Selective Control of Skeletal Muscle Differentiation by Akt1

The phosphatidylinositol 3-kinase-Akt pathway plays a central role in growth, development, and metabolism in both normal and neoplastic cells. In skeletal muscle, Akt has been implicated in regulating regeneration and hypertrophy and in counteracting atrophy. Here we provide evidence that Akt1 and not Akt2 is essential for muscle differentiation. Using a robust model of MyoD-mediated muscle development, in which dominant-negative Akt blocked differentiation, we show that targeted loss of Akt1 was equally inhibitory. Selective elimination of Akt1 had no effect on myoblast viability or proliferation but prevented differentiation by impairing the transcriptional actions of MyoD. In contrast, knockdown of Akt2 had no effect on myoblast survival or differentiation and minimally inhibited MyoD-regulated transcription. Our results define isoform-specific Akt-regulated signaling pathways in muscle cells that act through Akt1 to sustain muscle gene activation and promote differentiation. The phosphatidylinositol 3-kinase-Akt pathway plays a central role in growth, development, and metabolism in both normal and neoplastic cells. In skeletal muscle, Akt has been implicated in regulating regeneration and hypertrophy and in counteracting atrophy. Here we provide evidence that Akt1 and not Akt2 is essential for muscle differentiation. Using a robust model of MyoD-mediated muscle development, in which dominant-negative Akt blocked differentiation, we show that targeted loss of Akt1 was equally inhibitory. Selective elimination of Akt1 had no effect on myoblast viability or proliferation but prevented differentiation by impairing the transcriptional actions of MyoD. In contrast, knockdown of Akt2 had no effect on myoblast survival or differentiation and minimally inhibited MyoD-regulated transcription. Our results define isoform-specific Akt-regulated signaling pathways in muscle cells that act through Akt1 to sustain muscle gene activation and promote differentiation. The serine-threonine protein kinases of the Akt family have been subjects of intense scrutiny since their discovery in the early 1990s (1Brazil D.P. Yang Z.Z. Hemmings B.A. Trends Biochem. Sci. 2004; 29: 233-242Abstract Full Text Full Text PDF PubMed Scopus (735) Google Scholar, 2Woodgett J.R. Curr. Opin. Cell Biol. 2005; 17: 150-157Crossref PubMed Scopus (315) Google Scholar). The three Akts share a common structure and ∼85% amino acid identity and are activated by growth factors and hormones through similar biochemical mechanisms that are initiated by production of the lipid signaling molecule, phosphatidylinositol-3,4,5 trisphosphate (1Brazil D.P. Yang Z.Z. Hemmings B.A. Trends Biochem. Sci. 2004; 29: 233-242Abstract Full Text Full Text PDF PubMed Scopus (735) Google Scholar, 2Woodgett J.R. Curr. Opin. Cell Biol. 2005; 17: 150-157Crossref PubMed Scopus (315) Google Scholar). Initial studies of the effects of these proteins established roles for them in cell proliferation, survival, and metabolism (1Brazil D.P. Yang Z.Z. Hemmings B.A. Trends Biochem. Sci. 2004; 29: 233-242Abstract Full Text Full Text PDF PubMed Scopus (735) Google Scholar, 2Woodgett J.R. Curr. Opin. Cell Biol. 2005; 17: 150-157Crossref PubMed Scopus (315) Google Scholar), whereas more recent analyses in mice revealed distinct developmental functions for each Akt. Akt1 deficiency led to a reduction in somatic growth (3Cho H. Thorvaldsen J.L. Chu Q. Feng F. Birnbaum M.J. J. Biol. Chem. 2001; 276: 38349-38352Abstract Full Text Full Text PDF PubMed Scopus (837) Google Scholar); targeted loss of Akt2 caused insulin resistance and diabetes mellitus and growth impairment (4Cho H. Mu J. Kim J.K. Thorvaldsen J.L. Chu Q. Crenshaw E.B. Kaestner K.H. Bartolomei M.S. Shulman G.I. Birnbaum M.J. Science. 2001; 292: 1728-1731Crossref PubMed Scopus (1573) Google Scholar); and knock-out of Akt3 led to reduced brain size (5Easton R.M. Cho H. Roovers K. Shineman D.W. Mizrahi M. Forman M.S. Lee V.M. Szabolcs M. de Jong R. Oltersdorf T. Ludwig T. Efstratiadis A. Birnbaum M.J. Mol. Cell Biol. 2005; 25: 1869-1878Crossref PubMed Scopus (485) Google Scholar). Combined deficiency of Akt1 and Akt2 was associated with severe growth defects and neonatal death (6Peng X.D. Xu P.Z. Chen M.L. Hahn-Windgassen A. Skeen J. Jacobs J. Sundararajan D. Chen W.S. Crawford S.E. Coleman K.G. Hay N. Genes Dev. 2003; 17: 1352-1365Crossref PubMed Scopus (678) Google Scholar), whereas targeted loss of Akt1 and Akt3 was lethal in mid-gestation (7Yang Z.Z. Tschopp O. Di-Poi N. Bruder E. Baudry A. Dummler B. Wahli W. Hemmings B.A. Mol. Cell Biol. 2005; 25: 10407-10418Crossref PubMed Scopus (187) Google Scholar). Skeletal muscle represents a tissue that is responsive to Akt, and Akt action has been linked to skeletal muscle development, regeneration, and hypertrophy through several pathways that culminate in stimulation of protein synthesis, inhibition of atrophy, and prevention of cell death (8Glass D.J. Nat. Cell Biol. 2003; 5: 87-90Crossref PubMed Scopus (549) Google Scholar, 9Hoffman E.P. Nader G.A. Nat. Med. 2004; 10: 584-585Crossref PubMed Scopus (94) Google Scholar). In mice, both Akt1 and Akt2 appear to be important for muscle development as their combined deficiency resulted in severe hypoplasia (6Peng X.D. Xu P.Z. Chen M.L. Hahn-Windgassen A. Skeen J. Jacobs J. Sundararajan D. Chen W.S. Crawford S.E. Coleman K.G. Hay N. Genes Dev. 2003; 17: 1352-1365Crossref PubMed Scopus (678) Google Scholar) but also caused defects in other tissues and organs. Despite these observations, it has not been established which Akt might maintain myoblast survival, promote regeneration, or induce hypertrophy. In cultured muscle cells, Akt2 is induced during differentiation, whereas Akt1 is constant (10Vandromme M. Rochat A. Meier R. Carnac G. Besser D. Hemmings B.A. Fernandez A. Lamb N.J. J. Biol. Chem. 2001; 276: 8173-8179Abstract Full Text Full Text PDF PubMed Scopus (96) Google Scholar, 11Kaneko S. Feldman R.I. Yu L. Wu Z. Gritsko T. Shelley S.A. Nicosia S.V. Nobori T. Cheng J.Q. J. Biol. Chem. 2002; 277: 23230-23235Abstract Full Text Full Text PDF PubMed Scopus (46) Google Scholar, 12Gonzalez I. Tripathi G. Carter E.J. Cobb L.J. Salih D.A. Lovett F.A. Holding C. Pell J.M. Mol. Cell Biol. 2004; 24: 3607-3622Crossref PubMed Scopus (84) Google Scholar). A constitutively active Akt can stimulate muscle hypertrophy in transgenic mice and tissue culture (13Pallafacchina G. Calabria E. Serrano A.L. Kalhovde J.M. Schiaffino S. Proc. Natl. Acad. Sci. U. S. A. 2002; 99: 9213-9218Crossref PubMed Scopus (313) Google Scholar, 14Lai K.M. Gonzalez M. Poueymirou W.T. Kline W.O. Na E. Zlotchenko E. Stitt T.N. Economides A.N. Yancopoulos G.D. Glass D.J. Mol. Cell Biol. 2004; 24: 9295-9304Crossref PubMed Scopus (344) Google Scholar) and can inhibit atrophy induced by denervation (15Bodine S.C. Stitt T.N. Gonzalez M. Kline W.O. Stover G.L. Bauerlein R. Zlotchenko E. Scrimgeour A. Lawrence J.C. Glass D.J. Yancopoulos G.D. Nat. Cell Biol. 2001; 3: 1014-1019Crossref PubMed Scopus (1994) Google Scholar), but it is likely that this overexpressed protein does not faithfully mimic physiologically relevant signaling pathways. Here we have assessed the roles of Akt1 and Akt2 in muscle. We find through selective gene knockdown that Akt1 is necessary for initiation and maintenance of myoblast differentiation but that Akt2 is dispensable. Reduction of each Akt led to similar alterations in phosphorylation of several Akt substrates, but only loss of Akt1 inhibited activity of the myogenic transcription factor MyoD. Our results define functionally separable Akt-mediated signaling mechanisms that act primarily through Akt1 to promote and sustain muscle differentiation. Cell Culture—C3H10T1/2 mouse embryonic fibroblasts (ATCC number CCL226) were incubated on gelatin-coated tissue culture dishes (16Wilson E.M. Hsieh M.M. Rotwein P. J. Biol. Chem. 2003; 278: 41109-41113Abstract Full Text Full Text PDF PubMed Scopus (78) Google Scholar). Differentiation was initiated at ∼95% of confluent density by addition of differentiation medium (DM) 2The abbreviations used are: DM, differentiation medium; PI3, phosphatidylinositol 3; IGF, insulin-like growth factor; mTOR, mammalian target of rapamycin; Ad, adenovirus; Dox, doxycycline. (Dulbecco's modified Eagle's medium plus 2% horse serum (16Wilson E.M. Hsieh M.M. Rotwein P. J. Biol. Chem. 2003; 278: 41109-41113Abstract Full Text Full Text PDF PubMed Scopus (78) Google Scholar)). Construction and Use of Recombinant Adenoviruses—The following have been described: mouse MyoD (Ad-MyoD), dominant-negative Akt (Ad-AktDN), mouse IGF-II in the anti-sense orientation (Ad-IGF-IIAS), tetracycline-inhibited transcriptional activator (Ad-tTA), β-galactosidase (Ad-β-Gal) (16Wilson E.M. Hsieh M.M. Rotwein P. J. Biol. Chem. 2003; 278: 41109-41113Abstract Full Text Full Text PDF PubMed Scopus (78) Google Scholar, 17Wilson E.M. Tureckova J. Rotwein P. Mol. Biol. Cell. 2004; 15: 497-505Crossref PubMed Scopus (60) Google Scholar). Adenoviruses expressing short hairpin inhibitor RNAs targeting mouse Akt1 or Akt2 mRNAs were prepared after testing three sets of double-stranded oligoribonucleotides for each Akt. Akt-selective hairpin oligonucleotides plus a RNA polymerase III termination signal were cloned into an adenoviral shuttle plasmid 3′ to the human H1 RNA promoter (18Kuninger D. Stauffer D. Eftekhari S. Wilson E. Thayer M. Rotwein P. Hum Gene Ther. 2004; 15: 1287-1292Crossref PubMed Scopus (23) Google Scholar). The sense strand is as follows: mouse Akt1, 5′-CTAGTGTGAGGTTGACAGAGGAACTTCAAGAGAGTTCCTCTGTCAACCTCACTTTTTGGATCCA-3′; mouse Akt2, 5′-CTAGTGCCAACCTTGGCTGTTACATTCAAGAGATGTAACAGCCAAGGTTGGCTTTTTGGATCCA-3′. Akt-specific sequences are underlined. Adenoviruses were generated, purified on discontinuous cesium chloride gradients, and titered by optical density (16Wilson E.M. Hsieh M.M. Rotwein P. J. Biol. Chem. 2003; 278: 41109-41113Abstract Full Text Full Text PDF PubMed Scopus (78) Google Scholar). Infections with Ad-MyoD, Ad-IGF-IIAS, and Ad-AktDN were performed as described (19Wilson E.M. Rotwein P. J. Biol. Chem. 2006; 281: 29962-29971Abstract Full Text Full Text PDF PubMed Scopus (101) Google Scholar). For infections with Ad-β-gal, Ad-shAkt1, or Ad-shAkt2, viruses were added to cells at ∼25% of confluent density (multiplicities of infection of 500) followed 24 h later by infection with Ad-MyoD (multiplicity of infection of 125). After an additional 24 h in growth medium, cells were washed, and DM was added. Immunoblotting—Immunoblots were performed as described (19Wilson E.M. Rotwein P. J. Biol. Chem. 2006; 281: 29962-29971Abstract Full Text Full Text PDF PubMed Scopus (101) Google Scholar). Primary antibodies were obtained from Cell Signaling (Beverly, MA) unless otherwise indicated and were used at the following dilutions: anti-Akt (1:2000), anti-phospho-AktSer473 (1:1000), anti-Akt1 (1:2000), anti-Akt2 (1:1000), anti-FoxO3a (1:1000), anti-phospho-FoxO3a (1:500), anti-p70 S6 kinase (1:500), anti-phospho-p70 S6 kinase (1:1000), anti-phospho-GSK-3α/β (1:500), anti-MyoD (Pharmingen, 1:3000), anti-GSK-3α/β (Upstate Cell Signaling, Lake Placid, NY, 1:1000), anti-α-tubulin (Sigma-Aldrich, 1:5000), anti-myogenin (F5D, 1:100), and anti-troponin T (CT3, 1:1000, Developmental Studies Hybridoma Bank, Iowa City, IA). Conjugated secondary antibodies were from Molecular Probes (Eugene, OR, 1:5000). Akt enzymatic assays were performed as described (16Wilson E.M. Hsieh M.M. Rotwein P. J. Biol. Chem. 2003; 278: 41109-41113Abstract Full Text Full Text PDF PubMed Scopus (78) Google Scholar). Immunocytochemistry—All steps have been described (16Wilson E.M. Hsieh M.M. Rotwein P. J. Biol. Chem. 2003; 278: 41109-41113Abstract Full Text Full Text PDF PubMed Scopus (78) Google Scholar), as have calculation of the cell number, fusion index, and myotube area (17Wilson E.M. Tureckova J. Rotwein P. Mol. Biol. Cell. 2004; 15: 497-505Crossref PubMed Scopus (60) Google Scholar). Luciferase Reporter Gene Assays—Reporter plasmids and luciferase assays have been described (19Wilson E.M. Rotwein P. J. Biol. Chem. 2006; 281: 29962-29971Abstract Full Text Full Text PDF PubMed Scopus (101) Google Scholar). Cells were seeded at 1 × 105/12-well dish, and the next day, they were transfected with 0.2 μg of plasmid DNA using TransIT LT-1 (Mirus Corp, Madison, WI). Cells were infected sequentially 16 h later with Ad-β-gal, Ad-shAkt1, or Ad-shAkt2 or both Ad-shAkt1 and Ad-shAkt2 followed by Ad-MyoD, as above. RNA Isolation and Analysis—Reverse transcription-PCR was performed with whole cell RNA (16Wilson E.M. Hsieh M.M. Rotwein P. J. Biol. Chem. 2003; 278: 41109-41113Abstract Full Text Full Text PDF PubMed Scopus (78) Google Scholar) and the following primers: mouse Akt1 sense strand, 5′-GTCTCTAGGGTCCAGGGCCAAAGTC-3′, and antisense, 5′-CATCTAAAAGGACAAGTGCTAGGAG-3′; mouse Akt2 sense, 5′-CCAAGACAGTATTGGGCCCCTTGGA-3′, and antisense, 5′-AGCCTGGGATGCTCACTGCTAGGTC-3′; S17 sense, 5′-ATCCCCAGCAAGAAGCTTCGGAACA-3′, and antisense, 5′-TATGGCATAACAGATTAAACAGCTC-3′. Results were quantified by densitometry after agarose gel electrophoresis. Statistical Analysis—Data are presented as mean ± S.D. Statistical significance was determined by paired Student’s t test (p < 0.05). Inhibition of Akt Activity Blocks MyoD-mediated Muscle Differentiation—We previously defined an autocrine growth factor circuit involving IGF-II, the IGF-I receptor, and the PI3-kinase-Akt pathway that was necessary for differentiation of muscle cell lines (16Wilson E.M. Hsieh M.M. Rotwein P. J. Biol. Chem. 2003; 278: 41109-41113Abstract Full Text Full Text PDF PubMed Scopus (78) Google Scholar, 17Wilson E.M. Tureckova J. Rotwein P. Mol. Biol. Cell. 2004; 15: 497-505Crossref PubMed Scopus (60) Google Scholar) and mesenchymal stem cells acutely converted to myoblasts by MyoD (16Wilson E.M. Hsieh M.M. Rotwein P. J. Biol. Chem. 2003; 278: 41109-41113Abstract Full Text Full Text PDF PubMed Scopus (78) Google Scholar). The of Akt was assessed in which that muscle differentiation can be blocked by inhibition of IGF-II production or by dominant-negative Akt Adenoviruses or not only led to loss of of Akt phosphorylation at but also prevented of muscle and had no effect on MyoD blocked the normal in the enzymatic activity of Akt and inhibited differentiation as was in myoblasts IGF-II (16Wilson E.M. Hsieh M.M. Rotwein P. J. Biol. Chem. 2003; 278: 41109-41113Abstract Full Text Full Text PDF PubMed Scopus (78) Google Scholar, E.M. Rotwein P. J. Biol. Chem. 2006; 281: 29962-29971Abstract Full Text Full Text PDF PubMed Scopus (101) Google Scholar). mechanisms by which Akt muscle differentiation, we assessed the effects of on transcriptional actions of using a gene of a We that stimulation of the IGF-I by IGF-II was for MyoD-mediated muscle gene activation (19Wilson E.M. Rotwein P. J. Biol. Chem. 2006; 281: 29962-29971Abstract Full Text Full Text PDF PubMed Scopus (101) Google Scholar). promoter activity was reduced by by or but to inhibited production of these with Akt-mediated signaling muscle differentiation by impairing transcriptional actions of MyoD. Akt1 for and of Muscle the roles of Akt in differentiation, we short hairpin RNAs targeting Akt1 or Akt2 Ad-shAkt1 reduced of Akt1 in stem cells by but had no effect on for Akt2 or and Ad-shAkt2 Akt2 cells were infected with Ad-shAkt1 and Ad-shAkt2, for both Akts were of Akt1 caused a reduction of Akt1 protein but not the normal in Akt2 during differentiation (10Vandromme M. Rochat A. Meier R. Carnac G. Besser D. Hemmings B.A. Fernandez A. Lamb N.J. J. Biol. Chem. 2001; 276: 8173-8179Abstract Full Text Full Text PDF PubMed Scopus (96) Google Scholar, 11Kaneko S. Feldman R.I. Yu L. Wu Z. Gritsko T. Shelley S.A. Nicosia S.V. Nobori T. Cheng J.Q. J. Biol. Chem. 2002; 277: 23230-23235Abstract Full Text Full Text PDF PubMed Scopus (46) Google Scholar, 12Gonzalez I. Tripathi G. Carter E.J. Cobb L.J. Salih D.A. Lovett F.A. Holding C. Pell J.M. Mol. Cell Biol. 2004; 24: 3607-3622Crossref PubMed Scopus (84) Google Scholar) and not the of MyoD of Akt1 led to a reduction in and and caused an in myotube and and a in fusion with is likely that this muscle differentiation the of cells that were not infected with of Akt2 caused the of Akt2 protein but had effect on Akt1 or MyoD and not inhibit of or of Akt2 also not or fusion The of Ad-shAkt1 and Ad-shAkt2 led to the elimination of both Akt proteins and blocked differentiation to the as Ad-shAkt1 We results in the muscle cell Akt1 deficiency inhibited differentiation at and knockdown of Akt2 not of Akt1 or Akt2 not cell proliferation during in growth medium and not myoblast death during h in DM loss of both Akts led to reduction in cell number after 24 or h in DM the cell death the of MyoD in myoblast survival overexpressed Rotwein P. J. Cell Biol. PubMed Scopus Google Scholar), Akt has been to be a in muscle cell viability Rotwein P. J. Cell Biol. PubMed Scopus Google Scholar, K. T. J.R. K. Mol. Cell. Biol. PubMed Scopus Google Scholar). Signaling by Akt1 and Akt2 in Muscle mechanisms by which Akt1 and Akt2 differentiation, we the effects of reduction of each protein on Akt target Akt1 deficiency led to a in Akt and phosphorylation at loss of Akt2 of Akt1 or Akt2 led to in phosphorylation of and on and Akt1 loss caused a in of kinase and in phosphorylation by loss of Reduction of Akt1 led to a in phosphorylation of D. Cell. 2004; Full Text Full Text PDF PubMed Scopus Google Scholar), Akt to the in of during in alterations in Akt target proteins were in cells not loss of each Akt to to in signaling proteins and effects on of Akt1 of next Akt deficiency inhibited MyoD-mediated transcription. promoter activity was reduced by in cells Akt1, whereas loss of Akt2 caused a Muscle kinase promoter activity also by in cells Akt1, whereas Akt2 deficiency caused a of both Akts results similar to of Akt1 Akt1 and Akt2 distinct effects on muscle gene Akt1 to be for of muscle gene early in differentiation, loss differentiation. contrast, Akt2 is for early muscle gene but to of later Akts in Cell and results to a on actions of Akts in cell For viability of and their differentiation was reduced by loss of Akt1 but not by of Akt2 and be to normal by cell death with a inhibitor D.J. J. Biol. Chem. 2006; 281: Full Text Full Text PDF PubMed Scopus Google Scholar). We find that the inhibitor not the of muscle differentiation with Akt1 deficiency not that mechanisms in and contrast, differentiation to Akt1 and Akt2 as knockdown of protein was cell viability or proliferation T. J. Biol. Chem. 2005; Full Text Full Text PDF PubMed Scopus Google Scholar). In or Akt2 is essential for normal whereas Akt1 was Cho H. Mu J. Birnbaum M.J. J. Biol. Chem. 2003; 278: Full Text Full Text PDF PubMed Scopus Google Scholar, K. A. L. A. J.R. Cell 2006; Full Text Full Text PDF PubMed Scopus Google Scholar). In cells, Akt1 to and with and loss as as Akt2 also was D. M. P. N. S. J. Cell Biol. 2005; PubMed Scopus Google Scholar). The mechanisms by which Akt1 in these cells are not In Akt1 deficiency led to which was for cell D. M. P. N. S. J. Cell Biol. 2005; PubMed Scopus Google Scholar), whereas in Akt1 of the transcription factor by the M. I. P. S. A. Mol. Cell. 2005; Full Text Full Text PDF PubMed Scopus Google Scholar). on the cell or tissue Akt1 or Akt2 similar or distinct effects on differentiation, survival, or The steps by which Akts with pathways in cell to an Akts and Skeletal roles for Akt proteins in muscle growth and regeneration after (8Glass D.J. Nat. Cell Biol. 2003; 5: 87-90Crossref PubMed Scopus (549) Google Scholar, 9Hoffman E.P. Nader G.A. Nat. Med. 2004; 10: 584-585Crossref PubMed Scopus (94) Google Scholar). Akts inhibit transcription factors by of D. Cell. 2004; Full Text Full Text PDF PubMed Scopus Google Scholar). In muscle, this of and in atrophy M. C. A. C. Calabria E. A. K. Schiaffino S. A.L. Cell. 2004; Full Text Full Text PDF PubMed Scopus Google Scholar, T.N. D. B.A. F. Kline W.O. Gonzalez M. Yancopoulos G.D. Glass D.J. Mol. Cell. 2004; Full Text Full Text PDF PubMed Scopus Google Scholar). In transgenic mice, of active Akt led to hypertrophy K.M. Gonzalez M. Poueymirou W.T. Kline W.O. Na E. Zlotchenko E. Stitt T.N. Economides A.N. Yancopoulos G.D. Glass D.J. Mol. Cell Biol. 2004; 24: 9295-9304Crossref PubMed Scopus (344) Google Scholar), as of mouse or with a similar (13Pallafacchina G. Calabria E. Serrano A.L. Kalhovde J.M. Schiaffino S. Proc. Natl. Acad. Sci. U. S. A. 2002; 99: 9213-9218Crossref PubMed Scopus (313) Google Scholar, S.C. Stitt T.N. Gonzalez M. Kline W.O. Stover G.L. Bauerlein R. Zlotchenko E. Scrimgeour A. Lawrence J.C. Glass D.J. Yancopoulos G.D. Nat. Cell Biol. 2001; 3: 1014-1019Crossref PubMed Scopus (1994) Google Scholar). of active Akt in also reduced muscle atrophy after denervation and after effects blocked by the (13Pallafacchina G. Calabria E. Serrano A.L. Kalhovde J.M. Schiaffino S. Proc. Natl. Acad. Sci. U. S. A. 2002; 99: 9213-9218Crossref PubMed Scopus (313) Google Scholar, S.C. Stitt T.N. Gonzalez M. Kline W.O. Stover G.L. Bauerlein R. Zlotchenko E. Scrimgeour A. Lawrence J.C. Glass D.J. Yancopoulos G.D. Nat. Cell Biol. 2001; 3: 1014-1019Crossref PubMed Scopus (1994) Google Scholar), that selective pathways of Akt are to hypertrophy was in transgenic mice IGF-I in muscle L. A. N. J. Cell Biol. 2002; PubMed Scopus Google Scholar), and muscle was in mice with by an IGF-I with Akt phosphorylation L. A. N. J. Cell Biol. 2002; PubMed Scopus Google Scholar). Akts actions of IGF-I in muscle, but the Akt an We established that autocrine signaling by the IGF-I receptor, and Akt, was necessary for transcriptional actions of MyoD after of muscle differentiation (19Wilson E.M. Rotwein P. J. Biol. Chem. 2006; 281: 29962-29971Abstract Full Text Full Text PDF PubMed Scopus (101) Google Scholar). Inhibition of this pathway blocked gene transcription MyoD or DNA inhibition of signaling reduced of transcriptional and on at the and impairing and activation of RNA polymerase (19Wilson E.M. Rotwein P. J. Biol. Chem. 2006; 281: 29962-29971Abstract Full Text Full Text PDF PubMed Scopus (101) Google Scholar). with observations, these results Akt1 as a in MyoD-mediated transcriptional pathways in muscle cells, the mechanisms by which Akt1 activity of MyoD to be A model is in that Akt2 in during muscle differentiation (10Vandromme M. Rochat A. Meier R. Carnac G. Besser D. Hemmings B.A. Fernandez A. Lamb N.J. J. Biol. Chem. 2001; 276: 8173-8179Abstract Full Text Full Text PDF PubMed Scopus (96) Google Scholar, 11Kaneko S. Feldman R.I. Yu L. Wu Z. Gritsko T. Shelley S.A. Nicosia S.V. Nobori T. Cheng J.Q. J. Biol. Chem. 2002; 277: 23230-23235Abstract Full Text Full Text PDF PubMed Scopus (46) Google Scholar, 12Gonzalez I. Tripathi G. Carter E.J. Cobb L.J. Salih D.A. Lovett F.A. Holding C. Pell J.M. Mol. Cell Biol. 2004; 24: 3607-3622Crossref PubMed Scopus (84) Google Scholar) and that can inhibit differentiation (10Vandromme M. Rochat A. Meier R. Carnac G. Besser D. Hemmings B.A. Fernandez A. Lamb N.J. J. Biol. Chem. 2001; 276: 8173-8179Abstract Full Text Full Text PDF PubMed Scopus (96) Google Scholar) have led to the that Akt2 Akt1 plays a central role in muscle. In contrast, results that Akt1 is for and maintenance of muscle differentiation, we in In cells, of Akt2 led to reduction in a in fusion and whereas in cells, loss was associated with that act of Akt have been linked to the mechanisms are The protein kinase protein and is activated by Akt S. R. Cell. 2006; Full Text Full Text PDF PubMed Scopus Google Scholar). an cells with protein Chen J. J. Biol. Chem. 2005; Full Text Full Text PDF PubMed Scopus Google Scholar). The transcription factor was to fusion and myotube overexpressed J. 2003; PubMed Scopus Google Scholar), other results that inhibited differentiation M.L. J. T. J.R. D. J. Cell Biol. 2003; PubMed Scopus Google Scholar). signaling through the IGF-I has been associated with and D. 2005; PubMed Scopus (101) Google Scholar) and in M. A. A. Science. 2003; PubMed Scopus Google Scholar). the with IGF-I has been for muscle and to in (8Glass D.J. Nat. Cell Biol. 2003; 5: 87-90Crossref PubMed Scopus (549) Google Scholar, 9Hoffman E.P. Nader G.A. Nat. Med. 2004; 10: 584-585Crossref PubMed Scopus (94) Google Scholar). of this growth factor in muscle or other tissues a of signaling and their mechanisms of We for with