TDP43 proteinopathy exhibits disease, tissue, and context-specific cryptic splicing signatures. TDP43 proteinopathy exhibits disease, tissue, and context-specific cryptic splicing signatures

Mislocalization of the nuclear protein TDP43 is a hallmark of ALS and FTD and leads to de-repression and inclusion of cryptic exons, which represent promising biomarkers of TDP43 pathology. However, most cryptic exons to date have been identified from in vitro models, limiting our understanding of any tissue and/or cell-specific splices. We meta-analyzed published bulk RNA-Seq datasets representing 1,778 RNAseq profiles of ALS and FTD post mortem tissue, and in vitro models with experimentally depleted TDP43. We identified novel cryptic splices and mapped out their tissue-specificity, demonstrating subsets with distinct cortical and spinal cord enrichment. Novel events were validated by RNA-Seq and RT-qPCR in a new spinal cord cohort, and analysis of single-nucleus datasets localized cortical splices to layer-specific neuronal populations. This catalog of cryptic splices is the first step towards the development of biomarkers for cell type-specific TDP43 pathology. Overall design: Post mortem lumbar spinal cord samples were obtained (n=10 healthy controls, n=10 ALS), and bulk RNA was isolated. We performed differential splicing anlayses with SGSeq/DEXseq to characterize TDP43-associated cryptic splicing. **Raw data are not provided due to privacy concerns**

核蛋白TDP43的定位异常是肌萎缩侧索硬化症（ALS）和额颞叶痴呆（FTD）的标志性病理特征，可导致隐蔽外显子的去抑制与异常包含，这类隐蔽外显子是TDP43病理的潜在生物标志物。然而迄今为止，绝大多数隐蔽外显子均通过体外模型得以鉴定，这限制了我们对组织及/或细胞类型特异性可变剪接的认知。我们对已发表的批量RNA测序（bulk RNA-Seq）数据集进行了荟萃分析，这些数据集涵盖了1778份ALS和FTD死后组织的RNA测序图谱，以及经实验性敲低TDP43的体外模型样本。我们鉴定出了新型隐蔽剪接事件，并明确了其组织特异性，揭示出具有不同皮层与脊髓富集特征的亚型。通过对新增脊髓队列样本的RNA测序与实时定量聚合酶链反应（RT-qPCR）验证，我们确认了这些新型事件；而对单细胞核测序数据集的分析则将皮层剪接事件定位于特定层状的神经元群体中。本隐蔽剪接事件目录是开发针对细胞类型特异性TDP43病理的生物标志物的第一步。
整体实验设计：我们获取了死后腰椎脊髓样本（健康对照10例，ALS患者10例），并分离得到批量RNA。我们采用SGSeq/DEXseq工具开展差异剪接分析，以表征与TDP43相关的隐蔽剪接事件。**原始数据因隐私问题暂不提供**