Identifying regulatory uORFs as a targetable axis for hereditary disease

摘要(来自家长赠款) 这项建议的主要目标是通过计算识别、验证，然后针对治疗 在蛋白质编码基因的5‘非翻译区(UTR)内的干预调控元件， 称为上游开放阅读框(UORF)。在这样做的过程中，我们的目标是调节蛋白质 从选定的基因输出，提供了一种具有广泛潜力的新颖的翻译方法。 UORF是5‘UTRmRNA序列的片段，可以启动和终止翻译 蛋白质编码上游(CDS)起始密码子。罕见的uORF已被证明能产生 可能具有功能的微肽，而其他的则通过调节翻译来影响蛋白质的表达 下游蛋白质编码序列的速率。我们最近的研究表明，uORF展示了 保留其起始和结束密码子的强负选择，但不保留编码的密码子 序列，支持uORF通过进化保留的概念，因为它们 蛋白质基因产物的顺式调控作用。利用基因数据库，我们发现了变异 影响与疾病表型相关的uORF起始/终止密码子(Phewas分析)和 验证了他们对下游基因蛋白质(但不是RNA)水平的预期影响。 在大约50%的人类蛋白质编码基因中发现了潜在的uORF，其中许多基因 窝藏多个uORF。虽然阻断一些uORF会减少CDS蛋白的产量，但我们的 未发表的数据表明，靶向uORF可以用来增加CDS的翻译。作为一名 蛋白质水平升高的潜在案例，我们关注的是可遗传的肺动脉 高血压(PAH)，一种目前无法治愈的致命疾病，最常见的原因是BMPR2 单倍体功能不全。通过阻断BMPR2 uORF，我们能够提高BMPR2蛋白水平 高达220%。基于我们已发表和未发表的结果，我们提出了一套综合的 计算和实验方法，以系统地检测、区分优先顺序、验证和 靶向uORF用于治疗干预，重点关注与单倍体功能不全相关的基因 和ASOS作为一种干预。

Abstract (from parent grant) The broad goal of this proposal it to computationally identify, validate, then target for therapeutic intervention regulatory elements within the 5' untranslated regions (UTR) of protein coding genes, known as upstream open reading frames (uORFs). In so doing, we aim to modulate the protein output from selected genes, offering a novel, translational approach with broad potential. uORFs are segments of 5′UTR mRNA sequences that can initiate and terminate translation upstream of protein-coding (CDS) start codons. Rare uORFs have been shown to produce potentially functional micro-peptides, while others affect protein expression by tuning translation rates of downstream protein-coding sequences. Our recent work showed that uORFs exhibit strong negative selection for preserving their start and stop codons, but not the encoded sequence, favoring the notion that uORFs have been retained through evolution because of their cis regulatory effect on protein gene products. Using genetic databases, we showed that variants affecting uORF start/stop codons associate with disease phenotypes (PheWAS analysis) and validated their expected effect on the protein (but not RNA) levels of the downstream genes. Potential uORFs have been identified in ~50% of all human protein-coding genes, many of which harbor more than one uORF. While blocking some uORFs decreases CDS protein output, our unpublished data show that targeting uORFs can be used to increase CDS translation. As a potential case for increasing protein levels, we focused on heritable pulmonary arterial hypertension (PAH), a fatal condition with no current cure, most commonly driven by BMPR2 haploinsufficiency. By blocking a BMPR2 uORF, we were able to increase BMPR2 protein levels by up to 220%. Based on our published and unpublished results we propose an integrative set of computational and experimental approaches to systematically detect, prioritize, validate and target uORFs for therapeutic intervention, focusing on genes associated with haploinsufficiency and ASOs as an intervention.