Alcohol induced increases in sperm Histone H3 lysine 4 trimethylation correlate with increased placental CTCF occupancy and altered developmental programming

Using a mouse model, studies by our group reveal that paternal preconception alcohol intake affects offspring fetal-placental growth, with long-lasting consequences on adult metabolism. Here, we tested the hypothesis that chronic preconception male alcohol exposure impacts histone enrichment in sperm and that these changes are associated with altered developmental programming in the placenta. Using chromatin immunoprecipitation, we find alcohol-induced increases in sperm histone H3 lysine 4 trimethylation (H3K4me3) that map to promoters and presumptive enhancer regions enriched in genes driving neurogenesis and craniofacial development. Given the colocalization of H3K4me3 with the chromatin binding factor CTCF across both sperm and embryos, we next examined CTCF localization in the placenta. We find global changes in CTCF binding within placentae derived from the male offspring of alcohol-exposed sires. Furthermore, altered CTCF localization correlates with dysregulated gene expression across multiple gene clusters; however, these transcriptional changes only occur in male offspring. Finally, we identified a correlation between genomic regions exhibiting alcohol-induced increases in sperm H3K4me3 and increased CTCF binding in male placentae. Collectively, our analysis demonstrates that the chromatin landscape of sperm is sensitive to chronic alcohol exposure and that a subset of these affected regions exhibits increased placental CTCF enrichment. Overall design: Chromatin immunoprecipitation DNA-sequencing (ChIP-seq) for histone H3K4me3 in alcohol-exposed mouse sperm and chromatin binding factor CTCF in placentae of male offspring at GD14.5

本研究团队通过小鼠模型开展的研究显示，父代孕前饮酒可影响子代的胎儿-胎盘生长，并对成年代谢产生长期影响。本研究旨在验证下述假说：孕前长期雄性酒精暴露会影响精子中的组蛋白富集，且此类改变与胎盘发育编程异常相关。通过染色质免疫共沉淀（chromatin immunoprecipitation），我们发现酒精暴露诱导的精子组蛋白H3赖氨酸4三甲基化（H3K4me3）水平升高，其结合位点富集于驱动神经发生与颅面发育的基因启动子及假定增强子区域。鉴于H3K4me3与染色质结合因子CTCF在精子与胚胎中均存在共定位，我们后续检测了胎盘中CTCF的定位情况。结果显示，暴露于酒精的父代所产雄性子代的胎盘中，CTCF结合出现全局改变。此外，CTCF定位异常与多个基因簇的基因表达失调相关，但此类转录变化仅见于雄性子代。最后，我们发现精子中存在酒精诱导的H3K4me3富集升高的基因组区域，与雄性胎盘中CTCF结合增强的区域存在相关性。综上，本研究表明精子的染色质景观对慢性酒精暴露敏感，且其中部分受影响区域在胎盘中呈现CTCF富集增加。总体实验设计：对酒精暴露小鼠精子中的组蛋白H3K4me3进行染色质免疫共沉淀测序（chromatin immunoprecipitation DNA-sequencing，ChIP-seq），并对妊娠第14.5天（GD14.5）雄性子代胎盘中的染色质结合因子CTCF进行染色质免疫共沉淀测序