BACKGROUND & AIMS: A genome-wide association study associated 5 genetic variants with hepatic steatosis (identified by computerized tomography) in individuals of European ancestry. We investigated whether these variants were associated with measures of hepatic steatosis (HS) in non-Hispanic White (NHW), non-Hispanic Black (NHB), and Mexican-American (MA) participants in the US population-based National Health and Nutrition Examination Survey (NHANES) III, Phase 2. METHODS: We analyzed data from 4804 adults (1825 NHW, 1442 NHB, and 1537 MA; 51.7% female; mean age of 42.5 y at examination); the weighted prevalence of HS was 37.3%. We investigated whether ultrasound-measured HS, with and without increased levels of alanine aminotransferase (ALT), or level of ALT alone, was associated with rs738409 ( PNPLA3), rs2228603 ( NCAN), rs12137855 ( LYPLAL1), rs780094 ( GCKR), and rs4240624 ( PPP1R3B) using regression modeling in an additive genetic model. We controlled for age, age-squared, sex, and alcohol consumption. RESULTS: The G allele of rs738409 ( PNPLA3) and T allele of rs780094 ( GCKR) were associated with HS with a high level of ALT (odds ratios [ORs], 1.36;P =.01 and 1.30;P =.03, respectively). The A allele of rs4240624 ( PPP1R3B) and the T allele of rs2228603 ( NCAN) were associated with HS (ORs, 1.28;P =0.03 and 1.40;P =0.04, respectively). Variants ofPNPLA3andNCAN were associated with level of ALT among all 3 ancestries. Some single nucleotide polymorphisms were associated with particular races or ethnicities: variants in PNPLA3,NCAN,GCKR, andPPP1R3B were associated with NHW and variants inPNPLA3were associated with MA. No variants were associated with NHB. CONCLUSION: We used data from NHANES III to validate the association between rs738409 ( PNPLA3), rs780094 ( GCKR), and rs4240624 ( PPP1R3B) with HS, with or without increased levels of ALT, among 3 different ancestries. Some but not all associations between variants in NCAN,LYPLAL1,GCKR, andPPP1R3B with HS (with and without increased level of ALT) were significant within subpopulations.