Neurobiological Mechanism of 15q11-13 Duplication Autism Spectrum Disorder

描述(申请人提供)：该项目的长期目标是比较和对比Ube3a基因剂量改变所产生的神经元mRNA表达和电路功能缺陷，这些缺陷可能是自闭症谱系障碍(ASD)母亲15q11-13重复和Angelman综合征(AS)行为表型的基础。15q11-13ASD和AS患者的细胞分别存在E3泛素连接酶Ube3a的过度和缺陷。UBE3A是唯一一个在大脑中选择性表达的母体等位基因。大多数从母亲那里遗传15q11-13重复的儿童都会患上自闭症，而且遗传的副本越多，损害就越严重。最近，我们发现，增加Ube3a基因剂量本身就构成了自闭症相关行为缺陷的三联体。相比之下，当孩子遗传母体Ube3a基因突变时，就会出现安杰曼综合症(AS)。重要的是，Ube3a还作为核受体共激活因子调节基因的表达，而不依赖于连接酶的活性。因此，我们假设，过量的Ube3a(15q DUP模型)作用于细胞核，扰乱特定神经元基因和编码蛋白的正常表达，从而改变电路功能，从而产生与自闭症相关的行为缺陷。逻辑推论是，核Ube3a的缺陷可能会改变基因表达，从而改变电路功能，并通过相同但相反的机制产生与Angelman相关的行为缺陷。因为大脑的电路是为定量评估感觉信息和定量的运动反应而设计的，所以很可能许多控制电路功能的分子在它们的数量发生变化时会导致行为的渐进性变化。我们建议Ube3a通过对神经元基因表达的剂量依赖调节来做到这一点。这种分子数量的变化可能是ASD等行为障碍内在异质性的基础。这笔R01拨款的具体目标如下：(1和2)确定Ube3a基因剂量是否通过其在核内的作用导致ASD(Ube3a过多)和AS(Ube3a不足)相关的行为缺陷；(3)确定Ube3a过多或不足对皮质神经元mRNAs的定量调控；以及(4)定量确定Ube3a过多和不足导致的皮质回路功能障碍。我们的方法包括新的核靶向Ube3a转基因，Ube3a的神经细胞类型特异性表达，全基因组转录本的分析，以及共同的Ube3a基因启动子结合位点的鉴定。切片电生理学和神经元形态将检测Ube3a基因剂量的全局和细胞类型特异性变化以及核靶向性变化对活体脑片皮质回路功能的影响。这一分析将有助于我们在这两种截然不同的人类神经疾病中从基因到电路再到行为的桥梁。该项目促进了该机构的使命，通过遗传模型进一步深入了解ASD和AS中涉及的神经细胞、电路和基因。新的分子洞察力和基因工具将促进这些终身行为治疗方法的开发 残疾人士。

DESCRIPTION (provided by applicant): The long-term objective of this project is to compare and contrast the defects in neuronal mRNA expression and circuit functions produced by altered Ube3a gene dosage that may underlie the behavioral phenotypes of Autism Spectrum Disorder (ASD) with maternal 15q11-13 duplication and Angelman's Syndrome (AS). Cells from 15q11-13 ASD and AS patients have excess and deficient E3 ubiquitin ligase, Ube3a, respectively. Ube3a is the sole gene expressed selectively from the maternal allele in brain. The majority of children who inherit 15q11-13 duplications from their mothers develop ASD and the more copies inherited, the more severe the impairment. Recently, we established that increasing Ube3a gene dosage alone reconstitutes the triad of autism-related behavioral deficits in mice. Angelman's Syndrome (AS), by contrast, results when children inherit maternal Ube3a gene mutations. Importantly, Ube3a also acts as a nuclear receptor co-activator regulating gene expression independent of ligase activity. Thus we hypothesized that excess Ube3a (15q dup model) acts in the nucleus to perturb the normal expression of specific neuronal genes and encoded proteins to alter circuit function and thereby generate autism-related behavioral deficits. The logical corollary is that deficiencies of nuclear Ube3a may alter gene expression to change circuit function and produce Angelman-related behavioral deficits through the same, but opposite mechanisms. Because the brain's circuitries are designed for quantitative assessment of sensory information and quantitative motor responses, it is likely that many molecules controlling circuit functions can cause graded changes in behavior when their quantities are altered. We propose Ube3a do so through a dose-dependent regulation of neuronal gene expression. Such molecular quantity variations likely underlie the intrinsic heterogeneity of behavioral disorders like ASD. The specific aims of this R01 grant follow: (1 and 2) determine if Ube3a gene dosage causes ASD (excess Ube3a) and AS (deficient Ube3a) related behavioral deficits through its actions within the nucleus; (3) identify the cortical neuron mRNAs quantitatively regulated by excesses or deficiencies of Ube3a; and (4) identify the cortical circui dysfunctions quantitatively induced by excesses and deficiencies of Ube3a. Our methods include a novel nuclear-targeted Ube3a transgene, a neuron cell-type specific expression of Ube3a, genome wide analysis of transcripts, and identification of consensus Ube3a-gene promoter binding sites. Slice electrophysiology and neuronal morphology will examine the effects of global and cell-type specific changes and nuclear targeted changes of Ube3a gene dosage on cortical circuit function in living brain slices. This analysis will facilitate our effors to bridge from genes to circuits to behavior in the two contrasting human neurological diseases. The project promotes the agency's mission to further a deeper understanding of the neuronal cells, circuits, and genes involved in ASD and AS via genetic models. The novel molecular insights and genetic tools will facilitate development of treatments for these life-long behavioral disabilities.