title: Serine 302 Phosphorylation of Mouse Insulin Receptor Substrate 1 (IRS1) Is Dispensable for Normal Insulin Signaling and Feedback Regulation by Hepatic S6 Kinase.
abstract: Constitutive activation of the mammalian target of rapamycin complex 1 and S6 kinase (mTORC1→ S6K) attenuates insulin-stimulated Akt activity in certain tumors in part through "feedback" phosphorylation of the upstream insulin receptor substrate 1 (IRS1). However, the significance of this mechanism for regulating insulin sensitivity in normal tissue remains unclear. We investigated the function of Ser-302 in mouse IRS1, the major site of its phosphorylation by S6K in vitro, through genetic knock-in of a serine-to-alanine mutation (A302). Although insulin rapidly stimulated feedback phosphorylation of Ser-302 in mouse liver and muscle, homozygous A302 mice (A/A) and their knock-in controls (S/S) exhibited similar glucose homeostasis and muscle insulin signaling. Furthermore, both A302 and control primary hepatocytes from which Irs2 was deleted showed marked inhibition of insulin-stimulated IRS1 tyrosine phosphorylation and PI3K binding after emetine treatment to raise intracellular amino acids and activate mTORC1 → S6K signaling. To specifically activate mTORC1 in mouse tissue, we deleted hepatic Tsc1 using Cre adenovirus. Although it moderately decreased IRS1/PI3K association and Akt phosphorylation in liver, Tsc1 deletion failed to cause glucose intolerance or promote hyperinsulinemia in mixed background A/A or S/S mice. Moreover, Tsc1 deletion failed to stimulate phospho-Ser-302 or other putative S6K sites within IRS1, whereas ribosomal S6 protein was constitutively phosphorylated. Following acute Tsc1 deletion from hepatocytes, Akt phosphorylation, but not IRS1/PI3K association, was rapidly restored by treatment with the mTORC1 inhibitor rapamycin. Thus, within the hepatic compartment, mTORC1 → S6K signaling regulates Akt largely through IRS-independent means with little effect upon physiologic insulin sensitivity.
authors: From the Division of Endocrinology, Department of Medicine, Boston Children's Hospital, Harvard Medical School, Boston, Massachusetts 02115.
authors: Kyle D
authors: KD
authors: Copps
authors: From the Division of Endocrinology, Department of Medicine, Boston Children's Hospital, Harvard Medical School, Boston, Massachusetts 02115.
authors: Nancy J
authors: NJ
authors: Hançer
authors: From the Division of Endocrinology, Department of Medicine, Boston Children's Hospital, Harvard Medical School, Boston, Massachusetts 02115.
authors: Wei
authors: W
authors: Qiu
authors: From the Division of Endocrinology, Department of Medicine, Boston Children's Hospital, Harvard Medical School, Boston, Massachusetts 02115 morris.white@childrens.harvard.edu.
authors: Morris F
authors: MF
authors: White
mesh_terms: D019943:Amino Acid Substitution; D000818:Animals; D016466:CHO Cells; D006224:Cricetinae; D003412:Cricetulus; D017353:Gene Deletion; D018149:Glucose Intolerance / Q000235:genetics / Q000378:metabolism; D007328:Insulin / Q000172:deficiency / Q000378:metabolism*; D055504:Insulin Receptor Substrate Proteins / Q000235:genetics / Q000378:metabolism*; D008099:Liver / Q000378:metabolism*; D000076222:Mechanistic Target of Rapamycin Complex 1; D051379:Mice; D008822:Mice, Transgenic; D046912:Multiprotein Complexes / Q000235:genetics / Q000378:metabolism; D020125:Mutation, Missense; D019869:Phosphatidylinositol 3-Kinases / Q000235:genetics / Q000378:metabolism; D010766:Phosphorylation / Q000502:physiology; D051057:Proto-Oncogene Proteins c-akt / Q000235:genetics / Q000378:metabolism; D019893:Ribosomal Protein S6 Kinases / Q000235:genetics / Q000378:metabolism*; D012694:Serine / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction / Q000502:physiology*; D058570:TOR Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism; D000077004:Tuberous Sclerosis Complex 1 Protein; D025521:Tumor Suppressor Proteins / Q000235:genetics / Q000378:metabolism
chemical_list: D007328:Insulin; D055504:Insulin Receptor Substrate Proteins; D000076222:Mechanistic Target of Rapamycin Complex 1; D046912:Multiprotein Complexes; D019869:Phosphatidylinositol 3-Kinases; D051057:Proto-Oncogene Proteins c-akt; D019893:Ribosomal Protein S6 Kinases; D012694:Serine; D058570:TOR Serine-Threonine Kinases; D000077004:Tuberous Sclerosis Complex 1 Protein; D025521:Tumor Suppressor Proteins; C526295:Irs1 protein, mouse; C000624651:Tsc1 protein, mouse
keywords: S6 kinase; amino acids; gene knock-in; insulin receptor substrate 1 (IRS-1); insulin resistance; insulin signaling; mammalian target of rapamycin (mTOR); serine/threonine phosphorylation; tuberous sclerosis complex (TSC)