Lee Kong Chian School of Medicine, Nanyang Technological University, Singapore 636921, Singapore.
Lee Kong Chian School of Medicine, Nanyang Technological University, Singapore 636921, Singapore; Translational Laboratory in Genetic Medicine (TLGM), Agency for Science, Technology and Research (A∗STAR), Singapore 138648, Singapore.
EBioMedicine. 2023 Aug;94:104720. doi: 10.1016/j.ebiom.2023.104720. Epub 2023 Jul 21.
BACKGROUND: In Huntington's disease (HD), a CAG repeat expansion mutation in the Huntingtin (HTT) gene drives a gain-of-function toxicity that disrupts mRNA processing. Although dysregulation of gene splicing has been shown in human HD post-mortem brain tissue, post-mortem analyses are likely confounded by cell type composition changes in late-stage HD, limiting the ability to identify dysregulation related to early pathogenesis. METHODS: To investigate gene splicing changes in early HD, we performed alternative splicing analyses coupled with a proteogenomics approach to identify early CAG length-associated splicing changes in an established isogenic HD cell model. FINDINGS: We report widespread neuronal differentiation stage- and CAG length-dependent splicing changes, and find an enrichment of RNA processing, neuronal function, and epigenetic modification-related genes with mutant HTT-associated splicing. When integrated with a proteomics dataset, we identified several of these differential splicing events at the protein level. By comparing with human post-mortem and mouse model data, we identified common patterns of altered splicing from embryonic stem cells through to post-mortem striatal tissue. INTERPRETATION: We show that widespread splicing dysregulation in HD occurs in an early cell model of neuronal development. Importantly, we observe HD-associated splicing changes in our HD cell model that were also identified in human HD striatum and mouse model HD striatum, suggesting that splicing-associated pathogenesis possibly occurs early in neuronal development and persists to later stages of disease. Together, our results highlight splicing dysregulation in HD which may lead to disrupted neuronal function and neuropathology. FUNDING: This research is supported by the Lee Kong Chian School of Medicine, Nanyang Technological University Singapore Nanyang Assistant Professorship Start-Up Grant, the Singapore Ministry of Education under its Singapore Ministry of Education Academic Research Fund Tier 1 (RG23/22), the BC Children's Hospital Research Institute Investigator Grant Award (IGAP), and a Scholar Award from the Michael Smith Health Research BC.
背景：在亨廷顿病（HD）中，亨廷顿（HTT）基因中的 CAG 重复扩展突变导致功能获得性毒性，破坏 mRNA 加工。尽管在人类 HD 死后脑组织中已经显示出基因剪接的失调，但死后分析可能受到晚期 HD 中细胞类型组成变化的混淆，限制了识别与早期发病机制相关的失调的能力。 方法：为了研究早期 HD 中的基因剪接变化，我们进行了替代剪接分析，并结合蛋白质基因组学方法，以鉴定在既定的同源 HD 细胞模型中与早期 CAG 长度相关的剪接变化。 结果：我们报告了广泛的神经元分化阶段和 CAG 长度依赖性剪接变化，并发现与突变 HTT 相关的剪接存在 RNA 处理、神经元功能和表观遗传修饰相关基因的富集。当与蛋白质组数据集集成时，我们在蛋白质水平上鉴定了其中的几个差异剪接事件。通过与人类死后和小鼠模型数据进行比较，我们从胚胎干细胞到死后纹状体组织中鉴定出了几种改变的剪接模式。 解释：我们表明，HD 中的广泛剪接失调发生在神经元发育的早期细胞模型中。重要的是，我们在我们的 HD 细胞模型中观察到与 HD 相关的剪接变化，这些变化也在人类 HD 纹状体和小鼠模型 HD 纹状体中被识别到，这表明剪接相关的发病机制可能在神经元发育的早期发生，并持续到疾病的后期阶段。总之，我们的研究结果突出了 HD 中的剪接失调，这可能导致神经元功能和神经病理学紊乱。 资金：这项研究得到了新加坡南洋理工大学李光前医学院、新加坡教育部在其新加坡教育部学术研究基金 1 级（RG23/22）下的南洋助理教授启动资助、BC 儿童医院研究所研究员资助奖（IGAP）和来自迈克尔史密斯健康研究不列颠哥伦比亚省的学者奖的支持。
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