CaM-regulated chromatin remodeling: mechanisms, generality and in vivo functions

描述（由申请人提供）：哺乳动物染色质重塑BAF复合物控制着多种生物过程，通常是对环境信号的响应。众所周知，这些信号可以调节BAF向靶基因的募集。例如，巨噬细胞中的TLR4信号传导后，BAF复合物被招募到许多TLR4靶基因上，在那里它重塑其染色质以促进基因表达。出乎意料的是，我们发现BAF募集不足以重塑TLR4靶基因的一个子集；另外还需要钙/钙调蛋白（CaM）传递的第二种信号。CaM直接结合BAF复合物刺激重塑而不影响BAF的募集。这是CaM第一次被证明调节染色质重塑子，也是第一次发现一个信号通路独立于募集而调节重塑子。在这项资助中，我们将解决CaM如何刺激重塑（Aim 1），这目前是一个完全的谜。我们的假设是，重塑的一些抑制因子与CaM依赖的BAF靶基因相连，CaM使用BAF复合物搭便车到靶基因，在那里它使抑制因子失活，从而允许染色质重塑。我们将使用分子遗传学方法在巨噬细胞中测试这些想法。鉴于CaM和BAF都广泛表达，该资助还将验证在巨噬细胞中发现的CaM- BAF信号通路具有普遍重要性的观点（Aim 2）。我们的初步数据表明，CaM- BAF途径确实在T细胞中起作用，我们将通过携带阻断CaM- BAF相互作用的点突变的小鼠进一步探索这一点。我们的研究将揭示新的钙-CaM- BAF通路的机制，普遍性和生物学重要性，以及信号和染色质领域的兴趣。

DESCRIPTION (provided by applicant): The mammalian chromatin---remodeling BAF complex controls diverse biological processes, often in response to environmental signals. Such signals are well known to regulate BAF recruitment to target genes. For example, upon TLR4 signaling in macrophages, the BAF complex is recruited to many TLR4 target genes, where it remodels their chromatin to promote gene expression. Unexpectedly, we have found that BAF recruitment is insufficient for remodeling a subset of TLR4 target genes; a second signal, delivered by calcium/calmodulin (CaM), is additionally required. CaM directly binds the BAF complex to stimulate remodeling without affecting BAF recruitment. This is the first time CaM is shown to regulate a chromatin remodeler, and the first time a signaling pathway is known to regulate a remodeler independently of recruitment. In this grant, we will address how CaM stimulates remodeling (Aim 1), which is currently a total mystery. Our hypothesis is that some repressors of remodeling are tethered to CaM---dependent BAF target genes, and CaM uses the BAF complex to hitchhike to the target genes, where it inactivates the repressor to allow chromatin remodeling. We will test these ideas in macrophages, using molecular genetic methods. This grant will also test the idea that the CaM---BAF signaling pathway found in macrophages is of general importance, given that both CaM and BAF are widely expressed (Aim 2). Our preliminary data suggest the CaM---BAF pathway indeed operates in T cells, which we will further explore using mice bearing point mutations that block CaM---BAF interaction. Our study will shed light on the mechanisms, generality and biological importance of the novel Calcium---CaM---BAF pathway, and of interest to both signaling and chromatin fields.