Table_3_Genome-Wide DNA Methylation and Hydroxymethylation Changes Revealed Epigenetic Regulation of Neuromodulation and Myelination in Yak Hypothalamus.XLSX

Both 5-methylcytosine (5mC) and 5-hydroxymethylcytosine (5hmC) are important epigenetic modifications in neurodevelopment. However, there is little research examining the genome-wide patterns of 5mC and 5hmC in brain regions of animals under natural high-altitude conditions. We used oxidative reduced representation bisulfite sequencing (oxRRBS) to determine the 5mC and 5hmC sites in the brain, brainstem, cerebellum, and hypothalamus of yak and cattle. We reported the first map of genome-wide DNA methylation and hydroxymethylation in the brain, brainstem, cerebellum, and hypothalamus of yak (living at high altitudes) and cattle. Overall, we found striking differences in 5mC and 5hmC between the hypothalamus and other brain regions in both yak and cattle. Genome-wide profiling revealed that 5mC level decreased and 5hmC level increased in the hypothalamus than in other regions. Furthermore, we identified differentially methylated regions (DMRs) and differentially hydroxymethylated regions (DhMRs), most of which overlapped with each other. Interestingly, transcriptome results for these brain regions also showed distinctive gene levels in the hypothalamus. Finally, differentially expressed genes (DEGs) regulated by DMRs and DhMRs may play important roles in neuromodulation and myelination. Overall, our results suggested that mediation of 5mC and 5hmC on epigenetic regulation may broadly impact the development of hypothalamus and its biological functions.

5-甲基胞嘧啶（5-methylcytosine, 5mC）与5-羟甲基胞嘧啶（5-hydroxymethylcytosine, 5hmC）均为神经发育过程中关键的表观遗传修饰。然而，目前针对自然高海拔环境下动物脑区内5mC与5hmC的全基因组分布特征的相关研究仍较为匮乏。本研究采用氧化简化代表性亚硫酸氢盐测序（oxidative reduced representation bisulfite sequencing, oxRRBS）技术，对牦牛与黄牛的大脑、脑干、小脑及下丘脑组织中的5mC与5hmC位点进行了系统检测。本研究首次绘制了栖息于高海拔环境的牦牛与黄牛的大脑、脑干、小脑及下丘脑的全基因组DNA甲基化与羟甲基化图谱。整体分析结果显示，在牦牛与黄牛中，下丘脑与其余脑区的5mC与5hmC水平均存在显著差异：全基因组谱型分析表明，与其他脑区相比，下丘脑中的5mC水平降低，而5hmC水平升高。此外，本研究鉴定出了差异甲基化区域（differentially methylated regions, DMRs）与差异羟甲基化区域（differentially hydroxymethylated regions, DhMRs），其中绝大多数区域存在重叠。值得注意的是，上述脑区的转录组分析结果同样显示，下丘脑的基因表达水平具有显著特异性。最终，由DMRs与DhMRs调控的差异表达基因（differentially expressed genes, DEGs）可能在神经调节与髓鞘形成过程中发挥重要作用。综上，本研究结果提示，5mC与5hmC介导的表观遗传调控可能广泛影响下丘脑的发育及其生物学功能。