Functional exploration of Lrtm1 gene in myogenic differentiation

Abstract:

Objective：To observe Lrtm1 expression in C2C12 cells，to construct a lentiviral vector overexpressing Lrtm1 gene effectively in C2C12 cells，to determine its role in myoblast differentiation and to explore the mechanism by which the expression level of Lrtm1 protein is regulated. Methods：The differentiation of C2C12 cells was induced and the nucleus was stained by Hoechst. The mouse Lrtm1 gene was recombined with lentiviral vectors pLJM1-GFP. C2C12 cells which overexpress ectopic Lrtm1 were selected. After treating with C2C12-Lrtm1-DDK cell lines with MG132，the expression of endogenous Lrtm1 protein was observed. Results：Myotubes had formed during its differentiation. The expression of Lrmt1 and Myosin mRNA increased during differentiation.The recombinant pLJM1-Lrtm1-DDK vector was successfully constructed and stable cell lines were screened. The results of real-time PCR at 144 h showed that the Myogenin mRNA level in the Lrtm1-DDK group（272.1±18.22） was lower than that in the GFP group（332.41±41.21）（P<0.05）. The Myosin mRNA level was significantly lower in Lrtm1-DDK group（76.11±10.47） than in GFP group（145.51±10.02） （P＜0.05）. Lrtm1 protein was degraded by the proteasome in C2C12 cells. Conclusion：Lrtm1 gene is highly expressed during myogenic differentiation. Overexpression of Lrtm1 partially inhibits the expression of Myogenin and Myosin. Lrtm1 protein is degraded by the proteasome.