Non-alcoholic fatty liver disease (NAFLD) is a multifactorial metabolic syndrome which may progressively lead to nonalcoholic steatohepatitis, cirrhosis, hepatic failure and hepatocellular carcinoma. NAFLD is the most common liver disease in the world. So far there has been no disclosure on the genetic basis of NAFLD inheritance in families. In this study, in seven distinct multi-generation families we found the causative gene for a rare Mendelian form of NAFLD. In a large Italian family with nine affected members with NAFLD and/or dyslipidemia, whole exome sequencing identified a heterozygous stop codon mutation in ABHD5. While we identified a monoallelic mutation in NAFLD in adults, biallelic ABHD5 mutations were previously associated with CDS, a rare, autosomal recessive disease with multi-systemic manifestations including severe ichthyosis and neonatal NAFLD. Therefore, to explore the potential presence of NAFLD in heterozygous carriers in families with CDS, we subjected individuals from six consanguineous Iranian families, including a total of 29 normal appearing ABHD5 carriers. Overall, 35 out of 38 ascertained ABHD5 mutation carriers showed evidence of NAFLD and/or dyslipidemia (estimated penetrance: 92%). The youngest carrier with NAFLD was 29-year old. For estimation of the disease-allele frequency of the ABHD5 locus related to this late-onset disease, we analyzed the DNA sequence variants of ABHD5 gene in normal databases such as gnomAD. The prevalence of ABHD5-associated NAFLD in the general population was estimated 1 in 1131 individuals. In conclusion, our findings disclose the existence of a rare familial form of NAFLD associated with a single gene mutation.