Introduction: The objective of this study was to identify genetic variants that are associated with adult leisure time exercise behavior using genome-wide association (GWA) in two independent samples. Methods: Exercise behavior was measured in 1644 unrelated Dutch and 978 unrelated American adults of European ancestry with detailed questions about type, frequency, and duration of exercise. Individuals were classified into regular exercisers or nonexercisers using a threshold of 4 MET•h (metabolic equivalents-hours per week). GWA analyses of ∼1.6 million observed and imputed Single Nucleotide Polymorphism (SNP) were conducted in both samples independently using logistic regression in SNPTEST, including sex, age, and body mass index as covariates. A meta-analysis of the results was performed using the weighted inverse variance method in METAL. Results: Thirty-seven novel SNPs in the PAPSS2 gene and in two intergenic regions on chromosomes 2q33.1 and 18p11.32 were associated with exercise participation (pooled P values <1.0 × 10 -5 ). Previously reported associations (ACE, CASR, CYP19A1, DRD2, LEPR, and MC4R genes) or linkage findings (2p22.3, 4q28, 4q31.21 7p13, 9q31, 11p15, 13q22, 15q13, 18q12.2, 18q21.1, 19p13.3, and 20q12) were not replicated, although suggestive evidence was found for association to rs12405556 in the LEPR gene (pooled P value 9.7 × 10 -4 ; American sample, P value 9.8 × 10 -5 ) and for association to rs8036270 in the GABRG3 gene (pooled P value 4.6 × 10 -5 ) in the linkage region 15q12-13. Conclusions: The heritability of leisure time exercise behavior is likely to be accounted for by many genetic variants with small effect size. These can be detected by GWA as was shown here for the PAPSS2 gene, but larger samples with genome-wide genotypes and high-quality exercise data are needed for further progress.