Exposure to environmental and lifestyle factors, such as cigarette smoking, affect the epigenome and might mediate risk for diseases and cancers. We have performed a genome-wide DNA methylation study to determine the effect of smoke and snuff (smokeless tobacco) on DNA methylation. A total of 95 sites were differentially methylated [false discovery rate (FDR) q-values < 0.05] in smokers and a subset of the differentially methylated loci were also differentially expressed in smokers. We found no sites, neither any biological functions nor molecular processes enriched for smoke-less tobacco-related differential DNA methylation. This suggests that methylation changes are not caused by the basic components of the tobacco but from its burnt products. Instead, we see a clear enrichment (FDR q-value < 0.05) for genes, including CPOX, CDKN1A and PTK2, involved in response to arsenic-containing substance, which agrees with smoke containing small amounts of arsenic. A large number of biological functions and molecular processes with links to disease conditions are also enriched (FDR q-value < 0.05) for smoke-related DNA methylation changes. These include ‘insulin receptor binding', and ‘negative regulation of glucose import' which are associated with diabetes, ‘positive regulation of interleukin-6-mediated signaling pathway', ‘regulation of T-helper 2 cell differentiation', ‘positive regulation of interleukin-13 production' which are associated with the immune system and ‘sertoli cell fate commitment' which is important for male fertility. Since type 2 diabetes, repressed immune system and infertility have previously been associated with smoking, our results suggest that this might be mediated by DNA methylation changes.