Inhibition of methionine aminopeptidase in C2C12 myoblasts disrupts cell integrity via increasing endoplasmic reticulum stress.

Proteasome-dependent protein degradation and the digestion of peptides by aminopeptidases are essential for myogenesis. Methionine aminopeptidases (MetAPs) are uniquely involved in, both, the proteasomal degradation of proteins and in the regulation of translation (via involvement in post-translational modification). Suppressing MetAP1 and MetAP2 expression inhibits the myogenic differentiation of C2C12 myoblasts. However, the molecular mechanism by which inhibiting MetAPs impairs cellular function remains to be elucidated. Here, we provide evidence for our hypothesis that MetAPs regulate proteostasis and that their inhibition increases ER stress by disrupting the post-translational modification, and thereby compromises cell integrity. Thus, using C2C12 myoblasts, we investigate the effect of inhibiting MetAPs on cell proliferation and the molecular mechanisms underpinning its effects. We found that exposure to bengamide B (a MetAP inhibitor) caused C2C12 myoblasts to lose their proliferative abilities via cell cycle arrest. The underlying mechanism involved the accumulation of abnormal proteins (due to the decrease in the N-terminal methionine removal function) which led to increased endoplasmic reticulum stress, decreased protein synthesis, and a protective activation of the autophagy pathway. To identify the MetAP involved in these effects, we use siRNAs to specifically knockdown MetAP1 and MetAP2 expressions. We found that only MetAP2 knockdown mimicked the effects seen with bengamide B treatment. Thus, we suggest that MetAP2, rather than MetAP1, is involved in maintaining the integrity of C2C12 myoblasts. Our results are useful in understanding muscle regeneration, obesity, and overeating disorders. It will help guide new treatment strategies for these disorders.