Characterization of a COPD-Associated NPNT Functional Splicing Genetic Variant in Human Lung Tissue via Long-Read Sequencing.
Characterization of a COPD-Associated NPNT Functional Splicing Genetic Variant in Human Lung Tissue via Long-Read Sequencing.
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通过长读长测序表征人肺组织中与 COPD 相关的 NPNT 功能剪接遗传变异。
DOI：
10.1101/2020.10.20.20203927
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发表时间：
2020
期刊：
影响因子：
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通讯作者：
Castaldi,PeterJ
中科院分区：
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作者：
Saferali,Aabida;Xu,Zhonghui;Sheynkman,GloriaM;Hersh,CraigP;Cho,MichaelH;Silverman,EdwinK;Laederach,Alain;Vollmers,Christopher;Castaldi,PeterJ
慢性阻塞性肺疾病（COPD）是世界范围内的主要死亡原因。全基因组关联研究（GWAS）已经确定了80多个与COPD和肺气肿相关的基因座，然而对于大多数这些基因座的致病变异和基因是未知的。在这里，我们利用来自基因型组织表达项目（GTEx）的肺剪接定量性状位点（sQTL）数据和来自肺组织研究联盟（LTRC）的短读测序数据来表征nephronectin （NPNT）中与COPD病例控制状态和肺功能相关的位点。我们发现rs34712979变异与inpnt的选择性剪接连接有关，特别是连接第2和第4外显子的连接（chr4:105898001-105927336） （p=4.02×10−38）。这种关联与COPD的GWAS数据和肺肺活量测量的后验概率为94%，表明相同的因果遗传变异inpntt与COPD风险、肺功能肺活量测量和剪接相关。对npntshort - read测序的研究表明，rs34712979产生了一个隐剪接受体位点，导致包含一个3个核苷酸的外显子扩展，编码蛋白质n端附近的丝氨酸残基。利用Oxford Nanopore Technologies （ONT）的长读测序技术，我们鉴定出了13NPNTisoforms，其中6个被预测为蛋白质编码。其中两种是全长同种异构体，仅在3个核苷酸外显子扩展上不同，其发生情况因基因型而异。总的来说，我们的数据表明rs34712979通过创建一个新的剪接受体来调节COPD风险和肺功能，该剪接受体导致在nephronectin蛋白序列中包含一个编码丝氨酸的3个核苷酸序列。我们的研究结果表明，enpnt剪接与COPD风险有关，并在nephronectin蛋白中发现了一种新的丝氨酸插入，值得进一步研究。
Chronic obstructive pulmonary disease (COPD) is a leading cause of death worldwide. Genome-wide association studies (GWAS) have identified over 80 loci that are associated with COPD and emphysema, however for most of these loci the causal variant and gene are unknown. Here, we utilize lung splice quantitative trait loci (sQTL) data from the Genotype-Tissue Expression project (GTEx) and short read sequencing data from the Lung Tissue Research Consortium (LTRC) to characterize a locus in nephronectin (NPNT) associated with COPD case-control status and lung function. We found that the rs34712979 variant is associated with alternative splice junction use inNPNT, specifically for the junction connecting the 2nd and 4th exons (chr4:105898001-105927336) (p=4.02×10−38). This association colocalized with GWAS data for COPD and lung spirometry measures with a posterior probability of 94%, indicating that the same causal genetic variants inNPNTunderlie the associations with COPD risk, spirometric measures of lung function, and splicing. Investigation ofNPNTshort read sequencing revealed that rs34712979 creates a cryptic splice acceptor site which results in the inclusion of a 3 nucleotide exon extension, coding for a serine residue near the N-terminus of the protein. Using Oxford Nanopore Technologies (ONT) long read sequencing we identified 13NPNTisoforms, 6 of which are predicted to be protein coding. Two of these are full length isoforms which differ only in the 3 nucleotide exon extension whose occurrence differs by genotype. Overall, our data indicate that rs34712979 modulates COPD risk and lung function by creating a novel splice acceptor which results in the inclusion of a 3 nucelotide sequence coding for a serine in the nephronectin protein sequence. Our findings implicateNPNTsplicing in contributing to COPD risk, and identify a novel serine insertion in the nephronectin protein that warrants further study.