Comprehensive evaluation of smoking exposures and their interactions on DNA methylation

Abstract

Background: Smoking impacts DNA methylation, but data are lacking on smoking-related differential methylation by sex or dietary intake, recent smoking cessation (<1 year), persistence of differential methylation from in utero smoking exposure, and effects of environmental tobacco smoke (ETS).

Methods: We meta-analysed data from up to 15,014 adults across 5 cohorts with DNA methylation measured in blood using Illumina's EPIC array for current smoking (2560 exposed), quit < 1 year (500 exposed), in utero (286 exposed), and ETS exposure (676 exposed). We also evaluated the interaction of current smoking with sex or diet (fibre, folate, and vitamin C).

Findings: Using false discovery rate (FDR < 0.05), 65,857 CpGs were differentially methylated in relation to current smoking, 4025 with recent quitting, 594 with in utero exposure, and 6 with ETS. Most current smoking CpGs attenuated within a year of quitting. CpGs related to in utero exposure in adults were enriched for those previously observed in newborns. Differential methylation by current smoking at 4-71 CpGs may be modified by sex or dietary intake. Nearly half (35-50%) of differentially methylated CpGs on the 450 K array were associated with blood gene expression. Current smoking and in utero smoking CpGs implicated 3049 and 1067 druggable targets, including chemotherapy drugs.

Interpretation: Many smoking-related methylation sites were identified with Illumina's EPIC array. Most signals revert to levels observed in never smokers within a year of cessation. Many in utero smoking CpGs persist into adulthood. Smoking-related druggable targets may provide insights into cancer treatment response and shared mechanisms across smoking-related diseases.

Funding: Intramural Research Program of the National Institutes of Health, Norwegian Ministry of Health and Care Services and the Ministry of Education and Research, Chief Scientist Office of the Scottish Government Health Directorates and the Scottish Funding Council, Medical Research Council UK and the Wellcome Trust.

Keywords: Dietary intake; Epigenomics; Illumina EPIC array; Secondhand smoke exposure; Sex difference; Smoking cessation.

Fig. 1

Schematic of analyses and results. Depicts the analyses conducted, the studies that contributed to each analysis, the results, and downstream analyses that were conducted.

Fig. 2

Miami plot of meta-analysed results for A) current smoking (2560 exposed vs 8521 unexposed) and B) in utero smoking exposure (286 exposed vs 1982 unexposed). In each Miami plot, the top portion of the graph shows the −log₁₀ p-value of all CpGs with a positive effect estimate. The bottom portion of the graph shows the −log₁₀ p-value of all CpGs with an inverse effect estimate. The top five CpGs with higher (top) or lower (bottom) differential methylation are annotated. Blue horizontal line is the FWER threshold (p = 9E-08) and the dashed line is the FDR threshold.

Fig. 3

Heatmap of enriched pathways for implicated genes from current smoking, recent quitting, in utero smoking exposure, and environmental tobacco smoke exposure models. Column indicates the smoking model. Rows are the specific pathways. Darker shade of red means more significant enrichment.