BACKGROUND/OBJECTIVES: Homocysteine (Hcy) is a key intermediate in methionine metabolism. A high plasma concentration of Hcy is an independent risk factor for cardiovascular diseases among other determinants. In this study, we aimed to investigate the interactions between methylenetetrahydrofolate reductase enzyme gene (MTHFR) polymorphisms and lifestyle variables (smoking, alcohol intake and physical activity) on Hcy concentrations in a young Brazilian population. SUBJECTS/METHODS: The study population comprised 3803 individuals from the Pelotas Birth Cohort, aged 22-23 years. Allelic discrimination assays and chemiluminescence immunoassays were performed for genotyping and serum Hcy measurements, respectively. Linear regression models were used to explore the effect of gene-lifestyle interactions on Hcy concentrations. RESULTS: Men carrying the MTHFR 677TT genotype, who were also smokers and drinkers (⩾15 g of alcohol per day), had the highest concentration of Hcy (P-value for the interaction o 0.001 for smoking and 0.002 for alcohol intake). In contrast, high folate concentrations attenuated the effects of the MTHFR C677T genotype on serum Hcy concentrations (P-value for interaction o 0.001). Also, among males, blood folate concentration was the only lifestyle variable able to modify the influence of MTHFR A1298C genotypes on Hcy concentrations (P-value for the interaction o 0.001). There was no strong evidence of an interaction between the MTHFR genotypes and the lifestyle variables in women. CONCLUSIONS: In summary, our study demonstrates a sex difference in Hcy concentrations among Brazilian young adults regarding MTHFR C677T-lifestyle interactions that are worsened under conditions of low blood folate. Identification of potentially modifiable factors related to an increase in homocysteine in young adults, especially in those who are genetically susceptible, is important to prevent negative health consequences in the future.