Variant Histone H2afv reprograms DNA methylation during early zebrafish development

The DNA methylome is re-patterned during discrete phases of vertebrate development. In zebrafish, there are 2 waves of global DNA demethylation and re-methylation: the first occurs before gastrulation when the parental methylome is changed to the zygotic pattern and the second occurs after formation of the embryonic body axis, during organ specification. The occupancy of the histone variant H2A.Z and regions of DNA methylation are generally anti-correlated, and it has been proposed that H2A.Z restricts the boundaries of highly methylated regions. While many studies have described the dynamics of methylome changes during early zebrafish development, the factors involved in establishing the DNA methylation landscape in zebrafish embryos have not been identified. We test the hypothesis that the zebrafish ortholog of H2A.Z (H2afv) restricts DNA methylation during development. We find that, in control embryos, bulk genome methylation decreases after gastrulation, with a nadir at the bud stage, and peaks during mid-somitogenesis; by 24 hours post -fertilization, total DNA methylation levels return to those detected in gastrula. Early zebrafish embryos depleted of H2afv have significantly more bulk DNA methylation during somitogenesis, suggesting that H2afv limits methylation during this stage of development. H2afv deficient embryos are small, with multisystemic abnormalities. Genetic interaction experiments demonstrate that these phenotypes are suppressed by depletion of DNA methyltransferase 1 (Dnmt1). This work demonstrates that H2afv is essential for global DNA methylation reprogramming during early vertebrate development and that embryonic development requires crosstalk between H2afv and Dnmt1.

脊椎动物发育的不同阶段中，DNA甲基化组（DNA methylome）会经历重编程重塑。在斑马鱼中，存在两轮全局性DNA去甲基化与重新甲基化的波动：第一轮发生在原肠胚形成前，此时亲本甲基化组转变为合子型甲基化模式；第二轮则在胚胎体轴形成后、器官特化阶段发生。组蛋白变体H2A.Z（histone variant H2A.Z）的染色质占据区域与DNA甲基化区域通常呈负相关，且有研究提出H2A.Z可限制高度甲基化区域的边界。尽管已有多项研究阐明了斑马鱼早期发育过程中甲基化组的动态变化，但斑马鱼胚胎中建立DNA甲基化图谱的相关调控因子仍未被探明。本研究验证了如下假说：斑马鱼H2A.Z的同源基因H2afv可在发育进程中限制DNA甲基化水平。研究结果显示，在对照胚胎中，全基因组甲基化水平在原肠胚形成后出现下降，在胚体期达到最低点，随后在体节发生中期升至峰值；至受精后24小时，总DNA甲基化水平恢复至原肠胚阶段的检测水平。在体节发生阶段，H2afv沉默的早期斑马鱼胚胎的全基因组甲基化水平显著升高，这表明H2afv可在该发育阶段限制甲基化进程。H2afv缺陷型胚胎体型偏小，并伴随多系统异常表型。遗传互作实验表明，通过沉默DNA甲基转移酶1（DNA methyltransferase 1，Dnmt1）可抑制上述表型。本研究证实，H2afv对于脊椎动物早期发育过程中的全局性DNA甲基化重编程至关重要，且胚胎发育依赖于H2afv与Dnmt1之间的交叉互作。