Analysis of Shank3 Complete and Temporal and Spatial Specific Knockout Mice

Shank3 完全敲除小鼠和时空特异性敲除小鼠的分析

• 批准号: 8346356
• 负责人: YONG-HUI JIANG
• 金额: $ 48.14万
• 依托单位: DUKE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-06-01 至 2017-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8346356
• 关键词: 22q13.3; Affect; Age; Animal Model; Behavior; Behavior assessment; Behavioral; Biochemical; Brain region; Characteristics; Child; Clinical; Code; Complex; Corpus striatum structure; Defect; Development; Disease; Etiology; Excitatory Synapse; Exons; Functional disorder; Gene Mutation; Genes; Genetic; Glutamate Receptor; Glutamates; Heterogeneity; Hippocampus (Brain); Human; Impairment; Intellectual functioning disability; Knock-out; Knockout Mice; Knowledge; Language Delays; Lead; Mediating; Methods; Modeling; Molecular; Molecular Genetics; Mus; Mutant Strains Mice; Mutation; Neocortex; Neurobiology; Neurologic; Pathogenesis; Patients; Phenotype; Point Mutation; Positioning Attribute; Protein Isoforms; Protocols documentation; Public Health; RNA Splicing; Reporting; Scaffolding Protein; Signal Transduction; Staging; Synapses; Syndrome; Testing; Tissues; Triad Acrylic Resin; Work; autism spectrum disorder; base; clinical Diagnosis; density; design; gain of function; insight; interdisciplinary approach; loss of function; microdeletion; mouse model; novel therapeutic intervention; postsynaptic; promoter; receptor; response; social communication; tool

DESCRIPTION (provided by applicant): Autism spectrum disorder (ASD) is a major public health problem affecting 1 out 110 children. There is a fundamental gap in understanding the cellular and molecular mechanisms underlying the diverse clinical presentation of ASD. Understanding these mechanisms is critical for developing novel therapeutic approaches. The limitations inherent in human studies make it difficult to study cellular and molecular mechanisms, providing a rationale for the study of animal models. Recent genetic evidence implicates the SHANK3 gene in ASD. SHANK3 is a scaffolding protein that organizes a signaling complex at postsynaptic density of excitatory synapses. An array of SHANK3 isoforms result from 6 alternative promoters and splicing of coding exons. Deletion of the SHANK3 gene is a major contributor to ASD features in the 22q13.3 deletion Phelan-McDermid syndrome. Microdeletions of the entire SHANK3 gene and point mutations of SHANK3 disrupting specific isoforms have been identified in patients with ASD and intellectual disability. We and others reported Shank3 isoform-knockout mice with different exonic deletions. Reduced postsynaptic response of excitatory synapses in hippocampus, striatum, and neocortex as well as ASD-like behaviors is found in these mice, but there were also notable phenotypic differences. This phenotypic heterogeneity could be explained by the different impact that each mutation has on isoform-specific expression of Shank3. However, interpretation of these differences is complicated by the fact that the existing mutant mice are not Shank3 complete knockout and were analyzed in different brain regions and using different protocols. Because >95% of SHANK3 molecular defects in humans delete the entire SHANK3 gene, and these patients have more severe phenotypes than those with point mutations of SHANK3, we have generated Shank3 complete knockout mice by deleting exons 4-22, and also Shank3 conditional knockout mice, with floxed exons 4-22. These models are more valid for dissecting the cellular mechanism arising from SHANK3 deletion than existing isoform-knockout mice. The objective of this proposal is to analyze Shank3 complete knockout mice using an interdisciplinary approach. The central hypothesis is that the primary path by which SHANK3 molecular defects lead to ASD in humans is via cellular and synaptic defects of reduced glutamatergic receptor-mediated postsynaptic responses in different brain regions. With these mutant mice, we are uniquely positioned to study the following Specific Aims: 1) To model major clinical features of SHANK3 deletion in Shank3 complete knockout mice; 2). To delineate the cellular and molecular mechanism underlying Shank3 deficiency; 3) To analyze the temporal- and spatial-requirements of SHANK3 deficiency underlying the pathogenesis of ASD. The proposed studies are significant because analysis of Shank3 complete knockout mice may uncover the cellular and circuit mechanism of ASD caused by SHANK3 deletion. The knowledge gained from studying Shank3 will provide insights that may be generalizable and applicable to understanding the pathophysiology of other causes of ASD. PUBLIC HEALTH RELEVANCE: The proposed project is to understand the cellular and synaptic mechanism underlying autism spectrum disorder by analyzing Shank3 complete knockout mice. This project will lead to better understanding the pathogenesis and developing treatment for ASD.

描述（由申请人提供）：自闭症谱系障碍 (ASD) 是一个重大的公共卫生问题，影响着每 110 名儿童中就有 1 名。对于 ASD 不同临床表现背后的细胞和分子机制的理解存在着根本性的差距。了解这些机制对于开发新的治疗方法至关重要。人类研究固有的局限性使得研究细胞和分子机制变得困难，这为动物模型的研究提供了理论基础。最近的遗传证据表明 SHANK3 基因与自闭症谱系障碍有关。 SHANK3 是一种支架蛋白，在兴奋性突触的突触后密度处组织信号复合物。一系列 SHANK3 同工型由 6 个替代启动子和编码外显子剪接产生。 SHANK3 基因的缺失是 22q13.3 缺失 Phelan-McDermid 综合征中 ASD 特征的主要原因。在自闭症谱系障碍 (ASD) 和智力障碍患者中已经发现了整个 SHANK3 基因的微缺失和破坏特定亚型的 SHANK3 点突变。我们和其他人报道了具有不同外显子缺失的 Shank3 同工型敲除小鼠。在这些小鼠中发现海马、纹状体和新皮质的兴奋性突触的突触后反应减少以及 ASD 样行为，但也存在显着的表型差异。这种表型异质性可以通过每个突变对 Shank3 亚型特异性表达的不同影响来解释。然而，由于现有的突变小鼠并未完全敲除 Shank3，并且在不同的大脑区域并使用不同的方案进行分析，因此对这些差异的解释变得复杂。由于人类中>95%的SHANK3分子缺陷删除了整个SHANK3基因，并且这些患者比SHANK3点突变的患者具有更严重的表型，因此我们通过删除外显子4-22产生了Shank3完全敲除小鼠，也产生了Shank3条件敲除小鼠小鼠，具有固定的外显子 4-22。这些模型比现有的异构体敲除小鼠更能有效地剖析 SHANK3 缺失引起的细胞机制。本提案的目的是使用跨学科方法分析 Shank3 完全敲除小鼠。核心假设是，SHANK3 分子缺陷导致人类 ASD 的主要途径是通过不同大脑区域谷氨酸受体介导的突触后反应减少的细胞和突触缺陷。通过这些突变小鼠，我们具有独特的优势来研究以下具体目标：1）模拟 Shank3 完全敲除小鼠中 SHANK3 缺失的主要临床特征； 2）。描述 Shank3 缺陷的细胞和分子机制； 3) 分析自闭症谱系障碍发病机制中SHANK3缺陷的时间和空间需求。所提出的研究意义重大，因为对 Shank3 完全敲除小鼠的分析可能揭示 SHANK3 缺失引起的 ASD 的细胞和回路机制。通过研究 Shank3 获得的知识将提供可推广并适用于理解自闭症谱系障碍其他原因的病理生理学的见解。 公共健康相关性：拟议的项目是通过分析 Shank3 完全基因敲除小鼠来了解自闭症谱系障碍的细胞和突触机制。该项目将有助于更好地了解 ASD 的发病机制并开发治疗方法。