Coordinate gene expression via nucleosome remodeling and chromosomal looping

描述(由申请人提供)：在这项提案中，我们首次研究了一种名为核小体重塑的表观遗传学机制，以及它如何调节可卡因诱导记忆形成所需的协调基因表达。核小体是染色质的重复单位，是基因组DNA紧凑的基础。核小体重塑复合体通过将核小体重新定位在基因启动子上来改变染色质结构并调节表达。最近的人类外显子组测序研究发现，在散发性智力低下和散发性自闭症中，多态的BAF复合体(哺乳动物的SWI/SNF核小体重塑复合体)的亚单位经常发生突变。此外，神经元特异性BRG1相关因子(NBAF)核小体重塑复合体各亚单位的从头突变与Coffin-Siris和Nicolaides-Baraitser综合征有关，这两种综合征都与智能障碍有关。综上所述，这些研究表明nBAF功能是正常认知功能所必需的。虽然在其他领域(如酵母遗传学和癌症)，核小体重塑是一个重要的话题，但在神经科学中，核小体重塑很少受到关注。然而，一项重大发现是鉴定了第一个神经元特异性的BAF复合体，随后发现它调节前体细胞向终末分化神经元转化所需的基因表达。重要的是，nBAF复合体有一个亚基BAF53b，它参与使nBAF具有神经元特异性。该亚基是神经元和nBAF复合体的特异性亚基，是研究nBAF对突触生理和行为的潜在贡献的理想靶点。在这一点的基础上，我们建议测试这一假设，即BAF53b在发育过程中对神经元命运决定起关键作用后，继续调控基因表达，并以一种对成人记忆过程以及可卡因诱导的记忆形成至关重要的方式做到这一点。我们提出了三个具体目标来检验这一假设。在具体目标1中，我们将使用转基因小鼠来检测BAF53b在长期记忆中的作用。在具体目标2中，我们将使用下一代测序、RNA序列和染色体构象捕获3C来确定在记忆巩固过程中BAF53b和染色质环调控哪些基因表达谱。在具体目标3中，我们将确定可卡因如何通过依赖BAF53b的核小体重塑和染色质环在可卡因诱导的记忆形成过程中调节协调基因的表达。总之，在这些特定目标下的工作将阐明BAF53b和nBAF复合体对记忆过程的贡献，更具体地说，对可卡因诱导的记忆形成作为持续药物寻找行为的前体事件的贡献。

DESCRIPTION (provided by applicant): In this proposal, we examine for the first time an epigenetic mechanism called nucleosome remodeling and how it regulates coordinate gene expression required for cocaine-induced memory formation. The nucleosome is the repeating unit of chromatin and fundamental to the compaction of genomic DNA. Nucleosome remodeling complexes modify chromatin structure and regulate expression by repositioning nucleosomes at the promoters of genes. Recent human exome sequencing studies have identified subunits of the polymorphic BAF complexes (mammalian SWI/SNF nucleosome remodeling complex) that are frequently mutated in sporadic mental retardation and sporadic autism. Moreover, de novo mutations in various subunits of neuron-specific Brg1- associated factor (nBAF) nucleosome remodeling complex have been implicated in Coffin-Siris and Nicolaides-Baraitser syndromes, both of which are associated with intellectual disability. Together, these studies suggest that nBAF function is necessary for normal cognitive function. Although an important topic in other fields (e.g. yeast genetics and cancer), nucleosome remodeling has received little attention in neuroscience. However, a major discovery was the identification of the first neuron-specific BAF complex, which was subsequently found to regulate gene expression required for the conversion of precursor cells into terminally differentiated neurons. Importantly, the nBAF complex has a subunit, BAF53b, which participates in making nBAF neuron- specific. This subunit is both neuron and nBAF complex specific, making it an ideal target for investigating the potential contributions of nBAF to synaptic physiology and behavior. Building on this point, we propose to test the hypothesis that BAF53b, after playing a key role in neuronal fate decisions during development, continues to regulate gene expression and does so in a manner critical to adult memory processes as well as cocaine-induced memory formation. We propose three specific aims to test this hypothesis. In Specific Aim 1, we will use genetically modified mice to examine the role of BAF53b in long-term memory. In Specific Aim 2, we will use next generation sequencing, RNA seq, and chromosomal conformation capture 3C to determine what gene expression profiles are being regulated by BAF53b and chromatin looping during memory consolidation. In Specific Aim 3, we will determine how cocaine regulates coordinate gene expression via BAF53b-dependent nucleosome remodeling and chromatin looping during cocaine-induced memory formation. Together, the work under these specific aims will elucidate the contributions of BAF53b and the nBAF complex in general, to memory processes, and more specifically to cocaine-induced memory formation as a precursor event to persistent drug-seeking behavior.