Inflammation contributes to insulin resistance in diabetes and obesity. Mouse Pelle-like kinase (mPLK, homolog of human IL-1 receptor-associated kinase (IRAK)) participates in inflammatory signaling. We evaluated IRS-1 as a novel substrate for mPLK that may contribute to linking inflammation with insulin resistance. Wildtype mPLK, but not a kinase-inactive mutant (mPLK-KD), directly phosphorylated full-length IRS-1 in vitro. This in vitro phosphorylation was increased when mPLK was immunoprecipitated from tumor necrosis factor (TNF)-␣-treated cells. In NIH-3T3 IR cells, wild-type mPLK (but not mPLK-KD) co-immunoprecipitated with IRS-1. This association was increased by treatment of cells with TNF-␣. Using mass spectrometry, we identified Ser 24 in the pleckstrin homology (PH) domain of IRS-1 as a specific phosphorylation site for mPLK. IRS-1 mutants S24D or S24E (mimicking phosphorylation at Ser 24 ) had impaired ability to associate with insulin receptors resulting in diminished tyrosine phosphorylation of IRS-1 and impaired ability of IRS-1 to bind and activate PI-3 kinase in response to insulin. IRS-1-S24D also had an impaired ability to mediate insulin-stimulated translocation of GLUT4 in rat adipose cells. Importantly, endogenous mPLK/IRAK was activated in response to TNF-␣ or interleukin 1 treatment of primary adipose cells. In addition, using a phospho-specific antibody against IRS-1 phosphorylated at Ser 24 , we found that interleukin-1 or TNF-␣ treatment of Fao cells stimulated increased phosphorylation of endogenous IRS-1 at Ser 24 . We conclude that IRS-1 is a novel physiological substrate for mPLK. TNF-␣-regulated phosphorylation at Ser 24 in the pleckstrin homology domain of IRS-1 by mPLK/IRAK represents an additional mechanism for cross-talk between inflammatory signaling and insulin signaling that may contribute to metabolic insulin resistance.Biochemical, physiological, and epidemiological studies implicate pro-inflammatory cytokines (e.g. TNF-␣, 1 IL-1␤, and IL-6) in the development of insulin resistance and the pathophysiology of type 2 diabetes and obesity (1-8). These studies suggest an intriguing link between inflammation and metabolic dysregulation. Indeed, IB kinase ␤ (IKK␤), a critical mediator of inflammatory signaling pathways activating NF-B, has been identified as an important inhibitor of metabolic insulin signaling pathways (9 -12). Inactivation of IKK␤ signaling increases insulin sensitivity, whereas overexpression of IKK␤ or activation of IKK␤ by pro-inflammatory cytokines (e.g. TNF-␣) leads to insulin resistance (9, 12). Similarly, JNK is another inflammatory signaling molecule that may play a role in the insulin resistance of obesity (13). One potential explanation for these observations is cross-talk between inflammatory signaling and metabolic insulin signaling pathways.Metabolic actions of insulin such as enhanced glucose uptake into skeletal muscle and adipose tissue are regulated by activation of the insulin receptor tyrosine kinase and subsequent tyrosine phosphorylation o...

Angiotensin II, which stimulates AT(1) receptors, is a brain and peripheral stress hormone. We pretreated rats with the AT(1) receptor antagonist candesartan for 13 d via sc-implanted osmotic minipumps, followed by 24-h isolation in individual metabolic cages. We measured angiotensin II receptor-type binding and mRNAs and tyrosine hydroxylase mRNA by quantitative autoradiography and in situ hybridization, catecholamines by HPLC, and hormones by RIA. Isolation increased AT(1) receptor binding in hypothalamic paraventricular nucleus as well as anterior pituitary ACTH, and decreased posterior pituitary AVP. Isolation stress also increased AT(1) receptor binding and AT(1B) mRNA in zona glomerulosa and AT(2) binding in adrenal medulla, adrenal catecholamines, tyrosine hydroxylase mRNA, aldosterone, and corticosterone. Candesartan blocked AT(1) binding in paraventricular nucleus and adrenal gland; prevented the isolation-induced alterations in pituitary ACTH and AVP and in adrenal corticosterone, aldosterone, and catecholamines; abolished the increase in AT(2) binding in adrenal medulla; and substantially decreased urinary AVP, corticosterone, aldosterone, and catecholamines during isolation. Peripheral pretreatment with an AT(1) receptor antagonist blocks brain and peripheral AT(1) receptors and inhibits the hypothalamic-pituitary-adrenal response to stress, suggesting a physiological role for peripheral and brain AT(1) receptors during stress and a possible beneficial effect of AT(1) antagonism in stress-related disorders.

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