Articles by Sophie Saüc in JoVE
Isolation of Human Myoblasts, Assessment of Myogenic Differentiation, and Store-operated Calcium Entry Measurement Thomas Laumonier*1, Stéphane Koenig*2, Sophie Saüc2,3, Maud Frieden3 1Department of Orthopedic Surgery, Geneva University Hospitals & Faculty of Medicine, 2Department of Basic Neurosciences, Geneva Medical Center, 3Department of Cell Physiology and Metabolism, Geneva Medical Center Here, we describe a methodology to obtain a pure population of human myoblasts from adult muscle tissue. These cells are used to study in vitro skeletal muscle differentiation and, in particular, to study proteins involved in Ca2+ signaling.
Other articles by Sophie Saüc on PubMed
STIM1L Traps and Gates Orai1 Channels Without Remodeling the Cortical ER Journal of Cell Science. Apr, 2015 | Pubmed ID: 25736291 STIM proteins populate and expand cortical endoplasmic reticulum (ER) sheets to mediate store-operated Ca(2+) entry (SOCE) by trapping and gating Orai channels in ER-plasma membrane clusters. A longer splice variant, STIM1L, forms permanent ER-plasma membrane clusters and mediates rapid Ca(2+) influx in muscle. Here, we used electron microscopy, total internal reflection fluorescence (TIRF) microscopy and Ca(2+) imaging to establish the trafficking and signaling properties of the two STIM1 isoforms in Stim1(-/-)/Stim2(-/-) fibroblasts. Unlike STIM1, STIM1L was poorly recruited into ER-plasma membrane clusters and did not mediate store-dependent expansion of cortical ER cisternae. Removal of the STIM1 lysine-rich tail prevented store-dependent cluster enlargement, whereas inhibition of cytosolic Ca(2+) elevations or removal of the STIM1L actin-binding domain had no impact on cluster expansion. Finally, STIM1L restored robust but not accelerated SOCE and clustered with Orai1 channels more slowly than STIM1 following store depletion. These results indicate that STIM1L does not mediate rapid SOCE but can trap and gate Orai1 channels efficiently without remodeling cortical ER cisternae. The ability of STIM proteins to induce cortical ER formation is dispensable for SOCE and requires the lysine-rich tail of STIM1 involved in binding to phosphoinositides.
TRPC1 and TRPC4 Channels Functionally Interact with STIM1L to Promote Myogenesis and Maintain Fast Repetitive Ca(2+) Release in Human Myotubes Biochimica Et Biophysica Acta. 05, 2017 | Pubmed ID: 28185894 STIM1 and Orai1 are essential players of store-operated Ca(2+) entry (SOCE) in human skeletal muscle cells and are required for adult muscle differentiation. Besides these two proteins, TRPC (transient receptor potential canonical) channels and STIM1L (a longer STIM1 isoform) are also present on muscle cells. In the present study, we assessed the role of TRPC1, TRPC4 and STIM1L in SOCE, in the maintenance of repetitive Ca(2+) transients and in muscle differentiation. Knockdown of TRPC1 and TRPC4 reduced SOCE by about 50% and significantly delayed the onset of Ca(2+) entry, both effects similar to STIM1L invalidation. Upon store depletion, TRPC1 and TRPC4 appeared to interact preferentially with STIM1L compared to STIM1. STIM1L invalidation affected myoblast differentiation, with the formation of smaller myotubes, an effect similar to what we reported for TRPC1 and TRPC4 knockdown. On the contrary, the overexpression of STIM1L leads to the formation of larger myotubes. All together, these data strongly suggest that STIM1L and TRPC1/4 are working together in myotubes to ensure efficient store refilling and a proper differentiation program.