PDF(Pubmed)
Abstract：
Nonsense variants underlie many genetic diseases. The phenotypic impact of nonsense variants is determined by Nonsense-mediated mRNA decay (NMD), which degrades transcripts with premature termination codons (PTCs). NMD activity varies across transcripts and cellular contexts via poorly understood mechanisms. Here, by leveraging human genetic datasets, we uncover that the amino acid preceding the PTC dramatically affects NMD activity in human cells. We find that glycine codons in particular support high levels of NMD and are enriched before PTCs but depleted before normal termination codons (NTCs). Gly-PTC enrichment is most pronounced in human genes that tolerate loss-of-function variants. This suggests a strong biological impact for Gly-PTC in ensuring robust elimination of potentially toxic truncated proteins from non-essential genes. Biochemical assays revealed that the peptide release rate during translation termination is highly dependent on the identity of the amino acid preceding the stop codon. This release rate is the most critical feature determining NMD activity across our massively parallel reporter assays. Together, we conclude that NMD activity is significantly modulated by the \"window of opportunity\" offered by translation termination kinetics. Integrating the window of opportunity model with the existing framework of NMD would enable more accurate nonsense variant interpretation in the clinic.
摘要：
无义介导的mRNA衰变（NMD）是一种质量控制机制，可通过降解具有过早终止密码子（PTC）的转录物来防止有害的截短蛋白质的积累。NMD效率在许多情况下都不同，但是影响这种变异性的因素仍然知之甚少。这里,与正常终止密码子（NTC）之前的Gly密码子缺失相比，我们发现常见无义变体中PTC之前的甘氨酸（Gly）密码子富集。与丙氨酸-PTC环境相比，Gly-PTC环境具有更高的NMD活性，这种效应在具有长3个UTR的NMD底物上更强。我们使用大规模平行报告子试验来测试-2和-1密码子的所有可能组合，PTC,和+4个核苷酸，以全面评估PTC序列上下文如何影响NMD效率。随机森林分类器显示，翻译终止过程中的肽基-tRNA水解速率是区分高NMD活性和低NMD活性的最重要特征。我们通过体外生化测定表明，与其他密码子相比，Gly-TC上下文的终止率最慢。此外，Gly-PTC富集在耐受功能丧失变体的基因中最为明显，这表明Gly-PTC背景下增强的NMD塑造了PTC的演变。基于这些发现,我们认为NMD效率受肽基tRNA水解速率提供的“机会窗口”调节，因此，翻译终止动力学。
