Résumé :
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The myogenesis process involves the expression of muscle-specific transcription factors such as myogenin and MEF2, and is essentially regulated by fluctuations of the cytosolic Ca2+ concentration. Previous work demonstrated that hyperpolarization of the membrane potential is one of the first events to occur. This hyperpolarization of the myoblast results from a dephosphorylation at tyrosine 242 of the inward rectifying K+ channels Kir2.1 leading to its activation. The resulting increased driving force for Ca2+ caused by hyperpolarization promotes Ca2+ influx through Ca2+-channels. Arnaudeau and co-workers have demonstrated that store-operated Ca2+-entry (SOCE) channels are required for cell differentiation. In line with this, the molecular players of SOCE, STIM1, STIM2 (Stromal Interacting Molecule) and Orai1 were reported to be fundamental in the induction of the differentiation process of post-natal myoblasts. Besides STIM & Orai proteins, the family of TRPC (canonical transient receptor potential) channels was shown to be part of SOCE in several cellular systems. In this study, we investigated the role of TRPCs channels in human myogenesis process. We first showed that TRPC1, C3 and TRPC4 are endogenously expressed in human myoblast, and the knockdown of each of them reduced SOCE by about 20%. We also observed a strong reduction of myoblast proliferation in TRPC3 knockdown cells, while the reduction of TRPC1 and TRPC4 did not impact on myoblast proliferation. Second, we investigated the involvement of TRPCs channels on myoblast differentiation and fusion. The silencing of TRPC1, C3 and C4 reduced MEF2 expression by 30 to 50% after 48h in differentiation medium, while the myogenin expression was not affected. Futhermore, the efficiency of myoblast fusion was severely affected as shown by a reduction of the fusion index by 40% upon siTRPC3 treatment and by 65% after the silencing of TRPC1 or TRPC4. Our results show that TRPCs account for about 20% of the SOCE, and are required for the proper expression of the transcription factor MEF2. Strikingly, the fusion of myoblast appears to be the most affected event upon silencing of TRPCs channels.
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