Remodeling-independent function of the BAF Complex in T Cells and Beyond

描述（由申请人提供）：染色质重塑复合物可以使用ATP水解的能量来破坏核小体并调节基因表达。BAF复合物是一种典型的哺乳动物重塑蛋白，含有约10个亚基，包括重塑所必需的催化亚基Brg。我们发现，在T细胞中删除Brg会损害CD4表达和免疫耐受。令人惊讶的是，无催化活性的Brg点突变体有效地挽救了这两种缺陷，表明Brg和因此BAF复合物具有重塑独立功能（RIF），这与10个BAF亚基中的6个在体外被重塑的事实一致。RIF的分子机制是一个谜。为了解决这个问题，我们将确定RIF对靶基因的物理结构的影响，使用CD4基因座作为模型（目的1）。作为一种补充方法，我们将使用Brg缺失的肿瘤细胞系来鉴定BAF亚基及其介导RIF的功能结构域，重点是肌动蛋白和肌动蛋白相关蛋白BAF 53，一对令人困惑的亚基（Aim 2）。我们的研究挑战了关于染色质建模者的十年之久的教条，并且引起了普遍的兴趣，因为有证据表明组蛋白修饰酶也具有RIF样活性，并且因为核肌动蛋白的作用吸引了几代生物学家，但在很大程度上逃避了他们。

DESCRIPTION (provided by applicant): Chromatin remodeling complexes can use energy of ATP hydrolysis to disrupt nucleosomes and regulate gene expression. BAF complex is a prototypical mammalian remodeler, containing ~10 subunits including the catalytic subunit Brg essential for remodeling. We found that deleting Brg in T cells impairs CD4 expression and immune tolerance. Surprisingly, a catalytically inactive Brg point mutant effectively rescues both defects, indicating Brg and hence BAF complex possess Remodeling Independent Function (RIF), which is consistent with the fact that six of 10 BAF subunits are dispensable for remodeling in vitro. The molecular mechanism of RIF is a mystery. To attack this problem, we will determine the effects of RIF on the physical structure of target genes, using the CD4 locus as a model (Aim 1). As a complementary approach, we will use Brg-deleted tumor cell lines to identify the BAF subunits and their functional domains mediating RIF, focusing on actin and the actin-related protein BAF53, a puzzling pair of subunits (Aim 2). Our study challenges a decade-old dogma about chromatin-modelers, and is of general interest because evidence is emerging that histone modifying enzymes also possess RIF-like activities and because the roles of nuclear actin have fascinated but largely eluded generations of biologists.