Suppression of protein kinase C theta contributes to enhanced myogenesis In vitro via IRS1 and ERK1/2 phosphorylation

Joseph S. Marino; Terry D. Hinds; Rachael A. Potter; Eric Ondrus; Jeremy L. Onion; Abigail Dowling; Thomas J. McLoughlin; Edwin R. Sanchez; Jennifer W. Hill

doi:10.1186/1471-2121-14-39

Suppression of protein kinase C theta contributes to enhanced myogenesis In vitro via IRS1 and ERK1/2 phosphorylation

Joseph S. Marino, Terry D. Hinds, Rachael A. Potter, Eric Ondrus, Jeremy L. Onion, Abigail Dowling, Thomas J. McLoughlin, Edwin R. Sanchez, Jennifer W. Hill

Research output: Contribution to journal › Article › peer-review

14 Scopus citations

Abstract

Background: Differentiation and fusion of skeletal muscle myoblasts into multi-nucleated myotubes is required for neonatal development and regeneration in adult skeletal muscle. Herein, we report novel findings that protein kinase C theta (PKCθ) regulates myoblast differentiation via phosphorylation of insulin receptor substrate-1 and ERK1/2.Results: In this study, PKCθ knockdown (PKCθ^shRNA) myotubes had reduced inhibitory insulin receptor substrate-1 ser1095 phosphorylation, enhanced myoblast differentiation and cell fusion, and increased rates of protein synthesis as determined by [³H] phenylalanine incorporation. Phosphorylation of insulin receptor substrate-1 ser632/635 and extracellular signal-regulated kinase1/2 (ERK1/2) was increased in PKCθ^shRNA cells, with no change in ERK5 phosphorylation, highlighting a PKCθ-regulated myogenic pathway. Inhibition of PI3-kinase prevented cell differentiation and fusion in control cells, which was attenuated in PKCθ^shRNA cells. Thus, with reduced PKCθ, differentiation and fusion occur in the absence of PI3-kinase activity. Inhibition of the ERK kinase, MEK1/2, impaired differentiation and cell fusion in control cells. Differentiation was preserved in PKCθ^shRNA cells treated with a MEK1/2 inhibitor, although cell fusion was blunted, indicating PKCθ regulates differentiation via IRS1 and ERK1/2, and this occurs independently of MEK1/2 activation.Conclusion: Cellular signaling regulating the myogenic program and protein synthesis are complex and intertwined. These studies suggest that PKCθ regulates myogenic and protein synthetic signaling via the modulation of IRS1and ERK1/2 phosphorylation. Myotubes lacking PKCθ had increased rates of protein synthesis and enhanced myotube development despite reduced activation of the canonical anabolic-signaling pathway. Further investigation of PKCθ regulated signaling may reveal important interactions regulating skeletal muscle health in an insulin resistant state.

Original language	English
Article number	39
Journal	BMC Cell Biology
Volume	14
Issue number	1
DOIs	https://doi.org/10.1186/1471-2121-14-39
State	Published - Sep 21 2013

Bibliographical note

Funding Information:
The authors thank Francis X. Pizza (UT-Main Campus) for technical advice, and Lance Stechschulte, and Leah Wuescher (UT-College of Medicine) for technical assistance. The authors would like to thank Susan Tsivitse Arthur (UNC Charlotte) for critical review of this manuscript. This work was supported by the University of Toledo deArce-Koch award to JWH.

Funding

The authors thank Francis X. Pizza (UT-Main Campus) for technical advice, and Lance Stechschulte, and Leah Wuescher (UT-College of Medicine) for technical assistance. The authors would like to thank Susan Tsivitse Arthur (UNC Charlotte) for critical review of this manuscript. This work was supported by the University of Toledo deArce-Koch award to JWH.

Funders	Funder number
University of Toledo deArce-Koch
National Institute of Diabetes and Digestive and Kidney Diseases	P30DK020572

Keywords

Insulin receptor substrate
Myoblast differentiation
Myoblast fusion
Protein kinase C

ASJC Scopus subject areas

Cell Biology

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title = "Suppression of protein kinase C theta contributes to enhanced myogenesis In vitro via IRS1 and ERK1/2 phosphorylation",

abstract = "Background: Differentiation and fusion of skeletal muscle myoblasts into multi-nucleated myotubes is required for neonatal development and regeneration in adult skeletal muscle. Herein, we report novel findings that protein kinase C theta (PKCθ) regulates myoblast differentiation via phosphorylation of insulin receptor substrate-1 and ERK1/2.Results: In this study, PKCθ knockdown (PKCθshRNA) myotubes had reduced inhibitory insulin receptor substrate-1 ser1095 phosphorylation, enhanced myoblast differentiation and cell fusion, and increased rates of protein synthesis as determined by [3H] phenylalanine incorporation. Phosphorylation of insulin receptor substrate-1 ser632/635 and extracellular signal-regulated kinase1/2 (ERK1/2) was increased in PKCθshRNA cells, with no change in ERK5 phosphorylation, highlighting a PKCθ-regulated myogenic pathway. Inhibition of PI3-kinase prevented cell differentiation and fusion in control cells, which was attenuated in PKCθshRNA cells. Thus, with reduced PKCθ, differentiation and fusion occur in the absence of PI3-kinase activity. Inhibition of the ERK kinase, MEK1/2, impaired differentiation and cell fusion in control cells. Differentiation was preserved in PKCθshRNA cells treated with a MEK1/2 inhibitor, although cell fusion was blunted, indicating PKCθ regulates differentiation via IRS1 and ERK1/2, and this occurs independently of MEK1/2 activation.Conclusion: Cellular signaling regulating the myogenic program and protein synthesis are complex and intertwined. These studies suggest that PKCθ regulates myogenic and protein synthetic signaling via the modulation of IRS1and ERK1/2 phosphorylation. Myotubes lacking PKCθ had increased rates of protein synthesis and enhanced myotube development despite reduced activation of the canonical anabolic-signaling pathway. Further investigation of PKCθ regulated signaling may reveal important interactions regulating skeletal muscle health in an insulin resistant state.",

keywords = "Insulin receptor substrate, Myoblast differentiation, Myoblast fusion, Protein kinase C",

author = "Marino, {Joseph S.} and Hinds, {Terry D.} and Potter, {Rachael A.} and Eric Ondrus and Onion, {Jeremy L.} and Abigail Dowling and McLoughlin, {Thomas J.} and Sanchez, {Edwin R.} and Hill, {Jennifer W.}",

note = "Funding Information: The authors thank Francis X. Pizza (UT-Main Campus) for technical advice, and Lance Stechschulte, and Leah Wuescher (UT-College of Medicine) for technical assistance. The authors would like to thank Susan Tsivitse Arthur (UNC Charlotte) for critical review of this manuscript. This work was supported by the University of Toledo deArce-Koch award to JWH.",

year = "2013",

month = sep,

day = "21",

doi = "10.1186/1471-2121-14-39",

language = "English",

volume = "14",

number = "1",

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T1 - Suppression of protein kinase C theta contributes to enhanced myogenesis In vitro via IRS1 and ERK1/2 phosphorylation

AU - Marino, Joseph S.

AU - Hinds, Terry D.

AU - Potter, Rachael A.

AU - Ondrus, Eric

AU - Onion, Jeremy L.

AU - Dowling, Abigail

AU - McLoughlin, Thomas J.

AU - Sanchez, Edwin R.

AU - Hill, Jennifer W.

N1 - Funding Information: The authors thank Francis X. Pizza (UT-Main Campus) for technical advice, and Lance Stechschulte, and Leah Wuescher (UT-College of Medicine) for technical assistance. The authors would like to thank Susan Tsivitse Arthur (UNC Charlotte) for critical review of this manuscript. This work was supported by the University of Toledo deArce-Koch award to JWH.

PY - 2013/9/21

Y1 - 2013/9/21

N2 - Background: Differentiation and fusion of skeletal muscle myoblasts into multi-nucleated myotubes is required for neonatal development and regeneration in adult skeletal muscle. Herein, we report novel findings that protein kinase C theta (PKCθ) regulates myoblast differentiation via phosphorylation of insulin receptor substrate-1 and ERK1/2.Results: In this study, PKCθ knockdown (PKCθshRNA) myotubes had reduced inhibitory insulin receptor substrate-1 ser1095 phosphorylation, enhanced myoblast differentiation and cell fusion, and increased rates of protein synthesis as determined by [3H] phenylalanine incorporation. Phosphorylation of insulin receptor substrate-1 ser632/635 and extracellular signal-regulated kinase1/2 (ERK1/2) was increased in PKCθshRNA cells, with no change in ERK5 phosphorylation, highlighting a PKCθ-regulated myogenic pathway. Inhibition of PI3-kinase prevented cell differentiation and fusion in control cells, which was attenuated in PKCθshRNA cells. Thus, with reduced PKCθ, differentiation and fusion occur in the absence of PI3-kinase activity. Inhibition of the ERK kinase, MEK1/2, impaired differentiation and cell fusion in control cells. Differentiation was preserved in PKCθshRNA cells treated with a MEK1/2 inhibitor, although cell fusion was blunted, indicating PKCθ regulates differentiation via IRS1 and ERK1/2, and this occurs independently of MEK1/2 activation.Conclusion: Cellular signaling regulating the myogenic program and protein synthesis are complex and intertwined. These studies suggest that PKCθ regulates myogenic and protein synthetic signaling via the modulation of IRS1and ERK1/2 phosphorylation. Myotubes lacking PKCθ had increased rates of protein synthesis and enhanced myotube development despite reduced activation of the canonical anabolic-signaling pathway. Further investigation of PKCθ regulated signaling may reveal important interactions regulating skeletal muscle health in an insulin resistant state.

AB - Background: Differentiation and fusion of skeletal muscle myoblasts into multi-nucleated myotubes is required for neonatal development and regeneration in adult skeletal muscle. Herein, we report novel findings that protein kinase C theta (PKCθ) regulates myoblast differentiation via phosphorylation of insulin receptor substrate-1 and ERK1/2.Results: In this study, PKCθ knockdown (PKCθshRNA) myotubes had reduced inhibitory insulin receptor substrate-1 ser1095 phosphorylation, enhanced myoblast differentiation and cell fusion, and increased rates of protein synthesis as determined by [3H] phenylalanine incorporation. Phosphorylation of insulin receptor substrate-1 ser632/635 and extracellular signal-regulated kinase1/2 (ERK1/2) was increased in PKCθshRNA cells, with no change in ERK5 phosphorylation, highlighting a PKCθ-regulated myogenic pathway. Inhibition of PI3-kinase prevented cell differentiation and fusion in control cells, which was attenuated in PKCθshRNA cells. Thus, with reduced PKCθ, differentiation and fusion occur in the absence of PI3-kinase activity. Inhibition of the ERK kinase, MEK1/2, impaired differentiation and cell fusion in control cells. Differentiation was preserved in PKCθshRNA cells treated with a MEK1/2 inhibitor, although cell fusion was blunted, indicating PKCθ regulates differentiation via IRS1 and ERK1/2, and this occurs independently of MEK1/2 activation.Conclusion: Cellular signaling regulating the myogenic program and protein synthesis are complex and intertwined. These studies suggest that PKCθ regulates myogenic and protein synthetic signaling via the modulation of IRS1and ERK1/2 phosphorylation. Myotubes lacking PKCθ had increased rates of protein synthesis and enhanced myotube development despite reduced activation of the canonical anabolic-signaling pathway. Further investigation of PKCθ regulated signaling may reveal important interactions regulating skeletal muscle health in an insulin resistant state.

KW - Insulin receptor substrate

KW - Myoblast differentiation

KW - Myoblast fusion

KW - Protein kinase C

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JO - BMC Cell Biology

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ER -

Suppression of protein kinase C theta contributes to enhanced myogenesis In vitro via IRS1 and ERK1/2 phosphorylation

Abstract

Bibliographical note

Funding

Keywords

ASJC Scopus subject areas

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