Objective-Regulatory complexes comprising myocardin and serum response factor (SRF) are critical for the transcriptional regulation of many smooth muscle-specific genes. However, little is known about the epigenetic mechanisms that regulate the activity of these complexes. In the current study, we investigated the role of SWI/SNF ATP-dependent chromatin remodeling enzymes in regulating the myogenic activity of myocardin. Methods and Results-We found that both Brg1 and Brm are required for maintaining expression of several smooth muscle-specific genes in primary cultures of aortic smooth muscle cells. Furthermore, the ability of myocardin to induce expression of smooth muscle-specific genes is abrogated in cells expressing dominant negative Brg1. In SW13 cells, which lack endogenous Brg1 and Brm1, myocardin is unable to induce expression of smooth muscle-specific genes. Whereas, reconstitution of wild-type, or bromodomain mutant forms Brg1 or Brm1, into SW13 cells restored their responsiveness to myocardin. SWI/SNF complexes were found to be required for myocardin to increase SRF binding to the promoters of smooth muscle-specific genes. Brg1 and Brm directly bind to the N terminus of myocardin, in vitro, through their ATPase domains and Brg1 forms a complex with SRF and myocardin in vivo in smooth muscle cells. Conclusion-These data demonstrate that the ability of myocardin to induce smooth muscle-specific gene expression is dependent on its interaction with SWI/SNF ATP-dependent chromatin remodeling complexes. Key Words: Brg1 Ⅲ Brm Ⅲ telokin Ⅲ calponin Ⅲ SRF S mooth muscle cells are important contractile components of the cardiovascular system that regulate blood pressure and flow. Vascular smooth muscle cells modulate their phenotype in response to extracellular cues during the development and progression of a variety of diseases including atherosclerosis and hypertension. These diseases are associated with decreased expression of proteins required for the normal contractile function of smooth muscle cells. 1 Understanding the mechanisms that control expression of contractile and regulatory proteins in smooth muscle cells is, therefore, an essential step toward determining how these processes are altered in pathological conditions. The interaction of serum response factor (SRF) with the coactivator myocardin is a critical determinant of vascular smooth muscle development. 2,3 Myocardin null mice lack differentiated smooth muscle cells in the dosal aorta and placental vasculature and die around E10. 2 Myocardin is thus critically required for the differentiation of these populations of vascular smooth muscle cells. Myocardin does not bind directly to DNA, but interacts with genes via its binding to SRF through a basic domain and polyglutamine-rich (poly Q) domain in myocardin. Myocardin-bound SRF binds to CArG elements within the promoters of many smooth musclespecific genes 4 and myocardin activates transcription of these genes through a strong transcriptional activation domain at its C terminus. 5 However, a...

Rodenberg JM, Hoggatt AM, Chen M, Touw K, Jones R, Herring BP. Regulation of serum response factor activity and smooth muscle cell apoptosis by chromodomain helicase DNA-binding protein 8. Serum response factor (SRF) is a widely expressed protein that plays a key role in the regulation of smooth muscle differentiation, proliferation, migration, and apoptosis. It is generally accepted that one mechanism by which SRF regulates these diverse functions is through pathwayspecific cofactor interactions. A novel SRF cofactor, chromodomain helicase DNA binding protein 8 (CHD8), was isolated from a yeast two-hybrid screen using SRF as bait. CHD8 is highly expressed in adult smooth muscle tissues. Coimmunoprecipitation assays from A10 smooth muscle cells demonstrated binding of endogenous SRF and CHD8. Data from GST-pulldown assays indicate that the NH2terminus of CHD8 can interact directly with the MADS domain of SRF. Adenoviral-mediated knockdown of CHD8 in smooth muscle cells resulted in attenuated expression of SRF-dependent, smooth muscle-specific genes. Knockdown of CHD8, SRF, or CTCF, a previously described binding partner of CHD8, in A10 VSMCs also resulted in a marked induction of apoptosis. Mechanistically, apoptosis induced by CHD8 knockdown was accompanied by attenuated expression of the anti-apoptotic proteins, Birc5, and CARD10, whereas SRF knockdown attenuated expression of CARD10 and Mcl-1, but not Birc5, and CTCF knockdown attenuated expression of Birc5. These data suggest that CHD8 plays a dual role in smooth muscle cells modulating SRF activity toward differentiation genes and promoting cell survival through interactions with both SRF and CTCF to regulate expression of Birc5 and CARD10.