Proteomic study of calpeptin-induced differentiation on calpain-interacting proteins of C2C12 myoblast.

Studies on skeletal muscle cell specification and development have demonstrated in the past that calpains interact with various transcriptional factors in regulating the cellular function. It has therefore, been assumed that transcriptional factors like myogenin, MyoD, Myf5, and MRF4 that are active during the myogenic differentiation might be affected and degraded by calpains. Therefore, to examine the biochemical adaptations of myoblasts during myocyte formation and muscle development comprehensively, the current study was designed to identify the effect of calpeptin (calpain inhibitors) on protein expression during differentiation of C2C12 mouse myoblast. Cells were proliferated to near 80% confluence under Dulbecco's modified eagle medium and differentiated further in 2% HS with 50 μM calpeptin. Incubated cells were collected at 0, 12, and 72 h and later the cell proteins were focused onto pH 4-7 IEF strip, followed by 12.5% SDS-PAGE. Obtained spots on the gels were compared and matched using commercial 2-DE analysis software and matched spots were identified by MALDI-ToF and/or Q-Tof systems. Conclusively, cell differentiation was observed to be active from 12 to 72 h however, calpeptin affected the differentiation process and cut down the rate of fusion by approximately 50%. Out of 41 proteins identified, 12 proteins were found to be upregulated where as 29 proteins were downregulated.