Smoking and blood DNA methylation: an epigenome-wide association study and assessment of reversibility

ABSTRACT

We conducted a genome-wide association study of blood DNA methylation and smoking, attempted replication of previously discovered associations, and assessed the reversibility of smoking-associated methylation changes. DNA methylation was measured in baseline peripheral blood samples for 5,044 participants in the Melbourne Collaborative Cohort Study. For 1,032 participants, these measures were repeated using blood samples collected at follow-up, a median of 11 years later. A cross-sectional analysis of the association between smoking and DNA methylation and a longitudinal analysis of changes in smoking status and changes in DNA methylation were conducted. We used our cross-sectional analysis to replicate previously reported associations for current (N = 3,327) and former (N = 172) smoking. A comprehensive smoking index accounting for the biological half-life of smoking compounds and several aspects of smoking history was constructed to assess the reversibility of smoking-induced methylation changes. This measure of lifetime exposure to smoking allowed us to detect more associations than comparing current with never smokers. We identified 4,496 cross-sectional associations at P < 10⁻⁷, including 3,296 annotated to 1,326 genes that were not previously implicated in smoking-associated DNA methylation changes at this significance threshold. We replicated the majority of previously reported associations (P < 10⁻⁷) for current and former smokers. In our data, we observed for former smokers a substantial degree of return to the methylation levels of never smokers, compared with current smokers (median: 74%, IQR = 63-86%), corresponding to small values (median: 2.75, IQR = 1.5–5.25) for the half-life parameter of the comprehensive smoking index. Longitudinal analyses identified 368 sites at which methylation changed upon smoking cessation. Our study demonstrates the usefulness of the comprehensive smoking index to detect associations between smoking and DNA methylation at CpGs across the genome, replicates the vast majority of previously reported associations, and quantifies the reversibility of smoking-induced methylation changes.

KEYWORDS:

• Epigenome-wide association study
• DNA Methylation
• Smoking
• blood
• reversibility
• replication

Disclosure statement

No potential conflict of interest was reported by the authors.

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Additional information

Funding

This work was supported by the Australian National Health and Medical Research Council (NHMRC) [grant 1088405]. MCCS cohort recruitment was funded by VicHealth and Cancer Council Victoria. The MCCS was further supported by Australian NHMRC grants 209057, 251553 and 504711 and by infrastructure provided by Cancer Council Victoria. Cases were ascertained through the Victorian Cancer Registry (VCR) and the Australian Cancer Database (Australian Institute of Health and Welfare). The nested case-control methylation studies were supported by the NHMRC grants 1011618, 1026892, 1027505, 1050198, 1043616 and 1074383. M.C.S. is an NHMRC Senior Research Fellow (1155163).