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Atypical protein kinase Cs are the Ras effectors that mediate repression of myogenic satellite cell differentiation
Yuri V Fedorov1, Nathan C Jones, Bradley B Olwin
1Department of Molecular, Cellular and Developmental Biology, University of Colorado, Boulder, Colorado 80309, USA.
Abstract:
Oncogenic Ha-Ras is a potent inhibitor of skeletal muscle cell differentiation, yet the Ras effector mediating this process remains unidentified. Here we demonstrate that the atypical protein kinases (aPKCs; lambda and/or zeta) are downstream Ras effectors responsible for Ras-dependent inhibition of myogenic differentiation in a satellite cell line. First, ectopic expression of Ha-RasG12V induces translocation of PKClambda from the cytosol to the nucleus, suggesting that aPKCs are activated by Ras in myoblasts. The aPKCs function as downstream Ras effectors since inhibition of aPKCs by expression of a dominant negative PKCzeta mutant or by treatment of cells with an inhibitor, GO6983, promotes myogenesis in skeletal muscle satellite cells expressing oncogenic Ha-Ras. Arresting cell proliferation synergistically enhances myogenic differentiation only when aPKCs are also inhibited. Thus, the repression of myogenic differentiation in a satellite cell line appears to be directly mediated by aPKCs acting as Ras effectors and indirectly mediated via stimulation of cell proliferation.
Insights
Oncogenic Ha-Ras inhibits muscle cell differentiation by activating atypical protein kinases (aPKCs). Inhibiting aPKCs promotes myogenesis, revealing a key pathway in skeletal muscle development.
Area of Science:
- Molecular Biology
- Cell Biology
- Biochemistry
Background:
- Oncogenic Ha-Ras is known to inhibit skeletal muscle cell differentiation.
- The specific Ras effector responsible for this inhibition has not been identified.
Purpose of the Study:
- To identify the downstream Ras effector mediating the inhibition of myogenic differentiation.
- To elucidate the role of atypical protein kinases (aPKCs) in Ras-dependent muscle cell differentiation.
Main Methods:
- Ectopic expression of oncogenic Ha-Ras (Ha-RasG12V) in a satellite cell line.
- Assessing the translocation of protein kinase C lambda (PKClambda) in response to Ha-Ras.
- Inhibiting aPKCs using a dominant-negative PKCzeta mutant and the chemical inhibitor GO6983.
- Evaluating the effects of Ha-Ras and aPKC inhibition on myogenic differentiation and cell proliferation.
Main Results:
- Oncogenic Ha-Ras expression induced PKClambda translocation to the nucleus, indicating aPKC activation.
- Inhibition of aPKCs promoted myogenesis in Ha-Ras-expressing skeletal muscle satellite cells.
- Simultaneous inhibition of aPKCs and cell proliferation synergistically enhanced myogenic differentiation.
Conclusions:
- Atypical protein kinases (aPKCs) are identified as downstream Ras effectors that directly inhibit myogenic differentiation.
- Ras-mediated repression of skeletal muscle differentiation occurs both directly via aPKCs and indirectly through the stimulation of cell proliferation.