Our work helped to move the understanding of the mechanisms by which MyoD activates transcription from muscle loci forward. First, we have shown that a subunit of the SWI/SNF chromatin remodeling complex, BAF60c, binds the promoter of an early marker of skeletal muscle differentiation, myogenin, in the absence of catalytic SWI/SNF subunits and before transcription is activated. At this time, BAF60c and MyoD bind the chromatin of myogenic genes together in progenitor cells and they mutually facilitate binding. This pioneering event poises the chromatin for rapid transcriptional activation. Second, a p38-dependent phosphorylation of BAF60c is the key event that senses differentiation at the chromatin level, recruiting an active SWI/SNF complex that will remodel the nucleosome, and allow transcriptional initiation. Our proposed 2-step model for remodeling myogenic loci is in accordance with a stochastic assembly model proposing intermediate interactions for large multisubunit complexes that have also been described for spliceosomes, DNA repair complexes and RNA polymerases with associated transcription factors.24,25
Knowledge of transcriptional regulation has advanced dramatically in the last decade; the identification of the histone code and the integration of genome-wide and proteomic techniques have been crucial for revealing new revolutionary mechanisms of transcriptional regulation. Nevertheless, many aspects of the transcriptional process remain unexplained. In the myogenic transcription some exciting questions remain to be addressed. How is chromatin architecture integrating regulatory factor binding and tethering to the chromatin? If loops are bringing together enhancer and promoter regions, who are the mediators and which are the genes or genomic domains being regulated in this fashion? What are the specific kinetics of regulatory factor binding that modulate transcriptional regulation? Does chromosomal 3D structure vary between proliferating and differentiating myoblasts?
Regarding BAF60c functions, it would be important to identify new BAF60c chromatin target sites; to monitor if BAF60c binds MyoD-independent loci; whether BAF60c chromatin binding is accompanied by other SWI/SNF subunits and its significance/impact for chromatin structure and regulation.
Furthermore, it is crucial to understand the role that different combinations of preassembled SWI/SNF components may play in regulating chromatin architecture and remodeling.
Immunoprecipitation assays followed by liquid chromatography-tandem mass spectrometry (LC-MS/MS) could be used to identify differentiation-stage specific BAF60c-complexes. This approach could also identify post-translational modifications in BAF60c and in other SWI/SNF subunits that can play an important role in transcriptional activation/repression. In addition, the identification of BAF60c-interacting enzymes that promote these modifications could also reveal new regulatory pathways that may be involved in myogenic transcription.
Finally, it would be very interesting to explore in detail how the nucleosomes are remodeled along the myogenin promoter and how nucleosome location genome-wide differs in precursor vs. differentiated cells. Do BAF60c-differential complexes have an impact on DNA accessibility and how?
In summary, continued efforts to investigate these molecular mechanisms are needed and will be crucial for a better understanding of transcriptional regulation.