Human Loss-of-Function <i>ZNRF3</i> Mutation Impairs Adipose Thermogenesis and Drives Subcutaneous Fat Expansion

Zinc and ring finger protein 3 (ZNRF3) functions as a transmembrane E3 ubiquitin ligase. Although genome-wide association studies have implicated ZNRF3 in body fat distribution, the evidence from human genetics and animal models has been lacking. Here, through whole-exome sequencing of 1,944 obese individuals and 2,161 healthy lean control individuals, we identified a rare loss-of-function mutation in human ZNRF3 (p.V228L) observed exclusively in 8 obese individuals. These carriers exhibit increased hip circumference and subcutaneous fat area and lower fasting glucose levels compared with noncarriers. In mice, adipocyte-specific Znrf3 deletion leads to progressive adiposity, with expansion of subcutaneous white adipose tissue (sWAT) and improved glucose tolerance after a long-term high-fat diet. Furthermore, Znrf3 deficiency impairs the browning capacity of sWAT and cold-induced thermogenesis. A knock-in mouse model harboring the homologous human p.V228L mutation recapitulates key features of carriers and knockout mice, including reduced sWAT thermogenic function, altered fat distribution, and improved glucose tolerance. Mechanistically, Znrf3 loss attenuates the cAMP/cAMP-dependent protein kinase signaling pathway, suppressing UCP1 expression and mitochondrial respiration. Collectively, these findings provide genetic and biological evidence that ZNRF3 regulates thermogenic capacity and remodeling of sWAT, offering new insights into obesity heterogeneity. Article Highlights A rare loss-of-function variant in ZNRF3 (p.V228L) is enriched in individuals with obesity and is associated with increased subcutaneous white adipose tissue (sWAT) accumulation and lower fasting glucose levels. Both adipocyte-specific Znrf3 knockout and global variant knock-in impair sWAT browning, increase sWAT expansion, and improve glucose tolerance in mice. These findings establish ZNRF3 as a genetic regulator of fat distribution and thermogenic capacity, informing precise phenotyping of obesity. GWAS has implicated ZNRF3 in human fat distribution, yet its role in adipose tissue biology remains unknown.