Twin birth changes DNA methylation of subsequent siblings

We asked if twin birth influences the DNA methylation of subsequent siblings. We measured whole blood methylation using the HumanMethylation450 array for siblings from two twin and family studies in Australia and Korea. We compared the means and correlations in methylation between pairs of siblings born before a twin birth (BT siblings), born on either side of a twin birth (B/AT pairs) and born after a twin birth (AT siblings). For the genome-wide average DNA methylation, the correlation for AT pairs (r_AT) was larger than the correlation for BT pairs (r_BT) in both studies, and from the meta-analysis, r_AT = 0.46 (95% CI: 0.26, 0.63) and r_BT = -0.003 (95% CI: -0.30, 0.29) (P = 0.02). B/AT pairs were not correlated (from the meta-analysis r_BAT = 0.08; 95% CI: -0.31, 0.45). Similar results were found for the average methylation of several genomic regions, e.g., CpG shelf and gene body. BT and AT pairs were differentially correlated in methylation for 15 probes (all P < 10^-7), and the top 152 differentially correlated probes (at P < 10^-4) were enriched in cell signalling and breast cancer regulation pathways. Our observations are consistent with a twin birth changing the intrauterine environment such that siblings both born after a twin birth are correlated in DNA methylation.