Uba1 promotes differentiation, migration, adhesion and spreading of C2C12 cells

Objective To investigate the regulatory function of ubiquitin-like modifier activating enzyme 1 (Uba1) in the growth and development of myoblasts.

Method C2C12 myoblasts were used as a model to establish cell models with Uba1 overexpression and interference. Myogenic differentiation was assessed by immunofluorescence staining for myosin heavy chain (MyHC). Cell migration was analyzed using a wound-healing assay, and cell morphological changes were observed through cell adhesion and spreading assays. The expression of differentiation and migration marker genes was further examined by qRT-PCR and Western blotting.

Result C2C12 cell models with stable Uba1 overexpression and interference were successfully generated. Overexpression of Uba1 significantly increased the number of MyHC-positive cells on day 5 of differentiation (P<0.01), and significantly upregulated the expression of the myogenic differentiation markers MyoG and MyHC at both the mRNA and protein levels (P<0.01). It also promoted cell migration, with a significantly increased migration distance 24 h after scratching (P<0.001) and upregulated the expression of migration-related genes Rac1, Fak, and Paxillin (P<0.05). Furthermore, Uba1 overexpression significantly increased the number of adherent cells (P<0.05) and the average cell spreading area (P<0.001). Conversely, interference of Uba1 produced the opposite effects on all the above parameters.

Conclusion Uba1 promotes the differentiation, migration, adhesion and spreading of C2C12 cells, thereby exerting a positive regulatory role in the growth and development of skeletal muscle.