Mediator and epigenetic control of neuronal gene expression and differentiation

描述（由研究者提供）：我们的长期目标是了解Mediator在脊椎动物中枢神经系统中基因表达和细胞命运规范控制中的作用和调控。中介是基因特异性转录因子和真核RNA聚合酶II一般转录机制之间的多蛋白界面。在这种情况下，Mediator可以引导激活蛋白和抑制蛋白的调节信号，影响控制多种生理过程的基因表达程序的变化，包括细胞生长和稳态、发育和分化。MED12是xq13编码的230 kDa的Mediator亚基，通过选择性调控神经元基因的表达，在神经元发育过程中发挥重要作用。MED12的遗传变异与神经精神疾病和x连锁智力迟钝（XLMR）有关。然而，MED12通过选择性基因调控控制神经元分化的分子基础，以及病理序列改变影响MED12功能导致行为和认知缺陷的途径尚不清楚。在这方面，我们最近发现了调解中的MED12界面和G9a组蛋白甲基转移酶之间的功能相互作用，这是由RE1沉默转录因子/神经元限制性沉默因子（REST/NRSF）施加的表观遗传沉默所必需的，REST/NRSF是抑制非神经元和神经祖细胞中神经元基因表达的神经元命运的关键决定因素。值得注意的是，我们发现导致两种XLMR疾病（FG综合征和Lujan综合征）的MED12错义突变破坏了其rest特异性协同抑制子功能，从而将人类rest依赖的神经元基因抑制与高阶认知功能联系起来。由于我们最近的研究暗示rest依赖的神经元基因抑制在神经祖细胞分化的表观遗传学限制中，我们的研究结果提供了一个可能的表观遗传学角度来解释MED12通过改变神经元发育在XLMR病因学中的作用。因此，我们假设MED12中的XLMR相关突变破坏了rest对神经元基因表达和神经祖细胞分化施加的表观遗传限制。为了支持这一假说，我们提出以下研究目标，以揭示MED12/Mediator在rest依赖性神经元基因表达和分化的表观遗传抑制中的作用和病理意义：(1)阐明MED12/Mediator在rest依赖性神经元外基因沉默中的机制基础；(2)阐明MED12/Mediator在神经祖细胞rest依赖性神经元基因表达和分化抑制中的作用和调控；(3)阐明MED12中xlmr相关突变对REST依赖抑制神经祖细胞神经元基因表达和分化的影响及其机制。我们期望这些研究对神经系统疾病的细胞替代疗法以及XLMR的病因学具有重要的人类健康意义。公共卫生相关性：我们期望这些研究对神经系统疾病的细胞替代疗法以及人类发育和认知缺陷的病因学具有重要意义。首先，本文提出的阐明MED12/Mediator控制神经元基因表达和分化的机制（Aim 1）和调控（Aim 2）的研究有望开辟新的领域，阐明神经干细胞生物学的基本方面，这将对指导基于细胞的治疗方法重建神经系统受损或病变区域至关重要。其次，本文提出的评估MED12中XLMR相关突变对其基本生化特性和与神经元基因抑制和神经干细胞分化相关的功能相互作用的影响的研究（Aim 3）应该揭示有关XLMR病因的新的机制见解，并可能确定治疗或补救干预的新途径。

DESCRIPTION (provided by investigator): Our long-term goal is to understand the role and regulation of Mediator in the control of gene expression and cell fate specification within the vertebrate central nervous system. Mediator is a multiprotein interface between gene-specific transcription factors and the eukaryotic RNA polymerase II general transcription machinery. In this capacity, Mediator serves to channel regulatory signals from activator and repressor proteins to affect changes in gene expression programs that control diverse physiological processes, including cell growth and homeostasis, development, and differentiation. MED12, an Xq13-encoded 230 kDa Mediator subunit, plays an essential role in neuronal development through selective regulation of neuronal gene expression. Genetic variation in MED12 has been linked to neuropsychiatric illness and X-linked mental retardation (XLMR). However, the molecular bases by which MED12 controls neuronal differentiation through selective gene regulation and the means by which pathologic sequence alterations impact MED12 function leading to behavioral and cognitive defects remain to be clarified. In this regard, we recently identified a functional interaction between the MED12 interface in Mediator and G9a histone methyltransferase required for epigenetic silencing imposed by the RE1 silencing transcription factor/neuron restrictive silencing factor (REST/NRSF), a key determinant of neuronal fate that suppresses neuronal gene expression in non-neuronal and neural progenitor cells. Notably, we found that missense mutations in MED12 responsible for two XLMR disorders, FG syndrome and Lujan syndrome, disrupt its REST-specific corepressor function, thus linking REST-dependent neuronal gene repression with higher-order cognitive function in humans. Because our recent studies implicate REST-dependent neuronal gene repression in epigenetic restriction of neural progenitor cell differentiation, our findings provide a possible epigenetic perspective to explain the role of MED12 in the etiology of XLMR through altered neuronal development. Thus, we hypothesize that XLMR- associated mutations in MED12 disrupt REST-imposed epigenetic restrictions on neuronal gene expression and neural progenitor cell differentiation. To provide support for this hypothesis, we propose the following aims to decipher the role and pathologic implications of MED12/Mediator in REST-dependent epigenetic suppression of neuronal gene expression and differentiation: (1) Elucidate the mechanistic basis of MED12/Mediator in REST-dependent extra-neuronal gene silencing; (2) Elucidate the role and regulation of MED12/Mediator in REST-dependent suppression of neuronal gene expression and differentiation in neural progenitor cells; (3) Elucidate the impact and mechanism of XLMR-associated mutations in MED12 on REST- dependent suppression of neuronal gene expression and differentiation in neural progenitor cells. We expect these studies to have important human health implications for cell replacement therapy in neurological disease as well as the etiology of XLMR. PUBLIC HEALTH RELEVANCE: We expect these studies to have important implications for cell replacement therapy in neurological disease as well as the etiology of developmental and cognitive defects in humans. First, studies proposed herein to elucidate the mechanism (Aim 1) and regulation (Aim 2) of MED12/Mediator in control of neuronal gene expression and differentiation are expected to break new ground and illuminate fundamental aspects of neural stem cell biology that will be essential to guide prospective cell-based therapeutic approaches to repopulate damaged or diseased areas of the nervous system. Second, studies proposed herein to evaluate the impact of XLMR-associated mutations in MED12 on its basic biochemical properties and functional interactions relevant to neuronal gene repression and neural stem cell differentiation (Aim 3) should reveal new mechanistic insight concerning the etiology of XLMR and possibly identify new avenues for therapeutic or remedial intercession.