Activation of serum- and glucocorticoid-induced protein kinase (Sgk) by cyclic AMP and insulin.
Sgk (serum- and glucocorticoid-induced protein kinase) is a serine/threonine-specific protein kinase that is transcriptionally regulated by serum, glucorticoids, and mineralocorticoids. Sgk regulates the amiloride-sensitive sodium channel in kidney principal cells. Insulin and insulin-like growth factor-1 stimulate activity of Sgk by a mechanism mediated by phosphoinositide-dependent kinases (PDK)-1 and -2. In this study, we demonstrate that incubation of transfected cells with 8-(4-chlorophenylthio)-cAMP (8CPT-cAMP; 0.2 mm) led to a 2-fold activation of recombinant Sgk expressed in COS7 cells. Furthermore, the combination of insulin plus 8CPT-cAMP elicited a larger response than either agent alone. The effect of insulin was inhibited by wortmannin (100 nm), but not by the cyclic AMP-dependent protein kinase (PKA) inhibitor, H89 (10 microm). As expected, the effect of 8CPT-cAMP was completely blocked by H89. Surprisingly, the effect of 8CPT-cAMP was also inhibited by wortmannin, suggesting that phosphorylation of Sgk by PDK-1 and/or -2 is required for activation by 8CPT-cAMP. Mutational analysis led to similar conclusions. The Thr(369) --> Ala mutant, lacking the PKA phosphorylation site, was activated by insulin but not 8CPT-cAMP. In contrast, the Ser(422) --> Ala mutant, lacking a PDK-2 phosphorylation site, was inactive and resistant to activation by either insulin or 8CPT-cAMP. In summary, Sgk is subject to complex regulatory mechanisms. In addition to regulation at the level of gene expression, the enzymatic activity of Sgk is regulated by multiple protein kinases, including PKA, PDK-1, and PDK-2. Cross-talk among these signaling pathways may play an important role in the pathogenesis of the hypertension associated with hyperinsulinemia, obesity, and insulin resistance.
Perrotti
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N
Diabetes Branch, Division of Intramural Research, NIDDK, Bethesda, Maryland 20892, USA.
He
R A
RA
Phillips
S A
SA
Haft
C R
CR
Taylor
S I
SI
D000730:Androstadienes / Q000494:pharmacology; D000818:Animals; D001769:Blood; D019556:COS Cells; D000242:Cyclic AMP / Q000494:pharmacology*; D004790:Enzyme Induction; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005938:Glucocorticoids / Q000494:pharmacology; D017874:Immediate-Early Proteins; D007328:Insulin / Q000494:pharmacology*; D007546:Isoquinolines / Q000494:pharmacology; D016296:Mutagenesis; D009687:Nuclear Proteins; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000037:antagonists & inhibitors / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism; D013449:Sulfonamides; D000077191:Wortmannin; D000099184:Serum-Glucocorticoid Regulated Kinases
D000730:Androstadienes; D000242:Cyclic AMP; D004791:Enzyme Inhibitors; D005938:Glucocorticoids; D017874:Immediate-Early Proteins; D007328:Insulin; D007546:Isoquinolines; D009687:Nuclear Proteins; D017346:Protein Serine-Threonine Kinases; D011994:Recombinant Proteins; D013449:Sulfonamides; D000077191:Wortmannin; D000099184:Serum-Glucocorticoid Regulated Kinases; C063509:N-(2-(4-bromocinnamylamino)ethyl)-5-isoquinolinesulfonamide