Inhibition of myogenesis by depletion of the glycogen-associated regulatory subunit of protein phosphatase-1 in rat skeletal muscle cells.
In this study, we examined the role of the glycogen-associated regulatory subunit of protein phosphatase-1 (PP-1(G)) in L6 rat skeletal muscle cell myogenesis. The level of PP-1(G) was depleted by transfection with an inducible antisense-oriented PP-1(G) gene. Western blot analysis of the PP-1(G)-depleted cell line revealed a >90% depletion of PP-1(G) protein and a 45% reduction in cellular PP-1 activity and abolished the ability of L6 myoblasts to differentiate into multinucleated myotubes. PP-1(G)-depleted cells also exhibited a marked reduction in the expression of the differentiation marker myogenin as well as creatine kinase. After 7 days in culture, PP-1(G)-depleted cells sustained myoblast levels of inhibitor of differentiation-2, whereas control L6 cells had a severely lower inhibitor of differentiation-2 level and progressed into myotubes. Myoblasts were unable to exit the cell cycle, as measured by the impaired induction of p27 cyclin-dependent kinase inhibitor, a >2-fold increase in DNA synthesis, and elevated levels of phosphorylated retinoblastoma protein (pRb). Replacement of the PP-1(G) gene restored PP-1(G) protein expression, PP-1 enzymatic activity, and the ability to differentiate into myotubes. We conclude that PP-1(G) plays a definite role in L6 myogenesis via its regulation of PP-1 catalytic activity.
Ragolia
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Diabetes Research Laboratory, Winthrop University Hospital, Mineola, New York 11501 and the School of Medicine, State University of New York, Stony Brook, New York 11794, USA. lragolia@winthrop.org
Zuo
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Begum
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D000818:Animals; D002352:Carrier Proteins / Q000235:genetics / Q000502:physiology*; D002460:Cell Line; D003402:Creatine Kinase / Q000378:metabolism; D004261:DNA Replication; D007700:Kinetics; D024510:Muscle Development; D018482:Muscle, Skeletal / Q000201:enzymology* / Q000254:growth & development*; D020319:Oligodeoxyribonucleotides, Antisense / Q000378:metabolism; D010749:Phosphoprotein Phosphatases / Q000235:genetics / Q000502:physiology*; D010766:Phosphorylation; D054645:Protein Phosphatase 1; D051381:Rats; D016160:Retinoblastoma Protein / Q000378:metabolism; D014162:Transfection
D002352:Carrier Proteins; D020319:Oligodeoxyribonucleotides, Antisense; D016160:Retinoblastoma Protein; D003402:Creatine Kinase; D010749:Phosphoprotein Phosphatases; C114052:Ppp1r3b protein, rat; D054645:Protein Phosphatase 1