Sharma Tapan, Olea-Flores Monserrat, Imbalzano Anthony N
Department of Biochemistry and Molecular Biotechnology, UMass Chan Medical School, Worcester, MA, United States.
Front Cell Dev Biol. 2023 Jul 7;11:1160227. doi: 10.3389/fcell.2023.1160227. eCollection 2023.
Skeletal muscle differentiation is a tightly regulated process, and the importance of the mammalian SWI/SNF (mSWI/SNF) chromatin remodeling family for regulation of genes involved in skeletal myogenesis is well-established. Our prior work showed that bromodomains of mSWI/SNF ATPases BRG1 and BRM contribute to myogenesis by facilitating the binding of mSWI/SNF enzymes to regulatory regions of myogenic and other target genes. Here, we report that pathway analyses of differentially expressed genes from that study identified an additional role for mSWI/SNF enzymes via the regulation of the Wnt signaling pathway. The Wnt pathway has been previously shown to be important for skeletal muscle development. To investigate the importance of mSWI/SNF enzymes for the regulation of the Wnt pathway, individual and dual knockdowns were performed for BRG1 and BRM followed by RNA-sequencing. The results show that BRG1, but not BRM, is a regulator of Wnt pathway components and downstream genes. Reactivation of Wnt pathway by stabilization of β-catenin could rescue the defect in myogenic gene expression and differentiation due to BRG1 knockdown or bromodomain inhibition using a specific small molecule inhibitor, PFI-3. These results demonstrate that BRG1 is required upstream of β-catenin function. Chromatin immunoprecipitation of BRG1, BRM and β-catenin at promoters of Wnt pathway component genes showed binding of BRG1 and β-catenin, which provides further mechanistic insight to the transcriptional regulation of these genes.
骨骼肌分化是一个受到严格调控的过程,哺乳动物SWI/SNF(mSWI/SNF)染色质重塑家族在调节参与骨骼肌生成的基因方面的重要性已得到充分证实。我们之前的研究表明,mSWI/SNF ATP酶BRG1和BRM的溴结构域通过促进mSWI/SNF酶与成肌及其他靶基因的调控区域结合,从而有助于肌生成。在此,我们报告,对该研究中差异表达基因进行的通路分析确定了mSWI/SNF酶通过调控Wnt信号通路发挥的额外作用。此前已证明Wnt通路对骨骼肌发育很重要。为了研究mSWI/SNF酶对Wnt通路调控的重要性,对BRG1和BRM进行了单独及双重敲低,随后进行RNA测序。结果表明,BRG1而非BRM是Wnt通路成分及下游基因的调节因子。使用特异性小分子抑制剂PFI-3稳定β-连环蛋白来重新激活Wnt通路,可以挽救因BRG1敲低或溴结构域抑制导致的成肌基因表达和分化缺陷。这些结果表明,BRG1在β-连环蛋白功能的上游发挥作用。对Wnt通路成分基因启动子处的BRG1、BRM和β-连环蛋白进行染色质免疫沉淀,结果显示BRG1和β-连环蛋白存在结合,这为这些基因的转录调控提供了进一步的机制性见解。