Department of Molecular and Cell Biology, University of California Berkeley, Berkeley, California, United States of America.
PLoS Genet. 2012;8(8):e1002882. doi: 10.1371/journal.pgen.1002882. Epub 2012 Aug 16.
Gene expression varies widely between individuals of a population, and regulatory change can underlie phenotypes of evolutionary and biomedical relevance. A key question in the field is how DNA sequence variants impact gene expression, with most mechanistic studies to date focused on the effects of genetic change on regulatory regions upstream of protein-coding sequence. By contrast, the role of RNA 3'-end processing in regulatory variation remains largely unknown, owing in part to the challenge of identifying functional elements in 3' untranslated regions. In this work, we conducted a genomic survey of transcript ends in lymphoblastoid cells from genetically distinct human individuals. Our analysis mapped the cis-regulatory architecture of 3' gene ends, finding that transcript end positions did not fall randomly in untranslated regions, but rather preferentially flanked the locations of 3' regulatory elements, including miRNA sites. The usage of these transcript length forms and motifs varied across human individuals, and polymorphisms in polyadenylation signals and other 3' motifs were significant predictors of expression levels of the genes in which they lay. Independent single-gene experiments confirmed the effects of polyadenylation variants on steady-state expression of their respective genes, and validated the regulatory function of 3' cis-regulatory sequence elements that mediated expression of these distinct RNA length forms. Focusing on the immune regulator IRF5, we established the effect of natural variation in RNA 3'-end processing on regulatory response to antigen stimulation. Our results underscore the importance of two mechanisms at play in the genetics of 3'-end variation: the usage of distinct 3'-end processing signals and the effects of 3' sequence elements that determine transcript fate. Our findings suggest that the strategy of integrating observed 3'-end positions with inferred 3' regulatory motifs will prove to be a critical tool in continued efforts to interpret human genome variation.
基因表达在人群个体之间存在广泛差异，调控变化可能是具有进化和医学相关性的表型的基础。该领域的一个关键问题是 DNA 序列变异如何影响基因表达，迄今为止大多数机制研究都集中在遗传变化对蛋白质编码序列上游调控区域的影响上。相比之下，RNA 3' 端加工在调控变异中的作用在很大程度上仍然未知，部分原因是识别 3' 非翻译区中功能元件的挑战。在这项工作中，我们对来自遗传上不同的人类个体的淋巴母细胞中的转录本末端进行了基因组调查。我们的分析绘制了 3' 基因末端的顺式调控结构图谱，发现转录本末端位置并非随机出现在非翻译区，而是优先侧翼 3' 调控元件的位置，包括 miRNA 位点。这些转录本长度形式和基序的使用在人类个体之间存在差异，多聚腺苷酸化信号和其他 3' 基序中的多态性是它们所在基因表达水平的重要预测因子。独立的单基因实验证实了多聚腺苷酸化变体对其各自基因的稳态表达的影响，并验证了介导这些不同 RNA 长度形式表达的 3' 顺式调控序列元件的调节功能。我们以免疫调节因子 IRF5 为重点，确定了 RNA 3' 端加工的自然变异对抗原刺激的调控反应的影响。我们的研究结果强调了在 3' 端变异遗传学中发挥作用的两种机制的重要性：使用不同的 3' 端加工信号和决定转录本命运的 3' 序列元件的作用。我们的发现表明，将观察到的 3' 端位置与推断的 3' 调控基序相结合的策略将被证明是解释人类基因组变异的持续努力的重要工具。
