Table_2_Putative Causal Variants Are Enriched in Annotated Functional Regions From Six Bovine Tissues.XLSX

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.

影响复杂性状的因果变异（causal variants）被认为广泛存在于基因组的功能区域中。鉴定因果变异有助于预测奶牛（dairy cows）的复杂性状表型，但目前牛基因组（bovine genome）的功能区域注释仍较为匮乏。可借助染色质免疫共沉淀测序（Chromatin immunoprecipitation followed by sequencing, ChIP-seq）技术，在全基因组范围内检测组蛋白修饰（histone modifications）以及与基因组相互作用的蛋白质（如转录因子（transcription factors）），从而实现功能区域的鉴定。本研究采用ChIP-seq技术，对2~3头泌乳奶牛（lactating dairy cows）的6种组织（心脏、肾脏、肝脏、肺脏、乳腺和脾脏）针对4种组蛋白修饰（H3K4Me1、H3K4Me3、H3K27ac及H3K27Me3）与1种转录因子（CTCF）展开检测，以此鉴定牛基因组的功能区域。本研究共生成86个ChIP-seq样本，成功鉴定出牛基因组内数百万个功能区域。研究人员借助ChromHMM软件对组蛋白修饰与CTCF的组合模式进行分析，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显著差异，这提示部分区域可能在组织特异性调控中发挥关键作用。部分ChIP峰的读段计数（read counts）与邻近基因的表达量存在相关性，这印证了功能区域的顺式调控（cis-regulatory）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了推定的因果变异（putative causal variants）。值得注意的是，与基因表达相关的区域尤其富集了潜在因果变异。该结果支持了"复杂性状由改变基因表达的变异所调控"这一假说。本研究构建了当前规模最大的牛ChIP-seq注释资源库之一，且首次涵盖了泌乳奶牛的乳腺组织数据。本研究通过将调控区域与表达数量性状位点（expression QTL）和性状数量性状位点（trait QTL）进行关联，提出了一种鉴定牛因果变异的全新策略。