Transcriptome analysis of Shank3-heterozygous and -homozygous mice in different brain regions

Shank3 is an abundant excitatory postsynaptic scaffolding proteins implicated in various neurodevelopmental and psychiatric disorders, including ASD, Phelan-McDermid syndrome, intellectual disability, and schizophrenia. Shank3-mutant mice with a homozygous deletion of exons 14-16 (Shank3-HM mice) show ASD-like behavioral deficits and altered synaptic and neuronal functions, but little is known about how different ages, brain regions, and gene dosages contribute to transcriptomic phenotypes in these mice. Here, we performed RNA-Seq-based transcriptomic analyses of the prefrontal cortex, hippocampus, and striatum in adult Shank3 heterozygous- and homozygous-mutant mice. In addition, juvenile and adult Shank3 homozygous-mutant forebrain transcriptomes were compared. Juvenile and adult forebrain transcriptomes from Shank3 homozygous-mutant mice showed the patterns that are opposite and similar to those observed in ASD: reverse-ASD and ASD-like patterns, respectively. Here, the juvenile reverse-ASD pattern involved synaptic gene upregulations and ribosomal and mitochondrial downregulations, whereas the adult ASD-like pattern involved opposite changes. Gene set enrichment analyses (GSEA) of brain regional transcripts in adult Shank3-HT and Shank3-HM mice revealed that the cortical, hippocampal, and striatal transcripts show distinctly altered biological functions and ASD-related/risk gene expressions. The cortex and striatum display ASD-like patterns whereas the hippocampus displays reverse-ASD patterns. The cortical ASD-like pattern more strongly involves ASD-risk genes whereas the striatal ASD-like pattern more strongly involves astrocyte/microglia genes. Shank3-HT and Shank3-HM transcripts in a given brain region display largely similar patterns in biological functions and ASD-related/risk gene expressions, suggestive of small gene dosage effects. These results suggest that heterozygous and homozygous Shank3 deletions in mice lead to age, brain region, and gene dosage-differential transcriptomic changes. Transcriptome analysis of adult Shank3-heterozygous, -homozygous mice in prefrontal cortex, hippocampus, and striatumShank3是一种丰富的兴奋性突触后支架蛋白，与多种神经发育及精神疾病密切相关，包括孤独症谱系障碍（ASD, Autism Spectrum Disorder）、Phelan-McDermid综合征、智力障碍与精神分裂症。携带外显子14-16纯合缺失的Shank3突变小鼠（Shank3-HM小鼠，Shank3 homozygous mutant mice）可表现出类ASD行为缺陷以及突触与神经元功能异常，但目前对于不同年龄、脑区及基因剂量如何影响此类小鼠的转录组表型仍所知有限。本研究针对成年Shank3杂合突变与纯合突变小鼠的前额叶皮层、海马体与纹状体开展了基于RNA测序（RNA-Seq, RNA Sequencing）的转录组分析，并对幼年与成年Shank3纯合突变小鼠的前脑转录组进行了比较。结果显示，Shank3纯合突变小鼠的幼年与成年前脑转录组分别呈现与ASD相反及相似的表达模式，即反向ASD模式与类ASD模式：其中幼年反向ASD模式表现为突触基因上调、核糖体与线粒体基因下调，而成人类ASD模式则呈现相反的表达变化。对成年Shank3杂合突变（Shank3-HT, Shank3 heterozygous mutant）与纯合突变小鼠的脑区转录本进行基因集富集分析（GSEA, Gene Set Enrichment Analysis）后发现，皮层、海马体与纹状体的转录本存在显著改变的生物学功能及ASD相关/风险基因表达异常：皮层与纹状体呈现类ASD模式，海马体则呈现反向ASD模式；皮层的类ASD模式更显著涉及ASD风险基因，而纹状体的类ASD模式则更显著涉及星形胶质细胞/小胶质细胞基因。特定脑区内Shank3杂合与纯合突变的转录本在生物学功能及ASD相关/风险基因表达上整体呈现相似模式，提示基因剂量效应较弱。上述结果表明，小鼠体内Shank3的杂合与纯合缺失会引发年龄、脑区及基因剂量依赖性的转录组改变。本研究针对成年Shank3杂合突变、纯合突变小鼠的前额叶皮层、海马体与纹状体开展了转录组分析。