Department of Molecular Biology and Biochemistry, Wesleyan University, Middletown, CT 06459, USA.
Department of Molecular Biology and Biochemistry, Wesleyan University, Middletown, CT 06459, USA; Department of Biochemistry and Molecular Biotechnology, University of Massachusetts Chan Medical School, Worcester, MA 01605, USA.
Biochim Biophys Acta Gene Regul Mech. 2022 Feb;1865(2):194801. doi: 10.1016/j.bbagrm.2022.194801. Epub 2022 Feb 23.
The mammalian SWItch/Sucrose Non-Fermentable (mSWI/SNF) families of ATP-dependent chromatin remodeling enzymes are established co-regulators of gene expression. mSWI/SNF complexes can be assembled into three major subfamilies: BAF (BRG1 or BRM-Associated Factor), PBAF (Polybromo containing BAF), or ncBAF (non-canonical BAF) that are distinguished by the presence of mutually exclusive subunits. The mechanisms by which each subfamily contributes to the establishment or function of specific cell lineages are poorly understood. Here, we determined the contributions of the BAF, ncBAF, and PBAF complexes to myoblast proliferation via knock down (KD) of distinguishing subunits from each complex. KD of subunits unique to the BAF or the ncBAF complexes reduced myoblast proliferation rate, while KD of PBAF-specific subunits did not affect proliferation. RNA-seq from proliferating KD myoblasts targeting Baf250A (BAF complex), Brd9 (ncBAF complex), or Baf180 (PBAF complex) showed mis-regulation of a limited number of genes. KD of Baf250A specifically reduced the expression of Pax7, which is required for myoblast proliferation, concomitant with decreased binding of Baf250A to and impaired chromatin remodeling at the Pax7 gene promoter. Although Brd9 also bound to the Pax7 promoter, suggesting occupancy by the ncBAF complex, no changes were detected in Pax7 gene expression, Pax7 protein expression or chromatin remodeling at the Pax7 promoter upon Brd9 KD. The data indicate that the BAF subfamily of the mSWI/SNF enzymes is specifically required for myoblast proliferation via regulation of Pax7 expression.
哺乳动物 SWItch/Sucrose Non-Fermentable (mSWI/SNF) 家族的 ATP 依赖性染色质重塑酶是基因表达的既定共调节因子。mSWI/SNF 复合物可以组装成三个主要亚家族：BAF（BRG1 或 BRM 相关因子）、PBAF（含多溴的 BAF）或 ncBAF（非典型 BAF），它们通过相互排斥的亚基的存在来区分。每个亚家族对特定细胞谱系的建立或功能的贡献机制知之甚少。在这里，我们通过从每个复合物中敲除区分亚基来确定 BAF、ncBAF 和 PBAF 复合物对成肌细胞增殖的贡献。BAF 或 ncBAF 复合物特有的亚基的敲除降低了成肌细胞的增殖率，而 PBAF 特异性亚基的敲除则不影响增殖。针对 Baf250A（BAF 复合物）、Brd9（ncBAF 复合物）或 Baf180（PBAF 复合物）的增殖性 KD 成肌细胞的 RNA-seq 显示出少数基因的失调。Baf250A 的敲除特异性降低了 Pax7 的表达，这是成肌细胞增殖所必需的，同时降低了 Baf250A 的结合，并损害了 Pax7 基因启动子的染色质重塑。虽然 Brd9 也与 Pax7 启动子结合，表明 ncBAF 复合物的占据，但在 Brd9 KD 时，Pax7 基因表达、Pax7 蛋白表达或 Pax7 启动子的染色质重塑没有变化。数据表明，mSWI/SNF 酶的 BAF 亚家族通过调节 Pax7 表达特异性地需要成肌细胞增殖。
