Downregulation of POFUT1 Impairs Secondary Myogenic Fusion Through a Reduced NFATc2/IL-4 Signaling Pathway

Past work has shown that the protein O-fucosyltransferase 1 (POFUT1) is involved in mammal myogenic differentiation program. Pofut1 knockdown (Po -) in murine C2C12 cells leads to numerous elongated and thin myotubes, suggesting significant defects in secondary fusion. Among the few pathways involved in this process, NFATc2/IL-4 is described as the major one. To unravel the impact of POFUT1 on secondary fusion, we used wild-type (WT) C2C12 and Po - cell lines to follow Myf6, Nfatc2, Il-4 and Il-4r alpha expressions during a 120 h myogenic differentiation time course. Secreted IL-4 was quantified by ELISA. IL-4R alpha expression and its labeling on myogenic cell types were investigated by Western blot and immunofluorescence, respectively. Phenotypic observations of cells treated with IL-4R alpha blocking antibody were performed. In Po -, we found a decrease in nuclei number per myotube and a downexpression of Myf6. The observed downregulation of Nfatc2 is correlated to a diminution of secreted IL-4 and to the low level of IL-4R alpha for reserve cells. Neutralization of IL-4R alpha on WT C2C12 promotes myonuclear accretion defects, similarly to those identified in Po -. Thus, POFUT1 could be a new controller of myotube growth during myogenesis, especially through NFATc2/IL-4 signaling pathway.