNOS confirmed the reciprocal increase in O-linked N-acetylglucosamine and decrease in O-linked serine 1177 phosphorylation in response to hyperglycemia
- In contrast, when myc-tagged human eNOS carried a mutation at the Akt phosphorylation site ( Ser1177 ), O-linked N-acetylglucosamine modification was unchanged by hyperglycemia and phospho- eNOS was undetectable
- Similar changes in eNOS activity and covalent modification were found in aortae from diabetic animals
- Chronic impairment of eNOS activity by this mechanism may partly explain the accelerated atherosclerosis of diabetes