Enhancement of insulin responsiveness by nitric oxide-mediated inactivation of protein-tyrosine phosphatases.
NO synthesis is a prerequisite for proper insulin sensitivity in insulin-targeted tissues; however, the molecular basis for this process remains unclear. Using a gain-of-function model of endothelial nitric-oxide synthase (eNOS)-transfected COS-7 cells, we have shown a critical role of NO in insulin responsiveness, as evidenced by an NO-dependent increase of tyrosine phosphorylation levels of the insulin receptor and its downstream effectors insulin receptor substrate-1 and PKB/AKT. We hypothesized that NO-induced inactivation of endogenous protein-tyrosine phosphatases (PTPs) would enhance insulin receptor-mediated signaling. To test this hypothesis, we devised a new method of the PTP labeling using a cysteine sulfhydryl-reacted probe. Under the acidic conditions employed in this study, the probe recognized the reduced and active forms but not the S-nitrosylated and inactive forms of endogenous PTPs. Our data suggest that phosphatases SHP-1, SHP-2, and PTP1B, but not TC-PTP, are likely S-nitrosylated at the active site cysteine residue concomitantly with a burst of NO production in signaling response to insulin stimulation. These results were further confirmed by phosphatase activity assays. We investigated further the role of NO as a regulator of insulin signaling by RNA interference that ablates endogenous eNOS expression in endothelial MS-1 cells. We have shown that eNOS-dependent NO production is essential for the activation of insulin signaling. Our findings demonstrate that NO mediates enhancement of insulin responsiveness via the inhibition of insulin receptor phosphatases.
Hsu
Ming-Fo
MF
Institute of Biological Chemistry, Academia Sinica, Taipei 11529, Taiwan.
Meng
Tzu-Ching
TC
D000818:Animals; D019556:COS Cells; D002522:Chlorocebus aethiops; D003545:Cysteine / Q000378:metabolism; D003924:Diabetes Mellitus, Type 2 / Q000378:metabolism*; D007328:Insulin / Q000378:metabolism*; D055504:Insulin Receptor Substrate Proteins / Q000378:metabolism; D009569:Nitric Oxide / Q000378:metabolism*; D020030:Nitric Oxide Donors / Q000494:pharmacology; D052250:Nitric Oxide Synthase Type III / Q000235:genetics / Q000378:metabolism; D009584:Nitrogen / Q000378:metabolism; D054562:Protein Tyrosine Phosphatase, Non-Receptor Type 1 / Q000378:metabolism; D054592:Protein Tyrosine Phosphatase, Non-Receptor Type 11 / Q000378:metabolism; D054578:Protein Tyrosine Phosphatase, Non-Receptor Type 2 / Q000378:metabolism; D053824:Protein Tyrosine Phosphatase, Non-Receptor Type 6 / Q000378:metabolism; D017027:Protein Tyrosine Phosphatases / Q000378:metabolism*; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D034741:RNA, Small Interfering; D015398:Signal Transduction / Q000502:physiology*; D014162:Transfection; D014443:Tyrosine / Q000378:metabolism
D007328:Insulin; D055504:Insulin Receptor Substrate Proteins; D020030:Nitric Oxide Donors; D034741:RNA, Small Interfering; D009569:Nitric Oxide; D014443:Tyrosine; D052250:Nitric Oxide Synthase Type III; D051057:Proto-Oncogene Proteins c-akt; D054562:Protein Tyrosine Phosphatase, Non-Receptor Type 1; D054592:Protein Tyrosine Phosphatase, Non-Receptor Type 11; D054578:Protein Tyrosine Phosphatase, Non-Receptor Type 2; D053824:Protein Tyrosine Phosphatase, Non-Receptor Type 6; D017027:Protein Tyrosine Phosphatases; D003545:Cysteine; D009584:Nitrogen