Abstract
NF-κB signaling is implicated as an important regulator of skeletal muscle homeostasis, but the mechanisms by which this transcription factor contributes to muscle maturation and turnover remain unclear. To gain insight into these mechanisms, gene expression profiling was examined in C2C12 myoblasts devoid of NF-κB activity. Interestingly, even in proliferating myoblasts, the absence of NF-κB caused the pronounced induction of several myofibrillar genes, suggesting that NF-κB functions as a negative regulator of late-stage muscle differentiation. Although several myofibrillar promoters contain predicted NF-κB binding sites, functional analysis using the troponin-I2 gene as a model revealed that NF-κB-mediated repression does not occur through direct DNA binding. In the search for an indirect mediator, the transcriptional repressor YinYang1 (YY1) was identified. While inducers of NF-κB stimulated YY1 expression in multiple cell types, genetic ablation of the RelA/p65 subunit of NF-κB in both cultured cells and adult skeletal muscle correlated with reduced YY1 transcripts and protein. NF-κB regulation of YY1 occurred at the transcriptional level, mediated by direct binding of the p50/p65 heterodimer complex to the YY1 promoter. Furthermore, YY1 was found associated with multiple myofibrillar promoters in C2C12 myoblasts containing NF-κB activity. Based on these results, we propose that NF-κB regulation of YY1 and transcriptional silencing of myofibrillar genes represent a new mechanism by which NF-κB functions in myoblasts to modulate skeletal muscle differentiation.
SUPPLEMENTAL MATERIAL
We thank members of the Guttridge laboratory for their support and insight throughout the course of this study, especially K. J. Ladner for technical assistance. We also thank T. Huang and A. Chang for assistance with ChIP assays and Y. Shi for the YY1 expression plasmid.
This work was supported by NIH grants CA97953 to D.C.G. and AR054244 to H.W.