Basic Fibroblast Growth Factor Utilizes Both Types of Component Subunits of Gs for Dual Signaling in Human Adipocytes

Abstract

Basic fibroblast growth factor (bFGF), a ligand of receptor protein-tyrosine kinases, promoted the dissociation of Gs and had antagonistic stimulatory and inhibitory effects on adenylyl cyclase and NADPH oxidase in human fat cell plasma membranes. The bFGF-induced activation of adenylyl cyclase was blocked by COOH-terminal anti-Gαs, indicating that it was mediated by Gαs. The inhibitory action of bFGF was mimicked by exogenously supplied Gβγ-subunits and was reversed by anti-Gβ1/2, or βARK-CT, a COOH-terminal β-adrenergic receptor kinase fragment that specifically binds free Gβγ, indicating that it was transduced by Gβγ complexes. The bFGF-induced inhibition of NADPH-dependent H2O2 generation was also reversed by peptide 100–119, an inhibitor of Gs activation by ligand-occupied β-adrenergic receptors, indicating that the Gβγ complexes mediating the inhibitory action of the growth factor are derived from Gs. The findings suggest a direct, non-kinase-dependent, coupling of bFGF receptor(s) to Gs and provide the first example of a ligand of receptor protein-tyrosine kinases that is capable of utilizing both types of component subunits of a single heterotrimeric G protein for dual signaling in a single cell type. Basic fibroblast growth factor (bFGF), a ligand of receptor protein-tyrosine kinases, promoted the dissociation of Gs and had antagonistic stimulatory and inhibitory effects on adenylyl cyclase and NADPH oxidase in human fat cell plasma membranes. The bFGF-induced activation of adenylyl cyclase was blocked by COOH-terminal anti-Gαs, indicating that it was mediated by Gαs. The inhibitory action of bFGF was mimicked by exogenously supplied Gβγ-subunits and was reversed by anti-Gβ1/2, or βARK-CT, a COOH-terminal β-adrenergic receptor kinase fragment that specifically binds free Gβγ, indicating that it was transduced by Gβγ complexes. The bFGF-induced inhibition of NADPH-dependent H2O2 generation was also reversed by peptide 100–119, an inhibitor of Gs activation by ligand-occupied β-adrenergic receptors, indicating that the Gβγ complexes mediating the inhibitory action of the growth factor are derived from Gs. The findings suggest a direct, non-kinase-dependent, coupling of bFGF receptor(s) to Gs and provide the first example of a ligand of receptor protein-tyrosine kinases that is capable of utilizing both types of component subunits of a single heterotrimeric G protein for dual signaling in a single cell type. fibroblast growth factor basic fibroblast growth factor guanosine 5′-O-(3-thiotriphosphate) guanyl 5′-(β,γ-imino)triphosphate 3-(N-morpholino)propanesulfonic acid 2-(N-morpholino)ethanesulfonic acid glutathioneS-transferase β-adrenergic receptor kinase guanosine 5′-O-2-(thio)diphosphate The fibroblast growth factors (FGFs)1 are a family of heparin-binding growth factors and oncogenes with at least 20 members (1Fernig D.G. Gallagher J.T. Prog. Growth Factor Res. 1994; 5: 353-377Abstract Full Text PDF PubMed Scopus (178) Google Scholar, 2Burke D. Wilkes D. Blundell T.L. Malcolm S. Trends Biochem. Sci. 1998; 23: 59-62Abstract Full Text PDF PubMed Scopus (153) Google Scholar, 3Szebenyi G. Fallon J.S. Int. Rev. Cytol. 1999; 185: 45-106Crossref PubMed Google Scholar). Basic fibroblast growth factor (bFGF, FGF-2) or a related heparin-binding peptide is produced by white and brown adipocytes (4Teichert-Kuliszewska K. Hamilton B.S. Deitel M. Roncari D.A.K. J. Clin. Invest. 1992; 90: 1226-1231Crossref PubMed Scopus (36) Google Scholar,5Yamashita H. Sato N. Kizaki T. Oh-ishi S. Segawa M. Saitoh D. Ohira Y. Ohno H. Cell Growth Differ. 1995; 6: 1457-1462PubMed Google Scholar). The growth factor stimulates the proliferation of preadipocytes and inhibits their conversion to adipocytes. The expression of bFGF is decreased appreciably during adipose differentiation and is increased in obesity (4Teichert-Kuliszewska K. Hamilton B.S. Deitel M. Roncari D.A.K. J. Clin. Invest. 1992; 90: 1226-1231Crossref PubMed Scopus (36) Google Scholar). Interestingly, another member of the FGF family, aFGF (FGF-1), accelerated the conversion of 3T3 L1-preadipocytes to adipocytes in the presence of insulin and was adipogenic in itself (6Krieger-Brauer H.I. Kather H. Biochem. J. 1995; 307: 549-556Crossref PubMed Scopus (119) Google Scholar). The members of the fibroblast growth factor family interact with cell surface low affinity heparan sulfate that facilitates binding to their receptors (1Fernig D.G. Gallagher J.T. Prog. Growth Factor Res. 1994; 5: 353-377Abstract Full Text PDF PubMed Scopus (178) Google Scholar, 2Burke D. Wilkes D. Blundell T.L. Malcolm S. Trends Biochem. Sci. 1998; 23: 59-62Abstract Full Text PDF PubMed Scopus (153) Google Scholar, 3Szebenyi G. Fallon J.S. Int. Rev. Cytol. 1999; 185: 45-106Crossref PubMed Google Scholar, 7Yayon A. Klagsbrun M. Esko J.D. Leder P. Ornitz D.M. Cell. 1991; 64: 841-848Abstract Full Text PDF PubMed Scopus (2075) Google Scholar). These consist of a family of high affinity receptor tyrosine kinases (FGF receptors 1–4) displaying overlapping affinities for the various FGFs (1Fernig D.G. Gallagher J.T. Prog. Growth Factor Res. 1994; 5: 353-377Abstract Full Text PDF PubMed Scopus (178) Google Scholar, 2Burke D. Wilkes D. Blundell T.L. Malcolm S. Trends Biochem. Sci. 1998; 23: 59-62Abstract Full Text PDF PubMed Scopus (153) Google Scholar, 3Szebenyi G. Fallon J.S. Int. Rev. Cytol. 1999; 185: 45-106Crossref PubMed Google Scholar, 7Yayon A. Klagsbrun M. Esko J.D. Leder P. Ornitz D.M. Cell. 1991; 64: 841-848Abstract Full Text PDF PubMed Scopus (2075) Google Scholar). Alternative splicing generates isoforms of receptors 1–3 that exhibit unique binding characteristics (1Fernig D.G. Gallagher J.T. Prog. Growth Factor Res. 1994; 5: 353-377Abstract Full Text PDF PubMed Scopus (178) Google Scholar, 2Burke D. Wilkes D. Blundell T.L. Malcolm S. Trends Biochem. Sci. 1998; 23: 59-62Abstract Full Text PDF PubMed Scopus (153) Google Scholar, 3Szebenyi G. Fallon J.S. Int. Rev. Cytol. 1999; 185: 45-106Crossref PubMed Google Scholar). In addition, an unrelated cysteine-rich transmembrane protein of unknown function has also been identified as a high affinity receptor for FGFs (8Burrus L.W. Zuber M.E. Luedecke B.A. Olwin B.B. Mol. Cell. Biol. 1992; 12: 5600-5609Crossref PubMed Scopus (104) Google Scholar, 9Mason I.J. Cell. 1994; 78: 547-552Abstract Full Text PDF PubMed Scopus (525) Google Scholar). Mature human adipocytes and 3T3 L1-preadipocytes contain a plasma membrane-bound H2O2 generating system that is under antagonistic control by various hormones, growth factors, and cytokines, including ligands of receptor protein-tyrosine kinases, such as insulin and various isoforms of PDGF and FGF (6Krieger-Brauer H.I. Kather H. Biochem. J. 1995; 307: 549-556Crossref PubMed Scopus (119) Google Scholar, 10Krieger-Brauer H.I. Kather H. J. Clin. Invest. 1992; 89: 1006-1013Crossref PubMed Scopus (184) Google Scholar, 11Krieger-Brauer H.I. Kather H. Biochem. J. 1995; 307: 543-548Crossref PubMed Scopus (67) Google Scholar, 12Krieger-Brauer H.I. Medda P.K. Kather H. J. Biol. Chem. 1997; 272: 10135-10143Abstract Full Text Full Text PDF PubMed Scopus (147) Google Scholar, 13Krieger-Brauer H.I. Medda P.K. Hebling U. Kather H. J. Biol. Chem. 1999; 274: 28308-28313Abstract Full Text Full Text PDF PubMed Scopus (13) Google Scholar, 14Krieger-Brauer H.I. Medda P.K. Sattel B. Kather H. J. Biol. Chem. 2000; 275: 2486-2490Abstract Full Text Full Text PDF PubMed Scopus (12) Google Scholar). Recent work revealed that the stimulatory effect of insulin on NADPH-dependent H2O2 generation is transduced by a G protein (Gαi2), whereas the inhibitory action of the β-adrenergic agonist, isoproterenol, which signals through a prototypical Gs-coupled receptor, is mediated by Gβγ derived from Gs (13Krieger-Brauer H.I. Medda P.K. Hebling U. Kather H. J. Biol. Chem. 1999; 274: 28308-28313Abstract Full Text Full Text PDF PubMed Scopus (13) Google Scholar, 14Krieger-Brauer H.I. Medda P.K. Sattel B. Kather H. J. Biol. Chem. 2000; 275: 2486-2490Abstract Full Text Full Text PDF PubMed Scopus (12) Google Scholar). In this work we examined whether the inhibitory effect of bFGF, another ligand of tyrosine kinase receptors, on NADPH-dependent H2O2 generation is also transmitted by a G protein. Antibodies directed against the C terminus of Gαs(RM/1), common anti-Gβ (SW/1), and anti-Gβ1/2 were from PerkinElmer Life Sciences, Bad Homburg, Germany (RM/1 and SW/1), and Upstate Biotechnology Inc., Lake Placid, NY, respectively. Recombinant Gαi2, Gαs, and protein A-agarose were from Calbiochem. Hybond polyvinylidene difluoride membranes were obtained from Amersham Pharmacia Biotech-Buchler, Braunschweig, Germany. Insulin, GTPγS, GDPβS, and GTP were from Roche Molecular Biochemicals, Mannheim, Germany. Isoproterenol and rbFGF were from Sigma AG, München, Germany. Human albumin and luminol were obtained from Behring Werke, Marburg, Germany. The GST fusion protein containing a carboxyl-terminal fragment (residues 546–670) of the β-adrenergic receptor kinase (βARK 1CT-GST) and the corresponding GST protein were a gift of Dr. R. Lefkowitz, Howard Hughes Medical Institute, Duke University Medical Center, Durham, NC (16Koch W.J. Inglese J. Stone W.C. Lefkowitz R.J. J. Biol. Chem. 1993; 268: 8256-8260Abstract Full Text PDF PubMed Google Scholar). Experimental details have been described in detail elsewhere (5Yamashita H. Sato N. Kizaki T. Oh-ishi S. Segawa M. Saitoh D. Ohira Y. Ohno H. Cell Growth Differ. 1995; 6: 1457-1462PubMed Google Scholar, 9Mason I.J. Cell. 1994; 78: 547-552Abstract Full Text PDF PubMed Scopus (525) Google Scholar, 10Krieger-Brauer H.I. Kather H. J. Clin. Invest. 1992; 89: 1006-1013Crossref PubMed Scopus (184) Google Scholar, 11Krieger-Brauer H.I. Kather H. Biochem. J. 1995; 307: 543-548Crossref PubMed Scopus (67) Google Scholar, 12Krieger-Brauer H.I. Medda P.K. Kather H. J. Biol. Chem. 1997; 272: 10135-10143Abstract Full Text Full Text PDF PubMed Scopus (147) Google Scholar). Briefly, adipose tissue was from nondiabetic subjects undergoing elective abdominal or cosmetic breast surgery. The specimens were cut into small pieces, and fat cells were isolated by the method of Rodbell (17Rodbell M. J. Biol. Chem. 1964; 239: 375-384Abstract Full Text PDF PubMed Google Scholar) in a HEPES-buffered Krebs-Henseleit solution, pH 7.4, containing 20 mm HEPES, 10 mmNaHCO3, 5 mm glucose, 20 g/liter albumin, and 1 mg/ml collagenase (CLS, Worthington). After 30 min, fat cells were washed and re-suspended in 10 volumes of an ice-cold lysing medium containing 20 mm MES, pH 6.0, 2 mmMgCl2, 1 mm CaCl2, 5 mmKCl, and 100 mg/liter soybean trypsin inhibitor. Mechanical shaking completed cell lysis, and fat cell ghosts were collected by low speed centrifugation (1,000 × g, 4 °C, 20 min). Similar to other effector systems, such as adenylyl cyclases, it is difficult to assess the effects of inhibitory ligands on NADPH-dependent H2O2 generation under basal conditions, where enzyme activity is low. Since direct activators of NADPH oxidase are presently unknown, enzyme activity was increased by insulin, which stimulates NADPH oxidase via Gαi2 (12Krieger-Brauer H.I. Medda P.K. Kather H. J. Biol. Chem. 1997; 272: 10135-10143Abstract Full Text Full Text PDF PubMed Scopus (147) Google Scholar). A two-step procedure was used, as reported elsewhere (6Krieger-Brauer H.I. Kather H. Biochem. J. 1995; 307: 549-556Crossref PubMed Scopus (119) Google Scholar, 10Krieger-Brauer H.I. Kather H. J. Clin. Invest. 1992; 89: 1006-1013Crossref PubMed Scopus (184) Google Scholar, 11Krieger-Brauer H.I. Kather H. Biochem. J. 1995; 307: 543-548Crossref PubMed Scopus (67) Google Scholar, 12Krieger-Brauer H.I. Medda P.K. Kather H. J. Biol. Chem. 1997; 272: 10135-10143Abstract Full Text Full Text PDF PubMed Scopus (147) Google Scholar, 13Krieger-Brauer H.I. Medda P.K. Hebling U. Kather H. J. Biol. Chem. 1999; 274: 28308-28313Abstract Full Text Full Text PDF PubMed Scopus (13) Google Scholar, 14Krieger-Brauer H.I. Medda P.K. Sattel B. Kather H. J. Biol. Chem. 2000; 275: 2486-2490Abstract Full Text Full Text PDF PubMed Scopus (12) Google Scholar). Plasma membranes from untreated or insulin-treated cells were first exposed to hormones and growth factors and were then assayed for NADPH oxidase activity. The activation step was carried out in 30 mm MOPS, pH 7.5, containing 120 mm KCl, CaCl2, mm 10 mm and human were first with various of hormones and growth factors for 5 to receptor were as After 20 min, ghosts were collected by and then re-suspended in 30 mm MES, pH containing 120 mm 4 mm 1 mm and 10 for of NADPH-dependent H2O2 were by of NADPH and 30 at by to a of 100 H.I. Kather H. J. Clin. Invest. 1992; 89: 1006-1013Crossref PubMed Scopus (184) Google Scholar). assess the effects of G protein on NADPH-dependent H2O2 membranes were first exposed to at for and were then to the two-step procedure described In to other G in the presence of under the in the M. S. 1994; PubMed Scopus Google Scholar, M. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). Plasma membranes were in of pH containing 120 mm KCl, mm mm CaCl2, and human albumin in the or presence of bFGF for 5 at was and were for another The membranes were by centrifugation and in for 30 at by an directed against the terminus of which heterotrimeric and M. S. 1994; PubMed Scopus Google Scholar, M. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). were carried out in 10 mm pH 7.4, containing mm 1 mm 1 mm mm 2 2 and 1 a of and The of of were at 4 The complexes were with protein A-agarose and washed with 1 of mm pH 7.4, containing mm 1 mm 1 mm and were by and to polyvinylidene difluoride membranes. was and anti-Gβ Life were by the from Amersham Pharmacia and by The of was to the of for cyclase activity of human fat cell plasma membranes was in mm pH 7.5, containing 1 mm mm mm 10 10 and albumin in a of 100 T. T. M. J. Biol. Chem. Full Text PDF PubMed Google Scholar). were by of of protein and for at in the or presence of or bFGF, as were by 100 of ice-cold acid was by of isolated adipocytes were in HEPES-buffered Krebs-Henseleit solution, pH 7.4, containing mm human albumin, and or bFGF, as for at in a of were by an of ice-cold and was as described 1 A the effects of bFGF on NADPH-dependent H2O2 generation in the presence of of in membranes from cells that had been with 10 Similar to with the adenylyl where activation by to on the of activation was had effect under the J. M. Sci. U. S. A. PubMed Scopus Google Scholar). The inhibitory action of bFGF at of 5 and was at 10 a of NADPH-dependent H2O2 generation was by in the presence of 1 a for bFGF in the presence of a of The bFGF-induced inhibition of NADPH-dependent H2O2 generation was at 1 and was at 30 Recent work that the effects of another inhibitor of NADPH-dependent H2O2 the β-adrenergic isoproterenol, are transduced by subunits derived from Gs (13Krieger-Brauer H.I. Medda P.K. Hebling U. Kather H. J. Biol. Chem. 1999; 274: 28308-28313Abstract Full Text Full Text PDF PubMed Scopus (13) Google Scholar, 14Krieger-Brauer H.I. Medda P.K. Sattel B. Kather H. J. Biol. Chem. 2000; 275: 2486-2490Abstract Full Text Full Text PDF PubMed Scopus (12) Google Scholar). the was that the effects of bFGF were also mediated by the inhibitory action of bFGF was reversed by an directed against an of subunits which had on basal or of NADPH-dependent H2O2 generation 2 effects were at an of in the presence of bFGF 2 The inhibitory effect of bFGF also reversed by 10 a fragment of the β-adrenergic receptor kinase that specifically binds free Gβγ (16Koch W.J. Inglese J. Stone W.C. Lefkowitz R.J. J. Biol. Chem. 1993; 268: 8256-8260Abstract Full Text PDF PubMed Google Scholar). of the bFGF-induced of NADPH-dependent H2O2 generation was at effects were at 10 2 The GST protein to the inhibitory effects of and bFGF the effects of bFGF, and insulin on Gs In to other heterotrimeric G Gs in the presence of under the M. S. 1994; PubMed Scopus Google Scholar, M. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). Plasma membranes were with in the or presence of bFGF, or 5 of Gs into component subunits was by an utilizing which heterotrimeric and Gαs. that the of in was in membranes that had been with bFGF, aFGF and insulin to the of Gs. In the of GTPγS, bFGF had on the of Gs. the growth factor a dissociation of Gs which is for a The that bFGF also adenylyl which was in the the effects of bFGF and on and NADPH-dependent H2O2 generation in human fat cell plasma membranes. bFGF a of that at that it was transmitted by receptors 4 with the β-adrenergic isoproterenol, which a in bFGF was to a activation of adenylyl cyclase in fat cell plasma membranes 4 whereas both were of in NADPH-dependent H2O2 generation 4 The of and bFGF to in where a bFGF-induced in was in the presence of a inhibitor the were by 10 bFGF under conditions, a that to a in the presence of 1 isoproterenol, with in 3T3 cells M. K. P. Sci. U. S. A. 2000; PubMed Scopus Google Scholar). 5 the effects of a COOH-terminal a the of and a of on the bFGF-induced of adenylyl cyclase activity and the inhibitory action of the growth factor on of NADPH-dependent H2O2 the used, both the COOH-terminal and the peptide the stimulatory effect of bFGF on the inhibitory effect of bFGF on NADPH-dependent H2O2 generation was reversed by peptide whereas the COOH-terminal had The effects of the peptide were in as as a peptide derived from had on the antagonistic effects of bFGF on and H2O2 In addition, the peptide the stimulatory effect of insulin, which is mediated via In this it is that bFGF, a ligand of receptor protein-tyrosine kinases, stimulates adenylyl cyclase and inhibits NADPH-dependent H2O2 generation in human fat cell plasma membranes. The by which bFGF antagonistic stimulatory and inhibitory effects on adenylyl cyclase and NADPH oxidase were to the plasma of and in the of indicating that of including the tyrosine kinase activity of bFGF receptor(s) were with the that bFGF promoted dissociation of Gs findings that FGF receptor(s) are to via a human adipocytes at least members of the FGF receptor family, and isoforms with and affinity on to directed against their carboxyl-terminal and both to via Gs. Antibodies directed against the of subunits and corresponding to their are to receptor and and are in at protein coupling G. J. Biol. Chem. 1995; Full Text Full Text PDF PubMed Scopus Google Scholar, N. K. P. G. J. J. Biol. Chem. 1997; 272: Full Text Full Text PDF PubMed Scopus Google Scholar, M. J. Biol. Chem. 1993; 268: Full Text PDF PubMed Google Scholar, J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). The stimulatory effect of bFGF on adenylyl cyclase blocked by COOH-terminal and a peptide corresponding to indicating that it was in transmitted by Gαs. The bFGF-induced inhibition of NADPH-dependent H2O2 generation also the of a G protein. In to stimulatory effects on adenylyl the inhibitory action of bFGF to mediated via Gβγ on that this effect was specifically reversed by anti-Gβ or a COOH-terminal β-adrenergic receptor kinase fragment that free Gβγ, and was mimicked by exogenously supplied Gβγ subunits H.I. Medda P.K. Sattel B. Kather H. J. Biol. Chem. 2000; 275: 2486-2490Abstract Full Text Full Text PDF PubMed Scopus (12) Google Scholar, R. B. J. 1997; PubMed Scopus Google Scholar). In to the stimulatory effect of bFGF on adenylyl inhibitory action on NADPH-dependent H2O2 generation was by COOH-terminal A for this that activation of Gs by bFGF receptors or subunits to for inhibition of NADPH-dependent H2O2 as has been for the of adenylyl cyclase R. W.J. J. Biol. Chem. 1994; Full Text PDF PubMed Google Scholar, T. R. T. J. Biol. Chem. 1998; Full Text Full Text PDF PubMed Scopus Google Scholar). the effects of COOH-terminal on are and on receptors, G and cell types (13Krieger-Brauer H.I. Medda P.K. Hebling U. Kather H. J. Biol. Chem. 1999; 274: 28308-28313Abstract Full Text Full Text PDF PubMed Scopus (13) Google Scholar, 14Krieger-Brauer H.I. Medda P.K. Sattel B. Kather H. J. Biol. Chem. 2000; 275: 2486-2490Abstract Full Text Full Text PDF PubMed Scopus (12) Google Scholar, G. J. Biol. Chem. 1995; Full Text Full Text PDF PubMed Scopus Google Scholar, N. K. P. G. J. J. Biol. Chem. 1997; 272: Full Text Full Text PDF PubMed Scopus Google Scholar, M. J. Biol. Chem. 1993; 268: Full Text PDF PubMed Google Scholar, J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). the effects of COOH-terminal to the of the Gβγ complexes the inhibitory action of we have that Gs is capable of Gβγ subunits to for the inhibition of NADPH-dependent H2O2 generation in the presence of H.I. Medda P.K. Sattel B. Kather H. J. Biol. Chem. 2000; 275: 2486-2490Abstract Full Text Full Text PDF PubMed Scopus (12) Google Scholar). bFGF was in adenylyl both were of in of NADPH-dependent H2O2 is that the effect of bFGF on both is an the adenylyl cyclase whereas NADPH oxidase activity was in the presence of the of bFGF and to as of adenylyl cyclase under and was in indicating that it was A for this that β-adrenergic receptors Gs for signaling the receptor(s) for that complexes derived from G other Gs in bFGF receptor it has been reported that bFGF receptor(s) also to G in cell types G. J. Biol. Chem. 1994; Full Text PDF PubMed Google Scholar, Olwin B.B. Mol. Cell. Biol. 1998; PubMed Scopus Google Scholar). the of the Gβγ complexes the inhibitory effects of bFGF, we a peptide of Recent work that this peptide the antagonistic effects of the β-adrenergic on adenylyl cyclase and NADPH oxidase that are transmitted by and (13Krieger-Brauer H.I. Medda P.K. Hebling U. Kather H. J. Biol. Chem. 1999; 274: 28308-28313Abstract Full Text Full Text PDF PubMed Scopus (13) Google Scholar, 14Krieger-Brauer H.I. Medda P.K. Sattel B. Kather H. J. Biol. Chem. 2000; 275: 2486-2490Abstract Full Text Full Text PDF PubMed Scopus (12) Google Scholar). The revealed that peptide reversed the effects of bFGF on NADPH-dependent H2O2 generation and as The peptide the stimulatory effect of insulin, which is mediated by In addition, a peptide derived from had on the antagonistic effects of bFGF on and H2O2 indicating that peptide is for Gαs, and the of receptors with other G the the that the subunits mediating the bFGF-induced inhibition of NADPH-dependent H2O2 generation were derived from Gs. G are to it is that G for signals of other types of receptors as including receptor from the stimulatory effect of insulin on NADPH-dependent H2O2 which is mediated by Gαi2 (12Krieger-Brauer H.I. Medda P.K. Kather H. J. Biol. Chem. 1997; 272: 10135-10143Abstract Full Text Full Text PDF PubMed Scopus (147) Google activation of adenylyl cyclase by growth factor through and of fibroblast by via B. G. J. Biol. Chem. Full Text PDF PubMed Google Scholar, R. J. J. Biol. Chem. 1993; 268: Full Text PDF PubMed Google Scholar). 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Chem. 2000; 275: 2486-2490Abstract Full Text Full Text PDF PubMed Scopus (12) Google the by which Gαi2 the stimulatory effects of and the activity of the human fat cell oxidase to that activation of heterotrimeric G by complexes is by of GTP for on the and this is to dissociation into and subunits J. Biol. Chem. 1998; Full Text Full Text PDF PubMed Scopus Google Cell. 1995; Full Text PDF PubMed Scopus Google Scholar). G are then by or both J. Biol. Chem. 1998; Full Text Full Text PDF PubMed Scopus Google Scholar, Cell. 1995; Full Text PDF PubMed Scopus Google Scholar). the by which insulin and bFGF NADPH-dependent H2O2 generation were to the plasma and were of it that NADPH oxidase of action or of Gαi2 and to the out S. J. 1995; PubMed Scopus Google Scholar, K. Y. M. T. J. 1997; 275: PubMed Scopus Google Scholar, T. M. J. Biol. Chem. Full Text PDF PubMed Google Scholar). The generation of H2O2 and other to a common signaling for hormones and that cell growth and differentiation S. J. 1995; PubMed Scopus Google Scholar, K. Y. M. T. J. 1997; 275: PubMed Scopus Google Scholar, T. M. J. Biol. Chem. Full Text PDF PubMed Google Scholar, Biol. 1995; PubMed Scopus Google Scholar, J. Biol. Chem. 1997; 272: Full Text Full Text PDF PubMed Scopus Google Scholar). work from this revealed that growth factors that NADPH-dependent H2O2 generation promoted in 3T3 whereas inhibitory such as bFGF, were (6Krieger-Brauer H.I. Kather H. Biochem. J. 1995; 307: 549-556Crossref PubMed Scopus (119) Google Scholar). it has been reported that inhibition of mediated via a Gs that 1992; PubMed Scopus Google Scholar). The that the of bFGF, an inhibitor of is transduced by subunits that are derived from Gs is with and in of the that the H2O2 produced in to hormones, growth factors, and in the of differentiation and of the The of the bFGF-induced of is In a of by bFGF was in the presence of a inhibitor. In addition, that bFGF has activity in human that in is by in that bFGF has effects on capable of the effects of other of adenylyl cyclase via activation of Gs J. S. Cell. PubMed Scopus Google Scholar). In the findings that bFGF, a ligand of receptor protein-tyrosine kinases, is capable of utilizing as as for dual signaling in a single cell via a and suggest a of or members of the FGF receptor family with Gs. These and that the human fat cell oxidase is under antagonistic control by and Gβγ subunits derived from G and provide in of the that this membrane-bound system an effector system for hormones, growth factors, and ligand binding to cell surface receptors to in the are to Sattel and Hebling for and Dr. G. for G protein and Dr. R. Lefkowitz, Duke University Medical Center, Durham, for the GST fusion protein containing a carboxyl-terminal fragment of the β-adrenergic receptor kinase (βARK 1CT-GST) and the corresponding GST protein.