CG Methylation Covaries with Differential Gene Expression between Leaf and Floral Bud Tissues of Brachypodium distachyon

DNA methylation has the potential to influence plant growth and development through its influence on gene expression. To date, however, the evidence from plant systems is mixed as to whether patterns of DNA methylation vary significantly among tissues and, if so, whether these differences affect tissue-specific gene expression. To address these questions, we analyzed both bisulfite sequence (BSseq) and transcriptomic sequence data from three biological replicates of two tissues (leaf and floral bud) from the model grass species Brachypodium distachyon. Our first goal was to determine whether tissues were more differentiated in DNA methylation than explained by variation among biological replicates. Tissues were more differentiated than biological replicates, but the analysis of replicated data revealed high (>50%) false positive rates for the inference of differentially methylated sites (DMSs) and differentially methylated regions (DMRs). Comparing methylation to gene expression, we found that differential CG methylation consistently covaried negatively with gene expression, regardless as to whether methylation was within genes, within their promoters or even within their closest transposable element. The relationship between gene expression and either CHG or CHH methylation was less consistent. In total, CG methylation in promoters explained 9% of the variation in tissue-specific expression across genes, suggesting that CG methylation is a minor but appreciable factor in tissue differentiation.DNA甲基化（DNA methylation）可通过调控基因表达，进而影响植物的生长与发育进程。然而迄今为止，来自植物系统的相关研究证据仍存在分歧：DNA甲基化模式是否会在不同组织间出现显著差异，且若存在此类差异，这些差异是否会影响组织特异性基因的表达。为解答上述疑问，我们对模式禾本科植物二穗短柄草（Brachypodium distachyon）的两种组织（叶片与花芽）的三份生物学重复样本，分析了亚硫酸氢盐测序（bisulfite sequence, BSseq）与转录组测序两类数据。我们的首要目标是明确：相较于生物学重复间的变异，不同组织在DNA甲基化层面的分化程度是否更高。结果显示，组织间的分化程度确实高于生物学重复间的变异，但针对重复样本的分析表明，差异甲基化位点（differentially methylated sites, DMSs）与差异甲基化区域（differentially methylated regions, DMRs）的推断存在超过50%的假阳性率。将甲基化水平与基因表达水平进行关联分析后，我们发现，无论CG甲基化发生在基因内部、基因启动子区域，还是紧邻基因的转座子（transposable element）区域，其差异均与基因表达呈稳定的负相关关系；而CHG或CHH甲基化与基因表达的关联则相对缺乏一致性。总体而言，启动子区域的CG甲基化可解释基因间组织特异性表达差异的9%，这表明CG甲基化在组织分化过程中虽仅发挥次要作用，但其贡献仍不容忽视。