Gene body DNA hydroxymethylation restricts magnitude of transcriptional changes during aging [RNA-seq]

DNA hydroxymethylation (5hmC) is the most abundant oxidative derivative of DNA methylation (5mC) and is typically enriched at enhancers and gene bodies of transcriptionally active and tissue-specific genes. Although aberrant genomic 5hmC has been implicated in many age-related diseases, the functional role of the modification in aging remains largely unknown. Here, we report that 5hmC is stably enriched in multiple aged organs. Using the liver and cerebellum as model organs, we show that 5hmC accumulates in gene bodies associated with tissue-specific function and thereby restricts the magnitude of gene expression changes during aging. Mechanistically, we found that 5hmC decreases binding affinity of splicing factors compared to unmodified cytosine and 5mC, and is correlated with age-related alternative splicing events, suggesting RNA splicing as a potential mediator of 5hmC's transcriptionally restrictive function. Furthermore, we show that various age-related contexts, such as prolonged quiescence and senescence, are partially responsible for driving the accumulation of 5hmC with age. We provide evidence that this age-related function is conserved in mouse and human tissues, and further show that the modification is altered by regimens known to modulate lifespan. Our findings reveal that 5hmC is a regulator of tissue-specific function and may play a role in regulating longevity. RNA-seq using young (2-4 months) and old (18-20 months) mouse liver (four biological replicates per group).DNA羟甲基化（5hmC）是DNA甲基化（5mC）最丰富的氧化衍生物，通常富集于转录活跃及组织特异性基因的增强子区域与基因体中。尽管异常基因组5hmC已被证实与多种年龄相关性疾病相关，但该修饰在衰老过程中的功能作用仍未被充分阐明。本研究发现，5hmC在多个衰老器官中稳定富集。以肝脏与小脑作为模式器官，我们证实5hmC会在与组织特异性功能相关的基因体中积累，从而限制衰老过程中基因表达变化的幅度。机制层面，我们发现相较于未修饰胞嘧啶与5mC，5hmC会降低剪接因子的结合亲和力，且该修饰与年龄相关可变剪接事件存在关联，这提示RNA剪接可能是5hmC发挥转录抑制功能的潜在介导因子。进一步研究表明，多种年龄相关情境（如长期静息状态与细胞衰老）是驱动5hmC随年龄增长积累的部分原因。我们提供的证据显示，这一年龄相关功能在小鼠与人类组织中均保守存在，且该修饰可被已知可调节寿命的干预方案所改变。本研究结果揭示，5hmC是组织特异性功能的调控因子，或在寿命调控中发挥作用。本研究对年轻（2-4月龄）与老年（18-20月龄）小鼠肝脏开展了RNA测序实验，每组设置4个生物学重复。