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Morris F. White - One of the best experts on this subject based on the ideXlab platform.
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IRS proteins and diabetic complications
Diabetologia, 2016Co-Authors: Deborah P. Lavin, Morris F. White, Derek P BrazilAbstract:IRS proteins are cellular adaptor molecules that mediate many of the key metabolic actions of insulin. When tyrosine is phosphorylated by the activated insulin receptor, IRS proteins recruit downstream effectors, such as phosphoinositide 3-kinase and mitogen-activated protein kinase, in order to elicit cellular responses such as glucose uptake, lipid metabolism and cell proliferation. There are two main IRS proteins in humans (IRS1 and IRS2), both of which are widely expressed. Given their central role in the insulin signalling pathway, it is not surprising that male mice lacking Irs1 or IRS2 present with elevated blood glucose or type 2 diabetes, respectively. For reasons yet to be identified, female IRS2 ^−/− mice do not develop type 2 diabetes. A number of organs are affected by complications of diabetes; macrovascular complications include stroke and coronary artery disease, while nephropathy, neuropathy and retinopathy fall into the category of microvascular complications. Given the serious consequences of these complications on patient morbidity and mortality, it is essential to identify the molecular pathogenesis underlying diabetic complications, with a view to improving therapeutic intervention and patient outcomes. A number of recently published papers have converged on the hypothesis that the loss of insulin signalling and IRS proteins is instrumental to the development and/or progression of diabetic complications. This review will summarise some highlights from the published work in which this hypothesis is discussed.
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insulin and metabolic stress stimulate multisite serine threonine phosphorylation of insulin receptor substrate 1 and inhibit tyrosine phosphorylation
Journal of Biological Chemistry, 2014Co-Authors: Nancy J Hancer, Wei Qiu, Christine E Cherella, Kyle D Copps, Morris F. WhiteAbstract:IRS1 and IRS2 are key substrates of the insulin receptor tyrosine kinase. Mass spectrometry reveals more than 50 phosphorylated IRS1 serine and threonine residues (Ser(P)/Thr(P) residues) in IRS1 from insulin-stimulated cells or human tissues. We investigated a subset of IRS1 Ser(P)/Thr(P) residues using a newly developed panel of 25 phospho-specific monoclonal antibodies (αpS/TmAbIrs1). CHO cells overexpressing the human insulin receptor and rat IRS1 were stimulated with insulin in the absence or presence of inhibitors of the PI3K → Akt → mechanistic target of rapamycin (mTOR) → S6 kinase or MEK pathways. Nearly all IRS1 Ser(P)/Thr(P) residues were stimulated by insulin and significantly suppressed by PI3K inhibition; fewer were suppressed by Akt or mTOR inhibition, and none were suppressed by MEK inhibition. Insulin-stimulated Irs1 tyrosine phosphorylation (Tyr(P)Irs1) was enhanced by inhibition of the PI3K → Akt → mTOR pathway and correlated with decreased Ser(P)-302Irs1, Ser(P)-307Irs1, Ser(P)-318Irs1, Ser(P)-325Irs1, and Ser(P)-346Irs1. Metabolic stress modeled by anisomycin, thapsigargin, or tunicamycin increased many of the same Ser(P)/Thr(P) residues as insulin, some of which (Ser(P)-302Irs1, Ser(P)-307Irs1, and four others) correlated significantly with impaired insulin-stimulated Tyr(P)Irs1. Thus, IRS1 Ser(P)/Thr(P) is an integrated response to insulin stimulation and metabolic stress, which associates with reduced Tyr(P)Irs1 in CHOIR/IRS1 cells.
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Myocardial Loss of IRS1 and IRS2 Causes Heart Failure and Is Controlled by p38α MAPK During Insulin Resistance
Diabetes, 2013Co-Authors: Qinglei Zhu, Morris F. White, Candice M. Thomas, Rajesh Kumar, Hao Feng, David E. Dostal, Kenneth M. Baker, Shaodong GuoAbstract:Cardiac failure is a major cause of death in patients with type 2 diabetes, but the molecular mechanism that links diabetes to heart failure remains unclear. Insulin resistance is a hallmark of type 2 diabetes, and insulin receptor substrates 1 and 2 (IRS1 and IRS2) are the major insulin-signaling components regulating cellular metabolism and survival. To determine the role of IRS1 and IRS2 in the heart and examine whether hyperinsulinemia causes myocardial insulin resistance and cellular dysfunction via IRS1 and IRS2, we generated heart-specific IRS1 and IRS2 gene double-knockout (H-DKO) mice and liver-specific IRS1 and IRS2 double-knockout (L-DKO) mice. H-DKO mice had reduced ventricular mass; developed cardiac apoptosis, fibrosis, and failure; and showed diminished Akt→forkhead box class O-1 signaling that was accompanied by impaired cardiac metabolic gene expression and reduced ATP content. L-DKO mice had decreased cardiac IRS1 and IRS2 proteins and exhibited features of heart failure, with impaired cardiac energy metabolism gene expression and activation of p38α mitogen-activated protein kinase (p38). Using neonatal rat ventricular cardiomyocytes, we further found that chronic insulin exposure reduced IRS1 and IRS2 proteins and prevented insulin action through activation of p38, revealing a fundamental mechanism of cardiac dysfunction during insulin resistance and type 2 diabetes.
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IRS2 and Irs4 synergize in non-LepRb neurons to control energy balance and glucose homeostasis.
Molecular metabolism, 2013Co-Authors: Marianna Sadagurski, X. Charlie Dong, Martin G. Myers, Morris F. WhiteAbstract:Insulin receptor substrates (Irs1, 2, 3 and Irs4) mediate the actions of insulin/IGF1 signaling. They have similar structure, but distinctly regulate development, growth, and metabolic homeostasis. IRS2 contributes to central metabolic sensing, partially by acting in leptin receptor (LepRb)-expressing neurons. Although Irs4 is largely restricted to the hypothalamus, its contribution to metabolic regulation is unclear because Irs4-null mice barely distinguishable from controls. We postulated that IRS2 and Irs4 synergize and complement each other in the brain. To examine this possibility, we investigated the metabolism of whole body Irs4(-/y) mice that lacked IRS2 in the CNS (bIRS2(-/-)·Irs4(-/y)) or only in LepRb-neurons (Lepr (∆IRS2) ·Irs4 (-/y) ). bIRS2(-/-)·Irs4(-/y) mice developed severe obesity and decreased energy expenditure, along with hyperglycemia and insulin resistance. Unexpectedly, the body weight and fed blood glucose levels of Lepr (∆IRS2) ·Irs4 (-/y) mice were not different from Lepr (∆IRS2) mice, suggesting that the functions of IRS2 and Irs4 converge upon neurons that are distinct from those expressing LepRb.
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Serine Phosphorylation Sites on IRS2 Activated by Angiotensin II and Protein Kinase C To Induce Selective Insulin Resistance in Endothelial Cells
Molecular and cellular biology, 2013Co-Authors: Kyoungmin Park, Morris F. White, Christian Rask-madsen, Akira Mima, Koji Mizutani, Jonathon N. Winnay, Yasutaka Maeda, Katharine E. D’aquino, Edward P. FeenerAbstract:Protein kinase C (PKC) activation, induced by hyperglycemia and angiotensin II (AngII), inhibited insulin-induced phosphorylation of Akt/endothelial nitric oxide (eNOS) by decreasing tyrosine phosphorylation of IRS2 (p-Tyr-IRS2) in endothelial cells. PKC activation by phorbol ester (phorbol myristate acetate [PMA]) reduced insulin-induced p-Tyr-IRS2 by 46% ± 13% and, similarly, phosphorylation of Akt/eNOS. Site-specific mutational analysis showed that PMA increased serine phosphorylation at three sites on IRS2 (positions 303, 343, and 675), which affected insulin-induced tyrosine phosphorylation of IRS2 at positions 653, 671, and 911 (p-Tyr-IRS2) and p-Akt/eNOS. Specific PKCβ2 activation decreased p-Tyr-IRS2 and increased the phosphorylation of two serines (Ser303 and Ser675) on IRS2 that were confirmed in cells overexpressing single point mutants of IRS2 (S303A or S675A) containing a PKCβ2-dominant negative or selective PKCβ inhibitor. AngII induced phosphorylation only on Ser303 of IRS2 and inhibited insulin-induced p-Tyr911 of IRS2 and p-Akt/eNOS, which were blocked by an antagonist of AngII receptor I, losartan, or overexpression of single mutant S303A of IRS2. Increases in p-Ser303 and p-Ser675 and decreases in p-Tyr911 of IRS2 were observed in vessels of insulin-resistant Zucker fatty rats versus lean rats. Thus, AngII or PKCβ activation can phosphorylate Ser303 and Ser675 in IRS2 to inhibit insulin-induced p-Tyr911 and its anti-atherogenic actions (p-Akt/eNOS) in endothelial cells.
Susan O Meakin - One of the best experts on this subject based on the ideXlab platform.
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trk receptor binding and neurotrophin fibroblast growth factor fgf dependent activation of the fgf receptor substrate frs 3
Biochimica et Biophysica Acta, 2006Co-Authors: Scott J Dixon, James I S Macdonald, Susan O Meakin, Kim N Robinson, Christopher J KubuAbstract:Abstract We have investigated the signaling properties of the fibroblast growth factor (FGF) receptor substrate 3 (FRS3), also known as SNT-2 or FRS2β, in neurotrophin-dependent differentiation in comparison with the related adapter FRS2 (SNT1 or FRS2α). We demonstrate that FRS3 binds all neurotrophin Trk receptor tyrosine kinases and becomes tyrosine phosphorylated in response to NGF, BDNF, NT-3 and FGF stimulation in transfected cells and/or primary cortical neurons. Second, the signaling molecules Grb2 and Shp2 bind FRS3 at consensus sites that are highly conserved among FRS family members and that Shp2, in turn, becomes tyrosine phosphorylated. While FRS3 over-expression in PC12 cells neither increases NGF-induced neuritogenesis nor activation of Map kinase/AKT, comparable to previous reports on FRS2, over-expression of a chimeric adapter containing the PH/PTB domains of the insulin receptor substrate (IRS) 2, in place of the PTB domain of FRS3 (IRS2-FRS3) supports insulin-dependent Map kinase activation and neurite outgrowth in PC12 cells. Collectively, these data demonstrate that FRS3 supports ligand-induced Map kinase activation and that the chimeric IRS2-FRS3 adapter is stimulating sufficient levels of activated MapK to support neurite outgrowth in PC12 cells.
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trk receptor binding and neurotrophin fibroblast growth factor fgf dependent activation of the fgf receptor substrate frs 3
Biochimica et Biophysica Acta, 2006Co-Authors: Scott J Dixon, James I S Macdonald, Susan O Meakin, Kim N Robinson, Christopher J KubuAbstract:We have investigated the signaling properties of the fibroblast growth factor (FGF) receptor substrate 3 (FRS3), also known as SNT-2 or FRS2beta, in neurotrophin-dependent differentiation in comparison with the related adapter FRS2 (SNT1 or FRS2alpha). We demonstrate that FRS3 binds all neurotrophin Trk receptor tyrosine kinases and becomes tyrosine phosphorylated in response to NGF, BDNF, NT-3 and FGF stimulation in transfected cells and/or primary cortical neurons. Second, the signaling molecules Grb2 and Shp2 bind FRS3 at consensus sites that are highly conserved among FRS family members and that Shp2, in turn, becomes tyrosine phosphorylated. While FRS3 over-expression in PC12 cells neither increases NGF-induced neuritogenesis nor activation of Map kinase/AKT, comparable to previous reports on FRS2, over-expression of a chimeric adapter containing the PH/PTB domains of the insulin receptor substrate (IRS) 2, in place of the PTB domain of FRS3 (IRS2-FRS3) supports insulin-dependent Map kinase activation and neurite outgrowth in PC12 cells. Collectively, these data demonstrate that FRS3 supports ligand-induced Map kinase activation and that the chimeric IRS2-FRS3 adapter is stimulating sufficient levels of activated MapK to support neurite outgrowth in PC12 cells.
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Trk receptor binding and neurotrophin/fibroblast growth factor (FGF)-dependent activation of the FGF receptor substrate (FRS)-3
Biochimica et Biophysica Acta (BBA) - Molecular Cell Research, 2006Co-Authors: Scott J Dixon, James I S Macdonald, Kim N Robinson, Christopher J Kubu, Susan O MeakinAbstract:We have investigated the signaling properties of the fibroblast growth factor (FGF) receptor substrate 3 (FRS3), also known as SNT-2 or FRS2beta, in neurotrophin-dependent differentiation in comparison with the related adapter FRS2 (SNT1 or FRS2alpha). We demonstrate that FRS3 binds all neurotrophin Trk receptor tyrosine kinases and becomes tyrosine phosphorylated in response to NGF, BDNF, NT-3 and FGF stimulation in transfected cells and/or primary cortical neurons. Second, the signaling molecules Grb2 and Shp2 bind FRS3 at consensus sites that are highly conserved among FRS family members and that Shp2, in turn, becomes tyrosine phosphorylated. While FRS3 over-expression in PC12 cells neither increases NGF-induced neuritogenesis nor activation of Map kinase/AKT, comparable to previous reports on FRS2, over-expression of a chimeric adapter containing the PH/PTB domains of the insulin receptor substrate (IRS) 2, in place of the PTB domain of FRS3 (IRS2-FRS3) supports insulin-dependent Map kinase activation and neurite outgrowth in PC12 cells. Collectively, these data demonstrate that FRS3 supports ligand-induced Map kinase activation and that the chimeric IRS2-FRS3 adapter is stimulating sufficient levels of activated MapK to support neurite outgrowth in PC12 cells.
Deborah J. Burks - One of the best experts on this subject based on the ideXlab platform.
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The influence of the lack of insulin receptor substrate 2 (IRS2) on the thyroid gland.
Scientific reports, 2019Co-Authors: María Carmen Iglesias-osma, Deborah J. Burks, Leonardo Catalano-iniesta, Virginia Sanchez-robledo, Marta Carretero-hernández, Enrique J. Blanco, María José García-barrado, Teresa Vicente-garcia, José CarreteroAbstract:Involvement of IRS2 in the proliferative effects of IGF-I of follicular thyroid cells has been described, but there are no evidences for in vivo participation of IRS2. This study aimed to analyse the in vivo relevance of IRS2 in the proliferation and apoptosis of thyroid cells by immunocytochemical studies for PCNA, Ki67, and active-caspase-3 in thyroid cells of IRS2 knockout (IRS2-KO) mice, jointly to TUNEL assay. Thyroid hormones were lower in IRS2-KO mice than in their wild-type (WT) counterparts. Increases in the area, perimeter and diameter of thyroid follicles of IRS2-KO mice were observed, which also showed increased proliferation rate of follicular cells and decreased percentage of apoptotic cells that was more evident in the central than in the marginal region of the gland. Sex-related differences were also found, since the follicular epithelium height was higher in male than in female mice. The percentage of proliferating cells showed significant changes in male but not in female mice, and apoptotic cells were more abundant in female than in male IRS2-KO animals, without significant differences between WT-animals. Therefore, our results suggest that IRS2 could be involved in the maintenance of thyroid cells population and in the normal physiology of the thyroid gland.
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Insulin/IRS2 signaling promotes epithelialization and sensitivity to FGF7 in vitro.
2019Co-Authors: Fátima Manzano-núñez, Deborah J. Burks, María José Arámbul-anthony, Amparo Galán Albiñana, Aranzazu Leal Tassias, Carlos Acosta Umanzor, Irene Borreda Gascó, Antonio Herrera, Jerónimo Forteza Vila, Luke A. NoonAbstract:(A) Schematic showing differentiation of bipotent HepaRG human LPCs in vitro produces epithelial “islands” containing hepatocyte-like cells. (B) RT-qPCR time course of HepaRG differentiation showing early induction of IRS2, and FGFR2-IIIb prior to hepatocyte differentiation (indicated by APOA2). (C) Colocalization of Alb, FGFR2, and IRS2 by immunostaining within hepatocyte islands during the early stages of HepaRG differentiation (day 14). IRS2 expression was visualized by stable transduction with a human IRS2 promoter (pIRS2) GFP reporter. Dotted line delimits “island.” (D–E) IRS2 is necessary for FGFR2-IIIb expression during HepaRG differentiation: (D) phase-contrast images of differentiated cells after stable silencing for IRS2 using shRNA lentivirus (sh-IRS2). Arrowheads highlight “islands” observed in control cultures expressing sh-scram but not in sh-IRS2 cells. Specificity of the knockdown phenotype was confirmed by a “rescue” in which Lenti-mIRS2 was constitutively expressed in sh-IRS2 cells. (E) RT-qPCR analysis of FGFR2-IIIb expression (n = 3). (F–G) Insulin/IRS2 is required for functional sensitivity to rFGF7 in HepaRG cells and is necessary for feed-forward induction of FGFR2-IIIb by FGF7 ligand. (F) Phase-contrast and immunofluorescence images of HepaRG cells (day 15) cultured with rhFGF7 for 11 days, showing an increase in epithelial islands positive for Alb/HNF4α and E-cad immunostaining and pAPOA2-GFP reporter expression (H = Hoechst). (G) RT-qPCR analysis for indicated genes following long-term (11-day) rhFGF7 treatment in the presence (+) or absence (−) of supplemented insulin in control (sh-luc) or IRS2 knockdown cells (sh-IRS2) (n = 3). Data information: underlying data are available in S1 Data. Data are presented as mean + SEM. *P < 0.05, **P < 0.01, and ***P < 0.001. (E and G) One-way ANOVA was used to compare means. Significance P values were calculated using Newman–Keuls multiple comparison test. Alb, albumin; APOA2, apolipoprotein A2; Cont, control; E-cad, E-cadherin; EF1, Human elongation factor-1 alpha promoter; FGF7, fibroblast growth factor 7; FGFR2, fibroblast growth factor receptor 2; GFP, green fluorescent protein; HNF4α, hepatocyte nuclear factor 4-alpha; IRS2, insulin receptor substrate 2; Lenti-mIRS2, murine IRS2 transgene not targeted by human-gene–silencing construct; mRNA, messenger RNA; pAPOA2, human APOA2 promoter; pIRS2, IRS2 promoter; rFgf7, recombinant Fgf7; rhFGF7, recombinant human FGF7; RT-qPCR, reverse transcriptase-quantitative PCR; RU, relative unit; sh-IRS2, shRNA-targeting IRS2; sh-luc, control luciferase; shRNA, short hairpin RNA; sh-scram, scrambled shRNA.
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Insulin Receptor Substrate 2 Is Required for Testicular Development
PloS one, 2013Co-Authors: Richard J. Griffeth, José Carretero, Deborah J. BurksAbstract:Insulin receptor substrate (IRS) proteins are key mediators of insulin and insulin-like growth factor (IGF) signalling. In mice, deletion of Irs1 is associated with profound growth retardation and increased longevity whereas IRS2-deficiency causes diabetes and female infertility. Clinical studies suggest that diabetes and obesity diminish male fertility. However, the role of IRS proteins in male reproduction is unknown. We observed that testis weight is reduced by 45% in IRS2-deficient mice as compared with control males. The weight of these organs in Irs1(-/-) males was similar to controls; however, since Irs1-deficient mice are 50% smaller, testis weight:body weight was increased in this model. Neonatal IRS2(-/-) mice also exhibited reduced testicular size, suggesting that impairments in this model occur during development. Histological examination of testicular cross sections from IRS2(-/-) mice revealed normal cellular associations without obvious abnormalities in the seminiferous epithelium. Reduced testicular weight was associated with fewer Sertoli cells, spermatogonia, spermatocytes, elongated spermatids, and epididymal spermatozoa. However, Leydig cell number and the concentration of serum testosterone were equivalent between IRS2-deficient and control males. Testicular weight was reduced similarly in non-diabetic and diabetic IRS2(-/-) mice, indicating that hyperglycemia does not compound the effects of IRS2 deletion on impaired testis development. Expression of Irs1, Irs3, and Irs4 was comparable between experimental groups. Collectively, our results demonstrate that IRS2 plays a critical role in testicular development, potentially by mediating IGF1 signalling during embryonic and early postnatal development.
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Insulin receptor substrate 2 (IRS2)-deficient mice show sensorineural hearing loss that is delayed by concomitant protein tyrosine phosphatase 1B (PTP1B) loss of function.
Molecular medicine (Cambridge Mass.), 2012Co-Authors: Silvia Murillo-cuesta, Deborah J. Burks, Águeda González-rodríguez, Ángela M. Valverde, Guadalupe Camarero, Lourdes Rodriguez-de La Rosa, Carlos Avendaño, Isabel Varela-nietoAbstract:The insulin receptor substrate (IRS) proteins are key mediators of insulin and insulinlike growth factor 1 (IGF-1) signaling. Protein tyrosine phosphatase (PTP)-1B dephosphorylates and inactivates both insulin and IGF-1 receptors. IRS2-deficient mice present altered hepatic insulin signaling and β-cell failure and develop type 2-like diabetes. In addition, IRS2 deficiency leads to developmental defects in the nervous system. IGF1 gene mutations cause syndromic sensorineural hearing loss in humans and mice. However, the involvement of IRS2 and PTP1B, two IGF-1 downstream signaling mediators, in hearing onset and loss has not been studied. Our objective was to study the hearing function and cochlear morphology of IRS2-null mice and the impact of PTP1B deficiency. We have studied the auditory brainstem responses and the cochlear morphology of systemic IRS2−/−Ptpn1+/+, IRS2+/+Ptpn1−/− and IRS2−/−Ptpn1−/− mice at different postnatal ages. The results indicated that IRS2−/−Ptpn1+/+ mice present a profound congenital sensorineural deafness before the onset of diabetes and altered cochlear morphology with hypoinnervation of the cochlear ganglion and aberrant stria vascularis, compared with wild-type mice. Simultaneous PTP1B deficiency in IRS2−/−Ptpn1−/− mice delays the onset of deafness. We show for the first time that IRS2 is essential for hearing and that PTP1B inhibition may be useful for treating deafness associated with hyperglycemia and type 2 diabetes.
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Inhibition of PTP1B restores IRS1-mediated hepatic insulin signaling in IRS2-deficient mice.
Diabetes, 2009Co-Authors: Águeda González-rodríguez, Deborah J. Burks, Jose A. Mas Gutierrez, Silvia M. Sanz-gonzález, Manuel Ros, Ángela M. ValverdeAbstract:OBJECTIVE Mice with complete deletion of insulin receptor substrate 2 (IRS2) develop hyperglycemia, impaired hepatic insulin signaling, and elevated gluconeogenesis, whereas mice deficient for protein tyrosine phosphatase (PTP)1B display an opposing hepatic phenotype characterized by increased sensitivity to insulin. To define the relationship between these two signaling pathways in the regulation of liver metabolism, we used genetic and pharmacological approaches to study the effects of inhibiting PTP1B on hepatic insulin signaling and expression of gluconeogenic enzymes in IRS2 −/− mice. RESEARCH DESIGN AND METHODS We analyzed glucose homeostasis and insulin signaling in liver and isolated hepatocytes from IRS2 −/− and IRS2 −/− /PTP1B −/− mice. Additionally, hepatic insulin signaling was assessed in control and IRS2 −/− mice treated with resveratrol, an antioxidant present in red wine. RESULTS In livers of hyperglycemic IRS2 −/− mice, the expression levels of PTP1B and its association with the insulin receptor (IR) were increased. The absence of PTP1B in the double-mutant mice restored hepatic IRS1-mediated phosphatidylinositol (PI) 3-kinase/Akt/Foxo1 signaling. Moreover, resveratrol treatment of hyperglycemic IRS2 −/− mice decreased hepatic PTP1B mRNA and inhibited PTP1B activity, thereby restoring IRS1-mediated PI 3-kinase/Akt/Foxo1 signaling and peripheral insulin sensitivity. CONCLUSIONS By regulating the phosphorylation state of IR, PTB1B determines sensitivity to insulin in liver and exerts a unique role in the interplay between IRS1 and IRS2 in the modulation of hepatic insulin action.
Kyle D Copps - One of the best experts on this subject based on the ideXlab platform.
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Hepatic Insulin Signaling Regulates Hepatokines to Coordinate Energy Expenditure and Weight Gain through IRS2 in Sim1 Neurons
Diabetes, 2018Co-Authors: Rongya Tao, Kyle D Copps, Oliver Stoehr, Orris F. WhiteAbstract:Insulin resistance is the underlying cause of type 2 diabetes; however, insulin resistance might also be a feed-back mechanism to protect cells and tissues from acute effects of over nutrition. Based upon our results with LDKO (hepatic specific Irs1L/LIRS2L/L CreAlbumin) mice, we found that hepatic insulin resistance elevated systemic oxygen consumption and energy expenditure, which was associated with a dramatic reduction of the RER (respiratory exchange ratio) and resistance to diet-induced obesity. Deletion of hepatic FoxO1 in LDKO-mice restored energy homeostasis and oxygen consumption, and increased the RER into the normal range. Contrary to the progressive peripheral insulin resistance of the LDKO-mice, IRS2 expression and insulin sensitivity increased in the PVH (paraventricular nucleus of hypothalamus). Interestingly, deletion of IRS2 in the PVH (IRS2L/L-mice intercrossed with CreSim1-mice) increased body weight and promoted obesity. To identify the molecular cross-talk between liver and brain, we used a neuronal cell line to screen for the effects of hepatokines that were dysregulated in LDKO-mice—but normalized in LTKO-mice. Among these dysregulated hepatokines, excess circulating NTF3 and GPX3 led to increased IRS2 expression and insulin sensitivity in neurons. Thus, our study suggests that hepatic FoxO1-dependent hepatokines—NTF3 or GPX3—modulate systemic energy expenditure and body weight through IRS2 expression in the PVH. Disclosure R. Tao: None. K.D. Copps: None. O. Stoehr: None. M.F. White: Advisory Panel; Self; Housey Pharmaceutical Research Laboratories.
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insulin and metabolic stress stimulate multisite serine threonine phosphorylation of insulin receptor substrate 1 and inhibit tyrosine phosphorylation
Journal of Biological Chemistry, 2014Co-Authors: Nancy J Hancer, Wei Qiu, Christine E Cherella, Kyle D Copps, Morris F. WhiteAbstract:IRS1 and IRS2 are key substrates of the insulin receptor tyrosine kinase. Mass spectrometry reveals more than 50 phosphorylated IRS1 serine and threonine residues (Ser(P)/Thr(P) residues) in IRS1 from insulin-stimulated cells or human tissues. We investigated a subset of IRS1 Ser(P)/Thr(P) residues using a newly developed panel of 25 phospho-specific monoclonal antibodies (αpS/TmAbIrs1). CHO cells overexpressing the human insulin receptor and rat IRS1 were stimulated with insulin in the absence or presence of inhibitors of the PI3K → Akt → mechanistic target of rapamycin (mTOR) → S6 kinase or MEK pathways. Nearly all IRS1 Ser(P)/Thr(P) residues were stimulated by insulin and significantly suppressed by PI3K inhibition; fewer were suppressed by Akt or mTOR inhibition, and none were suppressed by MEK inhibition. Insulin-stimulated Irs1 tyrosine phosphorylation (Tyr(P)Irs1) was enhanced by inhibition of the PI3K → Akt → mTOR pathway and correlated with decreased Ser(P)-302Irs1, Ser(P)-307Irs1, Ser(P)-318Irs1, Ser(P)-325Irs1, and Ser(P)-346Irs1. Metabolic stress modeled by anisomycin, thapsigargin, or tunicamycin increased many of the same Ser(P)/Thr(P) residues as insulin, some of which (Ser(P)-302Irs1, Ser(P)-307Irs1, and four others) correlated significantly with impaired insulin-stimulated Tyr(P)Irs1. Thus, IRS1 Ser(P)/Thr(P) is an integrated response to insulin stimulation and metabolic stress, which associates with reduced Tyr(P)Irs1 in CHOIR/IRS1 cells.
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regulation of insulin sensitivity by serine threonine phosphorylation of insulin receptor substrate proteins irs1 and IRS2
Diabetologia, 2012Co-Authors: Kyle D Copps, Morris F. WhiteAbstract:The insulin receptor substrate proteins IRS1 and IRS2 are key targets of the insulin receptor tyrosine kinase and are required for hormonal control of metabolism. Tissues from insulin-resistant and diabetic humans exhibit defects in IRS-dependent signalling, implicating their dysregulation in the initiation and progression of metabolic disease. However, IRS1 and IRS2 are regulated through a complex mechanism involving phosphorylation of >50 serine/threonine residues (S/T) within their long, unstructured tail regions. In cultured cells, insulin-stimulated kinases (including atypical PKC, AKT, SIK2, mTOR, S6K1, ERK1/2 and ROCK1) mediate feedback (autologous) S/T phosphorylation of IRS, with both positive and negative effects on insulin sensitivity. Additionally, insulin-independent (heterologous) kinases can phosphorylate IRS1/2 under basal conditions (AMPK, GSK3) or in response to sympathetic activation and lipid/inflammatory mediators, which are present at elevated levels in metabolic disease (GRK2, novel and conventional PKCs, JNK, IKKβ, mPLK). An emerging view is that the positive/negative regulation of IRS by autologous pathways is subverted/co-opted in disease by increased basal and other temporally inappropriate S/T phosphorylation. Compensatory hyperinsulinaemia may contribute strongly to this dysregulation. Here, we examine the links between altered patterns of IRS S/T phosphorylation and the emergence of insulin resistance and diabetes.
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insulin receptor substrates irs1 and IRS2 coordinate skeletal muscle growth and metabolism via the akt and ampk pathways
Molecular and Cellular Biology, 2011Co-Authors: Yun Chau Long, Morris F. White, Kyle D Copps, Zhiyong ChengAbstract:Coordination of skeletal muscle growth and metabolism with nutrient availability is critical for metabolic homeostasis. To establish the role of insulin-like signaling in this process, we used muscle creatine kinase (MCK)-Cre to disrupt expression of insulin receptor substrates Irs1 and IRS2 in mouse skeletal/cardiac muscle. In 2-week-old mice, skeletal muscle masses and insulin responses were slightly affected by Irs1, but not IRS2, deficiency. In contrast, the combined deficiency of Irs1 and IRS2 (MDKO mice) severely reduced skeletal muscle growth and Akt→mTOR signaling and caused death by 3 weeks of age. Autopsy of MDKO mice revealed dilated cardiomyopathy, reflecting the known requirement of insulin-like signaling for cardiac function (P. G. Laustsen et al., Mol. Cell. Biol. 27:1649-1664, 2007). Impaired growth and function of MDKO skeletal muscle were accompanied by increased Foxo-dependent atrogene expression and amino acid release. MDKO mice were resistant to injected insulin, and their isolated skeletal muscles showed decreased insulin-stimulated glucose uptake. Glucose utilization in MDKO mice and isolated skeletal muscles was shifted from oxidation to lactate production, accompanied by an elevated AMP/ATP ratio that increased AMP-activated protein kinase (AMPK)→acetyl coenzyme A carboxylase (ACC) phosphorylation and fatty acid oxidation. Thus, insulin-like signaling via Irs1/2 is essential to terminate skeletal muscle catabolic/fasting pathways in the presence of adequate nutrition.
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irs1 and IRS2 signaling is essential for hepatic glucose homeostasis and systemic growth
Journal of Clinical Investigation, 2006Co-Authors: Xiaocheng Dong, Kyle D Copps, Sunmin Park, Xueying Lin, Morris F. WhiteAbstract:Insulin receptor substrates, including Irs1 and IRS2, integrate insulin and IGF receptor signals with heterologous pathways to coordinate growth and metabolism. Since IRS2 is thought to be especially important in hepatic nutrient homeostasis, we deleted IRS2 [corrected] from hepatocytes of WT mice (called LKO) or genetically insulin-resistant Irs1-/- mice (called LKO::Irs1-/-). Viable LKO::Irs1-/- mice were 70% smaller than WT or LKO mice, and 40% smaller than Irs1-/- mice. Hepatic insulin receptors were functional in all the mice, but insulin signaling via the Akt-FoxO1 pathway was reduced in Irs1-/- and LKO liver, and undetected in LKO::Irs1-/- liver; however, Gsk3beta phosphorylation (Ser9) and hepatic glycogen stores were nearly normal in all of the mice. LKO and Irs1-/- mice developed insulin resistance and glucose intolerance that never progressed to diabetes, whereas LKO::Irs1-/- mice developed hyperglycemia and hyperinsulinemia immediately after birth. Regardless, few hepatic genes changed expression significantly in Irs1-/- or LKO mice, whereas hundreds of genes changed in LKO::Irs1-/- mice--including elevated levels of Pck1, G6pc, Ppargc1, Pparg, and Igfbp1. Thus, signals delivered by Irs1 or IRS2 regulate hepatic gene expression that coordinates glucose homeostasis and systemic growth.
Markus Schubert - One of the best experts on this subject based on the ideXlab platform.
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Neuronal insulin receptor substrate 2 (IRS2) expression is regulated by ZBP89 and SP1 binding to the IRS2 promoter
The Journal of endocrinology, 2009Co-Authors: Michael Udelhoven, Uschi Leeser, Wilhelm Krone, Mareike Pasieka, Markus SchubertAbstract:Since neuronal insulin receptor substrate 2 (IRS2)-mediated signals coordinate key processes in rodent physiology such as food intake, fertility, longevity, and aging-related behavior, we analyzed the mechanisms of neuronal IRS2 expression in neuroblastoma (SHSY5Y) and hypothalamic (GT1-7) cell lines. Using dual luciferase reporter assays and IRS2 promoter deletion constructs, we identified a regulatory cassette within the IRS2 promoter between -779 and -679 bp from the translational start which is responsible for approximately 50% of neuronal IRS2 promoter activity. Chromatin immunoprecipitation assays and electromobility shift assay revealed four overlapping ZBP89/specificity protein 1 (SP1) binding sites which alternatively bind to ZBP89 (ZNF148 as listed in the HUGO Database) or SP1. Activation of this cassette is inhibited by phosphoinositide-3-kinase (PI3K) via increased ZBP89 binding to the promoter. Serum starvation caused increased SP1 binding at one specific SP1 site and decreased binding to another, proving a regulatory interaction between the different binding sites within this promoter cassette to tightly control IRS2 expression. Mutants containing all the possible combinations of one, two, three, or all the four SP1 binding sites of the IRS2 promoter revealed that SP1 binding to one particular site is most important for promoter activation. Stable downregulation of ZBP89 using siRNA substantially increased IRS2 mRNA and protein expression. Thus, alternative binding of ZBP89 or SP1 to the described region in the IRS2 promoter regulates neuronal IRS2 expression in a PI3K-dependent manner.
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Identification of a region in the human IRS2 promoter essential for stress induced transcription depending on SP1, NFI binding and ERK activation in HepG2 cells.
Journal of molecular endocrinology, 2009Co-Authors: Michael Udelhoven, Uschi Leeser, Susanna Freude, Mm Hettich, Matthias Laudes, Jessika Schnitker, Wilhelm Krone, Markus SchubertAbstract:Recent studies have discovered changes in the insulin-/IGF1 signaling affecting glucose metabolism and the molecular pathogenesis of human hepatocellular cancer. Insulin/IGF1 receptor mediates its intracellular effects by recruitment of one out of the four different insulin receptor substrates (IRS). To investigate mechanisms of IRS2 expression, we analyzed transcriptional regulation of IRS2 in human HepG2 cells. We identified a region 688 bp upstream of the translation start codon responsible for w90% of basal human IRS2 promoter activity in HepG2 cells, and confirmed binding of specificity protein 1 (also called Sp1 transcription factor, SP1) and nuclear factor 1 (NFI) in this region. Mutation of both SP1 and NFI binding sites or inhibition of extracellular signal regulated kinase (ERK) suppressed IRS2 promoter activity almost completely, revealing a major role of MAP kinases (MAPK) for IRS2 transcription. Activating this cascade with oxidative stress increased IRS2 promoter activity and endogenous IRS2 expression substantially. IRS2 promoter activity rose even more after additional inhibition of p38MAPK indicating an inhibitory effect of p38MAPK on ERK mediated IRS2 transcription. Activation of the MAPK pathway using interleukin 1, beta (IL1B) increased IRS2 promoter activity similar to oxidative stress. In contrast IL1B decreases and inhibition of the MAPK pathway increases IRS1 promoter activity revealing opposed effects of IL1B and ERK on the expression of different IRS proteins. In conclusion we discovered a specific region (K688 to K611 bp) in the IRS2 promoter essential for basal promoter activity and oxidative stress induced transcription depending on ERK activation and SP1 and NFI binding in human hepatocytes.
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insulin receptor substrate 2 deficiency impairs brain growth and promotes tau phosphorylation
The Journal of Neuroscience, 2003Co-Authors: Markus Schubert, Jake A. Kushner, Deborah J. Burks, Carrie L. Flint, Derek P Brazil, Janet Farhangfallah, Pieter Dikkes, Xavier M Warot, Carlos Rio, Gabriel CorfasAbstract:Insulin resistance and diabetes might promote neurodegenerative disease, but a molecular link between these disorders is unknown. Many factors are responsible for brain growth, patterning, and survival, including the insulin-insulin-like growth factor (IGF)-signaling cascades that are mediated by tyrosine phosphorylation of insulin receptor substrate (IRS) proteins. IRS2 signaling mediates peripheral insulin action and pancreatic β-cell function, and its failure causes diabetes in mice. In this study, we reveal two important roles for IRS2 signaling in the mouse brain. First, disruption of the IRS2 gene reduced neuronal proliferation during development by 50%, which dissociated brain growth from Irs1-dependent body growth. Second, neurofibrillary tangles containing phosphorylated tau accumulated in the hippocampus of old IRS2 knock-out mice, suggesting that IRS2 signaling is neuroprotective. Thus, dysregulation of the IRS2 branch of the insulin-Igf-signaling cascade reveals a molecular link between diabetes and neurodegenerative disease.
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Pdx1 restores β cell function in IRS2 knockout mice
Journal of Clinical Investigation, 2002Co-Authors: Jake A. Kushner, Markus Schubert, Deborah J. Burks, Matthew A. Dow, Carrie L. Flint, Sanjoy Dutta, Marc Montminy, Morris F. WhiteAbstract:The homeodomain transcription factor Pdx1 is required for pancreas development, including the differentiation and function of beta cells. Mutations in Pdx1 or upstream hepatocyte nuclear factors cause autosomal forms of early-onset diabetes (maturity-onset diabetes of the young [MODY]). In mice, the IRS2 branch of the insulin/Igf signaling system mediates peripheral insulin action and pancreatic beta cell growth and function. To investigate whether beta cell failure in IRS2(-/-) mice might be related to dysfunction of MODY-related transcription factors, we measured the expression of Pdx1 in islets from young IRS2(-/-) mice. Before the onset of diabetes, Pdx1 was reduced in islets from IRS2(-/-) mice, whereas it was expressed normally in islets from wild-type or Irs1(-/-) mice, which do not develop diabetes. Whereas male IRS2(-/-)Pdx1(+/+) mice developed diabetes between 8 and 10 weeks of age, haploinsufficiency for Pdx1 caused diabetes in newborn IRS2(-/-) mice. By contrast, transgenic expression of Pdx1 restored beta cell mass and function in IRS2(-/-) mice and promoted glucose tolerance throughout life, as these mice survived for at least 20 months without diabetes. Our results suggest that dysregulation of Pdx1 might represent a common link between ordinary type 2 diabetes and MODY.
