The Function of MyoD Binding to the Id2 Promoter

Id2 (inhibitor of differentiation 2) is a helix-loop-helix (HLH) protein that inhibits the function of MyoD, a muscle-specific transcription factor that promotes expression of skeletal muscle-specific genes. Id2 is expressed in proliferating myoblasts and is down-regulated during differentiation. Genome-wide analysis of MyoD binding suggests that MyoD may regulate the expression of genes that are not muscle-specific, as evidenced by MyoD binding at the Id2 promoter in undifferentiated myoblasts. We hypothesized that MyoD influences the levels of Id2 mRNA expression. We compared Id2 promoter efficiency, cellular proliferation rate and Id2 mRNA accumulation in cells expressing MyoD from a transgene vs. parent non-muscle cells.  Id2 promoter activity was down-regulated from growth to differentiation-inducing conditions ~7-fold in C2C12 myoblasts and 2-fold in  10T1/2 fibroblasts as determined by dual luciferase promoter/reporter assay. The proliferation rate was not significantly different between cell lines expressing MyoD and cells not expressing MyoD, as determined by analysis of cell number in culture over time.  The doubling times of 10T ½ fibroblasts +/- MyoD and 3T3 fibroblasts +/- MyoD were 22.75  and 16.84  hours, respectively. qRT-PCR analysis showed  no consistent difference in levels of Id2 mRNA accumulation between pairs of MyoD expressing and non-expressing cells. These results suggest that MyoD may not be affecting Id2 expression in proliferating cells. Further research to determine the function of MyoD binding to the Id2 promoter is needed. Our research will provide a better understanding of the contributions of MyoD and Id2 in normal muscle development.