Epigenetic consequences of hormonal interactions between opposite‐sex twin fetuses

Previous studies reported inconsistent evidence about some phenotypic traits of females in human opposite‐sex twins (opposite‐sex females [OSF]) being distinct from females in same‐sex twins (SSF). Comparatively, less evidence showed significant differences between males in OS twins (opposite‐sex males [OSM]) and males in same‐sex twins (SSM). The twin testosterone transfer hypothesis suggests that prenatal exposure of testosterone in utero may be a possible explanation for the differential traits in OSF; however, the underlying mechanism is unknown. Here, we investigated the potential epigenetic effects of hormone interactions and their correlation to the observed phenotypic traits. In the study, DNA methylomic data from 54 newborn twins and histone modification data (H3K4me3, H3K4me1, H3K27me3, and H3K27ac) from 14 newborn twins, including same‐sex females (SSF), OS twins, and same‐sex males (SSM) were generated. We found that OSF were clearly distinguishable from SSF by DNA methylome, while OSM were distinguishable from SSM by H3K4me1 and H3K4me3. To be more specific, compared to SSF, OSF showed a stronger correlation to males (OSM and SSM) in genome‐wide DNA methylation. Further, the DNA methylomic differences between OSF and SSF were linked to the process involving cognitive functions and nervous system regulation. The differential H3K4me3 between OSM and SSM was linked to immune responses. These findings provide epigenetic evidence for the twin testosterone transfer hypothesis and offer novel insights on how prenatal hormone exposure in utero may be linked to the reported differential traits of OS twins.