Fig. 5

Early pre-implantation ethanol exposure induces sex-specific DNA methylation alterations in developing embryonic forebrain. Identification of male and female sex-specific DMRs in EtOH-exposed E10.5 forebrains is described in Fig. 4a. a Manhattan plot showing differences of CpG methylation by chromosomal position for male-specific (n = 971; left graph) and female-specific (n = 133; right graph) DMRs. Red dots represent DMRs with increased-methylation in EtOH-exposed forebrains (n = 31; right graph and n = 30; left graph); blue dots represent DMRs with decreased methylation in EtOH-exposed forebrains (n = 940; right graph and n = 103; left graph); grey dots represent tiles with less than 10% methylation differences between EtOH-exposed and control forebrains (n = 45 504; right graph and n = 46 342; left graph). b Proportion of sex-specific DMRs (upper graph; male n = 971, lower graph; female n = 133) associated with changes in CpG methylation levels between control and EtOH-exposed E10.5 forebrains. c Percentage of sex-specific DMR-associated tiles (male n = 971, female n = 133) across various genomic features: genomic annotation (top), repeat elements (middle) and CpG-rich context (bottom). d Percentage of CpG methylation levels of male- and female-specific DMRs based on the distribution of CpG sites in CpG islands, CpG shores and CpG shelves. e Functional enrichment analysis showing top enriched pathways for male-(top 10 pathways, n = 508 unique gene DMRs) and female-specific DMRs (n = 87 unique gene DMRs), based on Metascape analysis for pathways and p value. The size of the dot represents the number of DMR-associated genes in pathways, and gene ratio represents the number of DMR-associated genes with regards to the number of genes in a pathway. f Examples of CpG methylation levels of individual samples for sex-specific DMR-associated genes related to the top enriched pathways in e). Letters under gene name relate to the pathways in e). ǂ represents significant differences in CpG methylation levels of DMRs (e.g., ±  > 10% methylation difference, q < 0.01) between control and EtOH-exposed embryos (see methods section for details)