Table_4_Putative Causal Variants Are Enriched in Annotated Functional Regions From Six Bovine Tissues.DOCX

Genetic variants which affect complex traits (causal variants) are thought to be found in functional regions of the genome. Identifying causal variants would be useful for predicting complex trait phenotypes in dairy cows, however, functional regions are poorly annotated in the bovine genome. Functional regions can be identified on a genome-wide scale by assaying for post-translational modifications to histone proteins (histone modifications) and proteins interacting with the genome (e.g., transcription factors) using a method called Chromatin immunoprecipitation followed by sequencing (ChIP-seq). In this study ChIP-seq was performed to find functional regions in the bovine genome by assaying for four histone modifications (H3K4Me1, H3K4Me3, H3K27ac, and H3K27Me3) and one transcription factor (CTCF) in 6 tissues (heart, kidney, liver, lung, mammary and spleen) from 2 to 3 lactating dairy cows. Eighty-six ChIP-seq samples were generated in this study, identifying millions of functional regions in the bovine genome. Combinations of histone modifications and CTCF were found using ChromHMM and annotated by comparing with active and inactive genes across the genome. Functional marks differed between tissues highlighting areas which might be particularly important to tissue-specific regulation. Supporting the cis-regulatory role of functional regions, the read counts in some ChIP peaks correlated with nearby gene expression. The functional regions identified in this study were enriched for putative causal variants as seen in other species. Interestingly, regions which correlated with gene expression were particularly enriched for potential causal variants. This supports the hypothesis that complex traits are regulated by variants that alter gene expression. This study provides one of the largest ChIP-seq annotation resources in cattle including, for the first time, in the mammary gland of lactating cows. By linking regulatory regions to expression QTL and trait QTL we demonstrate a new strategy for identifying causal variants in cattle.

影响复杂性状的遗传变异（因果变异（causal variants））通常被认为富集于基因组的功能区域内。鉴定因果变异有助于预测奶牛的复杂性状表型，但当前牛基因组的功能区域注释仍较为匮乏。可通过染色质免疫共沉淀测序（Chromatin immunoprecipitation followed by sequencing，简称ChIP-seq）技术，检测组蛋白翻译后修饰（组蛋白修饰（histone modifications））以及与基因组相互作用的蛋白质（例如转录因子（transcription factors）），从而在全基因组范围内鉴定功能区域。本研究采用ChIP-seq技术，对2~3头泌乳奶牛的6种组织（心脏、肾脏、肝脏、肺脏、乳腺及脾脏）中的4种组蛋白修饰（H3K4Me1、H3K4Me3、H3K27ac及H3K27Me3）以及1种转录因子（CTCF）进行检测，以鉴定牛基因组中的功能区域。本研究共构建86个ChIP-seq样本，成功鉴定出牛基因组内数百万个功能区域。研究人员通过ChromHMM工具分析组蛋白修饰与CTCF的组合模式，并通过与全基因组范围内的活跃基因及非活跃基因进行比对，完成了功能区域的注释。不同组织间的功能标记存在差异，这凸显了可能在组织特异性调控中发挥关键作用的区域。部分ChIP峰的测序读段计数与邻近基因的表达水平存在相关性，这佐证了功能区域的顺式调控作用。与其他物种的研究结果一致，本研究鉴定出的功能区域显著富集推定的因果变异。值得注意的是，与基因表达相关的区域尤其富集潜在因果变异。这一结果支持了"复杂性状由改变基因表达的变异所调控"的假说。本研究构建了当前规模最大的牛ChIP-seq注释资源库之一，且首次覆盖了泌乳奶牛的乳腺组织。通过将调控区域与表达数量性状位点（expression QTL）及性状数量性状位点（trait QTL）进行关联，本研究提出了一种鉴定牛因果变异的全新策略。