Myogenin expression is independent of agrin treatment in C2C12 cell culture

Skeletal muscle differentiation is influenced by myogenic regulatory factors, including MyoD and myogenin. Previous studies have reported that MyoD is expressed prior to myogenin. Our objective was to investigate this process using C2C12 cell culture. When C2C12 myoblasts proliferate to 80% density, growth medium (GM) is replaced with differentiation medium (DM). Subsequently, myoblasts fuse into myotubes within 72 hours in DM. During myotube formation, acetylcholine receptors (AChRs) cluster spontaneously. Treatment with motor neuron derived agrin increases the frequency of AChR clusters through an agrin signaling pathway that also clusters other postsynaptic components of the neuromuscular synapse. Using immunofluorescence, we observed MyoD in myoblast nuclei while myoblasts were in GM, with elimination of MyoD within 72 hours in DM. In contrast, we did not observe myogenin while myoblasts were in GM, with or without agrin treatment, but myogenin was observed in myotube nuclei within 24 hours in DM without agrin treatment. We conclude that myogenin expression is independent of agrin treatment in C2C12 cell culture. Wade A. Grow was supported by Midwestern University intramural funds.