PDF(Pubmed)
Abstract：
Human genetic variants that introduce an AG into the intronic region between the branchpoint (BP) and the canonical splice acceptor site (ACC) of protein-coding genes can disrupt pre-mRNA splicing. Using our genome-wide BP database, we delineated the BP-ACC segments of all human introns and found extreme depletion of AG/YAG in the [BP+8, ACC-4] high-risk region. We developed AGAIN as a genome-wide computational approach to systematically and precisely pinpoint intronic AG-gain variants within the BP-ACC regions. AGAIN identified 350 AG-gain variants from the Human Gene Mutation Database, all of which alter splicing and cause disease. Among them, 74% created new acceptor sites, whereas 31% resulted in complete exon skipping. AGAIN also predicts the protein-level products resulting from these two consequences. We performed AGAIN on our exome/genomes database of patients with severe infectious diseases but without known genetic etiology and identified a private homozygous intronic AG-gain variant in the antimycobacterial gene SPPL2A in a patient with mycobacterial disease. AGAIN also predicts a retention of six intronic nucleotides that encode an in-frame stop codon, turning AG-gain into stop-gain. This allele was then confirmed experimentally to lead to loss of function by disrupting splicing. We further showed that AG-gain variants inside the high-risk region led to misspliced products, while those outside the region did not, by two case studies in genes STAT1 and IRF7. We finally evaluated AGAIN on our 14 paired exome-RNAseq samples and found that 82% of AG-gain variants in high-risk regions showed evidence of missplicing. AGAIN is publicly available from https://hgidsoft.rockefeller.edu/AGAIN and https://github.com/casanova-lab/AGAIN.
摘要：
将AG引入蛋白质编码基因的分支点（BP）和典型剪接受体位点（ACC）之间的内含子区域的人类遗传变异体可以破坏前mRNA剪接。利用我们的全基因组BP数据库，我们描绘了所有人类内含子的BP-ACC片段,发现[BP+8,ACC-4]高危区域的AG/YAG极度耗竭.我们开发了AGAIN作为一种全基因组计算方法，以系统地，精确地查明BP-ACC区域内的内含子AG增益变体。AGAIN从人类基因突变数据库中鉴定出350个AG-gain变体，所有这些都会改变剪接并导致疾病。其中,74%的人创建了新的受体站点，而31%导致完全外显子跳跃。AGAIN还预测了这两种后果导致的蛋白质水平产物。我们对严重感染性疾病患者的外显子组/基因组数据库进行了AGAIN，但没有已知的遗传病因，并在分枝杆菌病患者的抗分枝杆菌基因SPPL2A中鉴定了一个私有纯合内含子AG-gain变异。AGAIN还预测保留六个编码框内终止密码子的内含子核苷酸，将AG增益变成停止增益。然后通过实验证实该等位基因通过破坏剪接导致功能丧失。我们进一步表明，高风险区域内的AG-gain变体导致错误剪接的产品，而该地区以外的人没有，通过STAT1和IRF7基因的两个案例研究。我们最终在我们的14个配对外显子组-RNAseq样品上评估了AGAIN,并且发现高风险区域中82%的AG-gain变体显示了错误剪接的证据。AGAIN可从https://hgidsoft公开获得。洛克菲勒.edu/AGAIN和https://github.com/casanova-lab/AGAIN。
