GRK2 Negatively Regulates Glycogen Synthesis in Mouse Liver FL83B Cells

G-protein-coupled receptor (GPCR) kinases (GRKs) are serine/threonine kinases that desensitize agonist-occupied classical GPCRs. Although the insulin receptor (IR) is a tyrosine kinase receptor, the IR also couples to G-proteins and utilizes G-protein signaling components. The present study was designed to test the hypothesis that GRK2 negatively regulates IR signaling. FL83B cells, derived from mouse liver, were treated with insulin and membrane translocation of GRK2 was determined using immunofluoresecence and Western blotting. Insulin caused an increase in the translocation of GRK-2 from cytosol to the plasma membrane. To determine the role of GRK2 in IR signaling, GRK2 was selectively down-regulated (∼by 90%) in FL83B cells using a small interfering RNA technique. Basal as well as insulin-induced glycogen synthesis (measured by d-[U-14C]glucose incorporation) was increased in GRK2-deficient cells compared with control cells. Similarly, GRK2 deficiency increased the basal and insulin-stimulated phosphorylation of Ser21 in glycogen synthase kinase-3α. Insulin-induced tyrosine phosphorylation of the IR was similar in control and GRK2-deficient cells. Basal and insulin-stimulated phosphorylation of Tyr612 in insulin receptor subunit 1 was significantly increased while phosphorylation of Ser307 was decreased in GRK2-deficient FL83B cells compared with control cells. Chronic insulin treatment (24 h) in control cells caused an increase in GRK2 (56%) and a decrease in IR (50%) expression associated with the absence of an increase in glycogen synthesis, suggesting impairment of IR function. However, chronic insulin treatment (24 h) did not decrease IR expression or impair IR effects on glycogen synthesis in GRK2-deficient cells. We conclude that (i) GRK2 negatively regulates basal and insulin-stimulated glycogen synthesis via a post-IR signaling mechanism, and (ii) GRK2 may contribute to reduced IR expression and function during chronic insulin exposure. G-protein-coupled receptor (GPCR) kinases (GRKs) are serine/threonine kinases that desensitize agonist-occupied classical GPCRs. Although the insulin receptor (IR) is a tyrosine kinase receptor, the IR also couples to G-proteins and utilizes G-protein signaling components. The present study was designed to test the hypothesis that GRK2 negatively regulates IR signaling. FL83B cells, derived from mouse liver, were treated with insulin and membrane translocation of GRK2 was determined using immunofluoresecence and Western blotting. Insulin caused an increase in the translocation of GRK-2 from cytosol to the plasma membrane. To determine the role of GRK2 in IR signaling, GRK2 was selectively down-regulated (∼by 90%) in FL83B cells using a small interfering RNA technique. Basal as well as insulin-induced glycogen synthesis (measured by d-[U-14C]glucose incorporation) was increased in GRK2-deficient cells compared with control cells. Similarly, GRK2 deficiency increased the basal and insulin-stimulated phosphorylation of Ser21 in glycogen synthase kinase-3α. Insulin-induced tyrosine phosphorylation of the IR was similar in control and GRK2-deficient cells. Basal and insulin-stimulated phosphorylation of Tyr612 in insulin receptor subunit 1 was significantly increased while phosphorylation of Ser307 was decreased in GRK2-deficient FL83B cells compared with control cells. Chronic insulin treatment (24 h) in control cells caused an increase in GRK2 (56%) and a decrease in IR (50%) expression associated with the absence of an increase in glycogen synthesis, suggesting impairment of IR function. However, chronic insulin treatment (24 h) did not decrease IR expression or impair IR effects on glycogen synthesis in GRK2-deficient cells. We conclude that (i) GRK2 negatively regulates basal and insulin-stimulated glycogen synthesis via a post-IR signaling mechanism, and (ii) GRK2 may contribute to reduced IR expression and function during chronic insulin exposure. The primary biological effect of insulin is maintenance of glucose homeostasis. The effects of insulin are mediated by binding to the insulin receptor, a member of the tyrosine kinase receptor family (1Kasuga M. Zick Y. Blith D.L. Karlsson F.A. Haring H.U. Kahn C.R. J. Biol. Chem. 1982; 257: 9891-9894Abstract Full Text PDF PubMed Google Scholar). The insulin receptor is a heterotetrameric receptor, consisting of two extracellular α and two transmembrane β subunits (2Pilch P.F. Czech M.P. J. Biol. Chem. 1980; 255: 1722-1731Abstract Full Text PDF PubMed Google Scholar). Insulin binding to the α-subunit of insulin receptor causes autophosphorylation of the β-subunit at specific tyrosine residues (3Ullrich A. Bell J.R. Chen E.Y. Herrera R. Petruzzelli L.M. Dull T.J. Gray A. Coussens L. Liao Y.C. Tsubokawa M. Nature. 1985; 313: 756-761Crossref PubMed Scopus (1500) Google Scholar). Autophosphorylation of the β-subunit increases the tyrosine kinase activity of the receptor toward other protein substrates, resulting in the tyrosine phosphorylation of a family of insulin receptor substrate proteins including IRS1, 2The abbreviations used are: IRS, insulin receptor substrate; PI 3-kinase, phosphatidylinositol 3-kinase; GSK3, glycogen synthase kinase 3; GPCR, G-protein-coupled receptor; GRK, G-protein-coupled receptor kinases; IR, insulin receptor; siRNA, small interfering RNA; PBS, phosphate-buffered saline; ANOVA, analysis of variance. 2The abbreviations used are: IRS, insulin receptor substrate; PI 3-kinase, phosphatidylinositol 3-kinase; GSK3, glycogen synthase kinase 3; GPCR, G-protein-coupled receptor; GRK, G-protein-coupled receptor kinases; IR, insulin receptor; siRNA, small interfering RNA; PBS, phosphate-buffered saline; ANOVA, analysis of variance. IRS2, and Shc. The IRS proteins serve as docking proteins for other intracellular proteins containing Src homology 2 domains such as phosphatidylinositol 3-kinase (PI 3-kinase) and Grb2. The binding of these proteins to IRS proteins transmits the signal downstream, leading to the metabolic and mitogenic effects of insulin (4Kahn C.R. White M.F. J. Clin. Investig. 1988; 82: 1151-1161Crossref PubMed Scopus (327) Google Scholar). For example, the interaction of phosphorylated IRS1/2 with PI 3-kinase leads to activation of protein kinase B/Akt, producing many effects such as GLUT4 translocation, protein synthesis, and inactivation of glycogen synthase kinase 3 (GSK3). The GSK3, a serine/threonine kinase, phosphorylates glycogen synthase and inactivates it. As glycogen synthase is a rate-limiting enzyme in glycogen synthesis, inactivation of GSK3 by insulin removes the inhibitory effect of GSK3 on glycogen synthase and leads to increased glycogen synthesis (5Avruch J. Mol. Cell Biochem. 1998; 182: 31-48Crossref PubMed Scopus (319) Google Scholar, 6Virkamaki A. Ueki K. Kahn C.R. J. Clin. Investig. 1999; 103: 931-943Crossref PubMed Scopus (715) Google Scholar, 7Fang X. Yu S.X. Lu Y. Bast C.R. Woodgett J.R. Mills G.B. Proc. Natl. Acad. Sci. U. S. A. 2000; 97: 11960-11965Crossref PubMed Scopus (619) Google Scholar). In addition to signaling via its tyrosine kinase activity, the insulin receptor also couples to the G-proteins Gi and Gq (8Krieger-Brauer H.I. Medda P.K. Kather H. J. Biol. Chem. 1997; 272: 10135-10143Abstract Full Text Full Text PDF PubMed Scopus (145) Google Scholar, 9Imamura T. Huang J. Dalle S. Ugi S. Usui I. Luttrell L.M. Miller W.E. Lefkowitz R.J. Olefsky J.M. J. Biol. Chem. 2001; 276: 43663-43667Abstract Full Text Full Text PDF PubMed Scopus (110) Google Scholar). This IR/G-protein coupling is required for some actions of insulin. For example, insulin resistance develops in adipocytes when Gαi is down-regulated (10Green A. Walters D.J. Belt S.E. Am. J. Physiol. 1997; 273: E254-E261PubMed Google Scholar). Moreover, in 3T3L1 adipocytes IR signaling via Gαq/11 is required for insulin-induced GLUT4 translocation (11Kanzaki M. Watson R.T. Artemyev N.O. Pessin J.E. J. Biol. Chem. 2000; 275: 7167-7175Abstract Full Text Full Text PDF PubMed Scopus (62) Google Scholar). The signaling of receptors via G-proteins is regulated by G protein-coupled receptor kinases (GRKs), a family of serine/threonine kinases that participate in G protein-coupled receptor (GPCR) desensitization. GRK2 can produce GPCR desensitization either by phosphorylation-dependent or phosphorylation-independent mechanisms. Phosphorylation of GPCRs by GRKs results in the uncoupling of GPCRs from heterotrimeric G-proteins and receptor internalization (12Krupnick J.G. Benovic J.L. Annu. Rev. Pharmacol. Toxicol. 1998; 38: 289-319Crossref PubMed Scopus (853) Google Scholar). Recently, GRK2 also has been shown to decrease GPCR signaling by sequestering Gq/11 proteins via their RGS domain (13Pitcher J.A. Freedman N.J. Lefkowitz R.J. Annu. Rev. Biochem. 1998; 67: 653-692Crossref PubMed Scopus (1053) Google Scholar, 14Carman C.V. Parent J.L. Day P.W. Pronin A.N. Sternweis P.M. Wedegaertner P.B. Gilman A.G. Benovic J.L. Kozasa T. J. Biol. Chem. 1999; 274: 34483-34492Abstract Full Text Full Text PDF PubMed Scopus (277) Google Scholar). This phosphorylation-independent mechanism was recently shown to account for the negative regulation of insulin-stimulated glucose transport by GRK2 in 3T3L1 adipocytes (15Usui I. Imamura T. Satoh H. Huang J. Babendure J.L. Hupfeld C.J. Olefsky J.M. EMBO J. 2004; 23: 2821-2829Crossref PubMed Scopus (72) Google Scholar). The present study was designed to determine whether GRK2 modulates insulin receptor signaling and insulin-stimulated glycogen synthesis in mouse liver FL83B cells. Our data show that insulin induces the translocation of GRK2 from cytosol to the plasma membrane. Depleting cells of GRK2 caused an increase in basal as well as insulin-stimulated phosphorylation of GSK3α, leading to increased glycogen synthesis. GRK2 depletion also increased the basal and insulin-induced tyrosine phosphorylation of IRS1 at residue 612, whereas the phosphorylation at serine residue 307 was decreased in GRK2-deficient cells. Finally, we found that chronic insulin treatment (24 h) decreased IR expression and IR-stimulated glycogen synthesis. These changes in IR expression and IR-stimulated glycogen synthesis by chronic insulin treatment were not observed in GRK2-depleted cells. Materials—The FL83B cell line was obtained from American Tissue Culture Collection (Manassas, VA). Anti-GRK2 antibody, anti-insulin receptor β-subunit antibody, and anti-GRK3 antibody were obtained from Santa Cruz Biotechnology Inc. (Santa Cruz, CA). Anti-Cy3 antibody was purchased from Jackson ImmunoResearch Laboratories Inc. (West Grove, PA). Anti-GSK3, anti-pS21 GSK3α, anti-IRS1, anti-pS307 IRS1, and anti-pY612 IRS1 antibodies were purchased from BIOSOURCE (Camarillo, CA). Phosphorylated tyrosine 612 is the pYMXM consensus motif that binds to PI 3-kinase. GRK2 siRNA was purchased from Dharmacon Inc. (Chicago, IL). Oligofectamine, Opti-MEM I, and Dulbecco's modified Eagle's medium were purchased from Invitrogen. d-[U-14C]Glucose was purchased from Amersham Biosciences. Mounting media with 4′,6-diamidino-2-phenylindole was purchased from Vector Laboratories Inc. (Burlingame, CA). All other chemicals were purchased from Sigma. The protein measuring kit was purchased from Pierce. Cell Culture—The experiments were performed in FL83B cells, a hepatocyte cell line derived from mouse. The cells were grown in F12 medium supplemented with 10% calf serum and 1% bacitracin at 37 °C and were performed on cells, that were GRK-2 by was performed using The cells were with phosphate-buffered and medium was GRK2 siRNA and were in Opti-MEM for were and for an at The containing was to the cells and the cells were for medium containing serum and were and the cells were Western of cell was performed using antibody to determine the of For a of siRNA that has at to or was used of the GRK2 siRNA were on and grown to the cells were treated insulin for or with and with for The cells were treated with for with and with for the cells were with 10% serum for at by with primary antibody antibody, in serum and at °C in a The cells were with were in the The cells were with antibody for 1 at the cells were with The were on with containing 4′,6-diamidino-2-phenylindole to the and to in the The cells were using an was used to were and the were using from Inc. cells were treated insulin for or in containing and and for plasma membrane J.R. R. C.J. W.E. J. Cell Biol. PubMed Scopus Google Scholar). the were on for and a The of the resulting cell was to and The were at for at The was and at for at The resulting was in and used as plasma membrane The of the plasma membrane was using cells were treated insulin or or (24 h) at 37 a cell was in of protein were to and to The were with and with primary antibody receptor anti-pS307 IRS1, IRS1, or The were treated with or to and with The were on and the of was by using a The of the antibodies for GRK2 and insulin receptor β-subunit was by using the that in the of the protein not of glycogen in cells was determined using a C.R. J. Biochem. PubMed Scopus Google Scholar). cells were in media with the cells were treated or with insulin for by with d-[U-14C]glucose for 3 at 37 the the cells were with PBS, in and at °C for were for protein and were to the of and the were on for to of The were at for and d-[U-14C]glucose glycogen was determined by synthesis is as of glucose of in plasma and cell was using protein is on the with P.K. Biochem. 1985; PubMed Scopus Google Scholar). by test was used to analysis was using CA). of was of Insulin on GRK2 determine whether insulin induces GRK2 translocation from cytosol to the plasma we and Western The FL83B cells were treated with insulin for or GRK2 was with antibody and antibody and the cells were observed the Insulin treatment increased the translocation of GRK2 from cytosol to the plasma membrane GRK2 also was determined in the plasma membrane by Western blotting. Insulin treatment caused an increase in GRK2 in the plasma membrane by and at and The of the plasma membrane was by the of a antibody a in the whereas was in the plasma membrane not suggesting that the plasma membrane are of of GRK2 siRNA on GRK2 we the GRK2 siRNA technique. of FL83B cells with GRK2 siRNA for decreased the of GRK2 in the cell by This of siRNA was used in GRK2 siRNA did not the expression of a member of GRK2 suggesting that GRK2 siRNA is specific for treatment with used as a effect on the GRK2 expression of Insulin on in and GRK2-deficient treatment increased the of glucose glycogen by of protein basal in control cells, whereas the increase was of protein basal in GRK2-deficient cells The insulin-stimulated increase in glycogen synthesis in cells of protein was similar to that in control cells. However, the basal for glycogen synthesis in GRK2-deficient cells of was the basal in the control and cells that insulin receptor expression was not in GRK2-deficient cells as compared with the control cells of chronic insulin treatment on the expression of GRK2 and insulin liver FL83B cells were treated with insulin for The cell of protein for for was to Western analysis by with antibody and anti-GRK3 FL83B cells and siRNA were with insulin for The cells were in and of protein was to Western analysis using anti-insulin receptor The were and with were by are as significantly from of Insulin on Phosphorylation of and in and GRK2-deficient shown in antibody a the were the was and with of was with The data that the basal phosphorylation of Ser21 at was increased in GRK2 compared with control cells. Insulin treatment and caused an increase in the Ser21 phosphorylation of However, the insulin-stimulated Ser21 phosphorylation of was in GRK2-deficient cells Phosphorylation inactivates GSK3 activity leading to an increase in glycogen synthesis. the phosphorylation and of is with the glycogen synthesis in GRK2-deficient cells. The insulin-stimulated phosphorylation of in GRK2-deficient was similar to that in control cells not of Insulin on Phosphorylation of IR in and GRK2-deficient determine whether the increase in insulin-induced glycogen synthesis in GRK2-deficient cells is to changes in the phosphorylation of the IR, we performed and experiments In the antibody a protein the were the was and with IR antibody, a protein with the of the protein was with IR Insulin treatment increased phosphorylation of IR to a similar in control and GRK2-deficient cells. The basal phosphorylation was not in either cell This that GRK2 not the phosphorylation of IR in of Insulin on and Phosphorylation of IRS1 in and GRK2-deficient determine the phosphorylation of IRS1, we used specific The phosphorylation of on residue increases its with PI 3-kinase and increases insulin signaling D.L. Y. A. 2001; PubMed Scopus Google Scholar). The was with the IRS1 protein Insulin treatment increased Tyr612 phosphorylation of IRS1 in control and GRK2-deficient cells. The increase in IRS1 phosphorylation basal in GRK2-deficient cells was in control cells the basal Tyr612 phosphorylation in control cells was not as as in GRK2-deficient cells, suggesting that GRK2 deficiency also the basal phosphorylation of Tyr612 of We determined the phosphorylation of IRS1 by using a antibody that of The phosphorylation of IRS1 at its with IR and the insulin signaling L. A. A. A. M.F. J. Clin. Investig. 2001; PubMed Scopus Google Scholar). In control cells, the IRS1 is Ser307 phosphorylated at the basal was reduced by insulin Basal Ser307 phosphorylation in GRK2-deficient cells was significantly reduced compared with that in control Insulin treatment did not Ser307 phosphorylation in GRK2-deficient cells of Chronic Insulin on GRK2 and Insulin shown in and insulin treatment for increased GRK2 protein expression by insulin treatment effect on the expression of is a member of the This a effect of chronic insulin treatment on GRK-2 the other chronic insulin treatment caused a in insulin receptor expression in control cells. However, in GRK2-deficient cells, the effect of chronic insulin treatment on IR expression was This a role of GRK2 in insulin-induced regulation of the of Chronic Insulin on in and GRK2-deficient determine whether chronic insulin treatment glycogen synthesis and whether decrease is in GRK2-deficient cells, we performed glycogen synthesis in control and GRK2-deficient cells. In the control the insulin treatment for as significantly increased the glycogen synthesis, whereas the chronic insulin treatment (24 h) to increase glycogen synthesis This desensitization of the insulin a of cells, as A. S. T. A. Rev. PubMed Scopus Google Scholar). However, in cells, chronic treatment with insulin increased glycogen synthesis. This a role of GRK2 in the desensitization of insulin on glycogen synthesis. GPCR is a GRK2 to the plasma membrane and phosphorylates GPCRs at serine/threonine leading to receptor desensitization. GRK2 also binds to the Gαq/11 proteins and signaling the G-protein C.V. Parent J.L. Day P.W. Pronin A.N. Sternweis P.M. Wedegaertner P.B. Gilman A.G. Benovic J.L. Kozasa T. J. Biol. Chem. 1999; 274: 34483-34492Abstract Full Text Full Text PDF PubMed Scopus (277) Google Scholar). GRK2 has been to of the GPCR (12Krupnick J.G. Benovic J.L. Annu. Rev. Pharmacol. Toxicol. 1998; 38: 289-319Crossref PubMed Scopus (853) Google the results of the present study that GRK2 may an inhibitory role in glycogen synthesis by insulin receptor signaling in liver cells. Recently, a similar study that GRK2 negatively regulates insulin-stimulated glucose transport in adipocytes (15Usui I. Imamura T. Satoh H. Huang J. Babendure J.L. Hupfeld C.J. Olefsky J.M. EMBO J. 2004; 23: 2821-2829Crossref PubMed Scopus (72) Google Scholar). The insulin receptor is a member of the tyrosine kinase receptor However, the insulin receptor also G signaling (8Krieger-Brauer H.I. Medda P.K. Kather H. J. Biol. Chem. 1997; 272: 10135-10143Abstract Full Text Full Text PDF PubMed Scopus (145) Google Scholar, 9Imamura T. Huang J. Dalle S. Ugi S. Usui I. Luttrell L.M. Miller W.E. Lefkowitz R.J. Olefsky J.M. J. Biol. Chem. 2001; 276: 43663-43667Abstract Full Text Full Text PDF PubMed Scopus (110) Google Scholar, A. Walters D.J. Belt S.E. Am. J. Physiol. 1997; 273: E254-E261PubMed Google Scholar). In the present study increased GRK2 translocation to the plasma in to a the insulin receptor and two contribute to GRK2 translocation may by a subunit as a of the interaction of the insulin receptor with activation of protein kinase by the insulin receptor GRK2 translocation A. Lefkowitz R.J. PubMed Scopus Google Scholar, L. L. A. M.P. A. J. A. J. Biol. Chem. 1999; 274: Full Text Full Text PDF PubMed Scopus Google Scholar). To determine the of GRK2 translocation in insulin receptor cells were GRK2-deficient and insulin receptor signaling and function were the metabolic effects of we found that GRK2 deficiency caused an increase in basal glycogen synthesis compared with basal in control cells. Insulin a increase in glycogen synthesis basal in GRK2-deficient a similar is observed in control cells. basal glycogen synthesis is increased in GRK2-deficient cells, the that the of insulin-stimulated glycogen synthesis is can in two is that GRK2 deficiency may not insulin receptor function. insulin caused an increase in glycogen synthesis in GRK2-depleted cells that is that observed in control cells of that GRK2 deficiency the metabolic to insulin. GRK2 may liver glycogen synthesis and mechanisms. The increase in glycogen synthesis GRK2 depletion well with the inactivation of synthase kinase phosphorylates and glycogen the enzyme that the of residues basal is an inhibitory on glycogen Phosphorylation of on Ser21 leads to decreased phosphorylation of glycogen This causes a activation of glycogen synthase and increases glycogen X. Yu S.X. Lu Y. Bast C.R. Woodgett J.R. Mills G.B. Proc. Natl. Acad. Sci. U. S. A. 2000; 97: 11960-11965Crossref PubMed Scopus (619) Google Scholar). In the present GRK2 depletion caused a increase in basal Ser21 phosphorylation of was Insulin caused Ser21 phosphorylation of in the GRK2-deficient and control the effect was in the GRK2-deficient that depletion of GRK2 basal of glycogen In addition GRK2 depletion insulin-stimulated glycogen by the of insulin to the inhibitory of on glycogen synthase and glycogen In of the of insulin-stimulated glycogen by GRK2 we were to that GRK2 depletion effect on insulin-stimulated insulin receptor tyrosine GRK2 activation leads to phosphorylation of GPCRs on serine/threonine residues and receptor desensitization J.A. Lefkowitz R.J. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). phosphorylation of the insulin receptor causes a decrease in phosphorylation of insulin receptor and a decrease in receptor activity S. J.A. Biochem. 2001; PubMed Scopus Google Scholar, S. White M.F. Kahn C.R. J. Biol. Chem. 1988; Full Text PDF PubMed Google Scholar). we that the GRK2 depletion serine phosphorylation of the insulin receptor, phosphorylation of the insulin receptor, and insulin receptor signaling. However, insulin-stimulated phosphorylation was similar in control and GRK2-deficient cells. This that GRK2 may not insulin receptor signaling by phosphorylation of the insulin the changes in basal and insulin-induced glycogen synthesis by GRK2 depletion may from effects on receptor signaling such as suggesting that the function of IRS1 by GRK2 was obtained in GRK2-deficient cells. We observed that the phosphorylation of IRS1 at Tyr612 was whereas phosphorylation at Ser307 was decreased by GRK2 The increase in Tyr612 phosphorylation of IRS1 was observed basal and insulin-stimulated Tyr612 phosphorylation of IRS1 also was IRS1 phosphorylation at the binding of the subunit of PI 3-kinase, leading to activation of P.K. Biochem. 1985; PubMed Scopus Google Scholar). In to phosphorylation of Ser307 of was reduced by GRK2 This phosphorylation of IRS1 at Ser307 the of insulin on of IRS1 PubMed Scopus Google Scholar, I. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). For example, Ser307 phosphorylation of IRS1 IRS1 with the insulin receptor, phosphorylation on IRS1 and negatively regulates insulin signaling D.L. Y. A. 2001; PubMed Scopus Google Scholar). the results of the present study the hypothesis that GRK2 phosphorylation of Ser307 on IRS1, insulin interaction and insulin receptor phosphorylation of Tyr612 on In GRK2 negatively insulin receptor signaling, in the basal and in the of insulin. The of GRK2 on insulin receptor signaling IRS phosphorylation is to the of phosphorylation of is observed in of with 2 K. M. S. J. M. H. PubMed Scopus Google Scholar). However, is whether GRK2 IRS1 or whether GRK2 phosphorylates signaling that is for phosphorylation of via negative regulation also phosphorylates IRS1 at serine residues and insulin receptor signaling H. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). the decrease in activity observed in GRK2-deficient cells in reduced phosphorylation of IRS1 and contribute to the increase in IRS1 phosphorylation that we In addition to the kinase activity, GRK2 also GPCR signaling by binding to Gαq/11 protein and from the signal Insulin receptor is to to Gαq/11 and Gαi The Gαq/11 subunit is required for insulin signaling in 3T3L1 adipocytes and of Gαi leads to insulin resistance in K. M. S. J. M. H. PubMed Scopus Google Scholar). a study that GRK-2 in 3T3L1 adipocytes insulin-induced GLUT4 translocation by binding to Gαq/11 (15Usui I. Imamura T. Satoh H. Huang J. Babendure J.L. Hupfeld C.J. Olefsky J.M. EMBO J. 2004; 23: 2821-2829Crossref PubMed Scopus (72) Google Scholar). This was of GRK2 kinase activity and did not phosphorylation of the insulin receptor, in that GRK2 deficiency did not basal or insulin-induced of in the present study was the of GRK2 on the to chronic insulin receptor Chronic insulin the insulin receptors and insulin receptor signaling. Moreover, shown that chronic is associated with reduced insulin receptor in and A. S. T. A. Rev. PubMed Scopus Google Scholar). In with we observed that chronic insulin treatment decreased insulin receptor expression and was not associated with increased glycogen synthesis, suggesting insulin receptor desensitization. These were by increased expression of In GRK2-deficient cells chronic insulin treatment effect on insulin receptor expression and was associated with increased glycogen synthesis. GRK2-deficient cells an increase basal glycogen synthesis, contribute to the increase in glycogen synthesis insulin treatment in these cells. However, the absence of insulin receptor in GRK2-deficient cells that GRK2 may a role in insulin receptor expression and desensitization during insulin exposure. is that two the of the insulin receptor chronic insulin exposure. desensitization of the insulin receptor signal mediated in by the receptor mechanism IRS1 phosphorylation as The a regulation of receptor synthesis or leading to reduced receptor with chronic insulin Our of GRK2 expression with chronic insulin treatment is in that expression of GRK2 also is in such as and Moreover, of GRK2 in has to of S. PubMed Scopus Google Scholar, Scopus Google Scholar). GRK2 expression also is observed in from a of insulin resistance and M. M.F. Am. J. Physiol. PubMed Scopus Google Scholar). of GRK2 to chronic insulin receptor may contribute to such as insulin resistance and to the of GRK2 in of and to of GRK2 the In we a the of GRK2 and and glycogen synthesis in liver cells. of GRK2 IRS1 phosphorylation and increases IRS1 in to insulin receptor activation by is with a in insulin receptor function in of glycogen synthesis. Chronic insulin treatment causes in GRK2 may a role in the insulin receptor desensitization and observed in cells insulin exposure. as the present the an inhibitory role of GRK2 in the regulation of glycogen synthesis in liver cells, are to the of with