Computational and Experimental Modeling of Alternative Polyadenylation

项目摘要/摘要 交替多聚腺苷(APA)在大多数人的转录后调节中起着重要的作用 基因。APA的广泛重要性从一个关键的APA-NUDT21的表达变化中得到了很好的例证 调节器，我们在这项拨款的第一个周期中报告了，用于治疗胶质母细胞瘤、特发性肺疾病 纤维化症和神经精神障碍。最近，我们的工作揭示了一种新的机制，通过这种机制，3ʹ- 非编码区缩短可通过干扰竞争来抑制反式抑制基因(如PTEN) 内源RNA(Cerna)串扰，而不是通过诱导顺式基因的癌基因。除了这几个人 例如，APA在广泛的人类特征和疾病中的患病率和功能仍然存在 很大程度上是未知的。大多数人类特征/疾病被发现与数十万 大量全基因组关联研究中的非编码单核苷酸多态(SNPs)。 然而，这些SNPs的功能解释仍然是一个巨大的挑战，因为GWAS的数据不 展示SNP是如何工作的。为了更好地了解它们的影响，表达数量性状基因座(EQTL)有 已被广泛用于将GWASSNPs与基因表达联系起来。尽管在阐明eQTL方面做出了巨大的努力，但 许多GWASNPs的功能仍未得到解释。一个重要的原因是eQTL不考虑APA 监管。我们最近构建了第一个人类3‘端非编码区APA数量性状基因座(3’aQTL)，它 使用约8,000个GTEx v7 RNA-SEQ样本，包含约40万个与目标基因的APA相关的SNP 涉及46种组织类型(自然遗传学，2021年接受)。这些3‘aQTL可解释约16.1%的GWA值 SNP存在于15个常见性状/疾病中，它们与eQTL和剪接QTL有很大的区别。基于 这些令人兴奋的初步数据，我们假设APA的计算和实验建模将 极大地促进了对许多对APA调控重要的GWASSNPs的解释，这些SNPs是 富含3‘端非编码区和基因下游区域。因此，我们建议发展创新的生物信息学。 以及鉴定3‘aQTL和提名APA连锁疾病/性状易感基因的实验方法 在各种各样的细胞类型和环境因素下，随后在体内使用功能表征 我们独特的CRISPR工程系统。我们希望将APA确立为一种新兴的重要分子 表型解释了很大一部分GWAS风险SNPs，导致了对 APA和与APA相关的易感基因在广泛的人类特征和疾病中的遗传基础。

Project Summary / Abstract Alternative polyadenylation (APA) plays an important role in the post-transcriptional regulation of most human genes. The broad importance of APA is well exemplified by the altered expression of NUDT21, a key APA regulator that we reported in the first cycle of this grant, in diseases such as glioblastoma, diopathic pulmonary fibrosis, and neuropsychiatric disorders. More recently, our work has revealed a novel mechanism by which 3ʹ- UTR shortening can repress tumor suppressor genes (e.g., PTEN) in trans by disrupting competing endogenous RNA (ceRNA) crosstalk, rather than by inducing oncogenes in cis. Aside from these few examples, the prevalence and functions of APA in a wide spectrum of human traits and diseases remain largely unknown. Most human traits/diseases have been found as associated with hundreds of thousands of noncoding single-nucleotide polymorphisms (SNPs) in numerous genome-wide association studies (GWASs). However, functional interpretation of these SNPs remains a significant challenge because GWAS data do not show how the SNPs work. To better understand their effects, expression quantitative trait loci (eQTLs) have been widely used to link GWAS SNPs to gene expression. Despite massive efforts on elucidating eQTLs, the functions of many GWAS SNPs remain unexplained. An important reason is that eQTLs do not consider APA regulation. We have recently constructed the first human 3′UTR APA quantitative trait loci (3′aQTLs), which contain ~0.4 million SNPs associated with APA of target genes, using ~8,000 GTEx v7 RNA-seq samples across 46 tissue types (Nature Genetics, accepted in 2021). These 3′aQTLs can explain ~16.1% of GWAS SNPs in 15 common traits/diseases, and they are largely distinct from eQTLs and splicing QTLs. Based on these exciting preliminary data, we hypothesize that computational and experimental modeling of APA will substantially facilitate the interpretation of numerous GWAS SNPs important for APA regulation, which are enriched in 3′UTRs and gene downstream regions. Hence, we propose to develop innovative bioinformatics and experimental methods for identifying 3′aQTLs and nominating APA-linked disease/trait susceptibility genes in a wide variety of cell types and environmental factors, followed by in vivo functional characterization using our unique CRISPR engineering system. We expect to establish APA as an emerging and important molecular phenotype to explain a large fraction of GWAS risk SNPs, leading to significant novel biological insights into the genetic basis of APA and APA-linked susceptibility genes in a wide spectrum of human traits and diseases.