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Expression and activity of serum response factor is required for expression of the muscle-determining factor MyoD in
C Gauthier-Rouviere1, M Vandromme, D Tuil
1Cell Biology Unit, Centre de Recherche de Macromoléculaire, CNRS-INSERM, Montpellier, France.
Abstract:
To understand the mechanism by which the serum response factor (SRF) is involved in the process of skeletal muscle differentiation, we have assessed the effect of inhibiting SRF activity or synthesis on the expression of the muscle-determining factor MyoD. Inhibition of SRF activity in mouse myogenic C2C12 cells through microinjection of either the SRE oligonucleotide (which acts by displacing SRF proteins from the endogenous SRE sequences), purified SRF-DB (a 30-kDa portion of SRF containing the DNA-binding domain of SRF, which acts as a dominant negative mutant in vivo), or purified anti-SRF antibodies rapidly prevents the expression of MyoD. Moreover, the rapid shutdown of MyoD expression after in vivo inhibition of SRF activity is observed not only in proliferating myoblasts but also in myoblasts cultured under differentiating conditions. Additionally, by using a cellular system expressing a glucocorticoid-inducible antisense-SRF (from aa 74 to 244) we have shown that blocking SRF expression by dexamethasone induction of antisense SRF results in the lack of MyoD expression as probed by both immunofluorescence and Northern blot analysis. Taken together these data demonstrate that SRF expression and activity are required for the expression of the muscle-determining factor MyoD.
Insights
Serum response factor (SRF) is essential for skeletal muscle differentiation. Inhibiting SRF activity or synthesis prevents the expression of MyoD, a key muscle-determining factor.
Area of Science:
- Molecular Biology
- Cell Biology
- Developmental Biology
Background:
- Skeletal muscle differentiation is a complex process involving the precise regulation of gene expression.
- The serum response factor (SRF) is a transcription factor known to play roles in various cellular processes.
- Understanding the specific role of SRF in muscle development is crucial for elucidating differentiation mechanisms.
Purpose of the Study:
- To investigate the mechanism by which serum response factor (SRF) influences skeletal muscle differentiation.
- To determine the effect of inhibiting SRF activity or synthesis on the expression of the muscle-determining factor, MyoD.
Main Methods:
- Utilized mouse myogenic C2C12 cells for experiments.
- Inhibited SRF activity via microinjection of SRE oligonucleotide, dominant-negative SRF mutant (SRF-DB), or anti-SRF antibodies.
- Assessed MyoD expression using immunofluorescence and Northern blot analysis.
- Employed a cellular system with glucocorticoid-inducible antisense-SRF to block SRF synthesis.
Main Results:
- Inhibition of SRF activity rapidly prevented MyoD expression in both proliferating and differentiating myoblasts.
- Blocking SRF synthesis using antisense SRF also resulted in a lack of MyoD expression.
- These findings were consistent across different methods of SRF inhibition.
Conclusions:
- SRF expression and activity are demonstrated to be necessary for the expression of MyoD.
- SRF plays a critical role in the transcriptional regulation of MyoD during skeletal muscle differentiation.
- This study highlights SRF as a key regulator in the myogenic pathway.