A gain-of-function variant in NNT causes premature diffuse familial sebaceous hyperplasia.

Premature diffuse familial sebaceous hyperplasia (PDFSH) constitutes a distinct clinical variant of sebaceous hyperplasia, characterized by three hallmark features: early disease onset, characteristic sparing of perioral and periocular regions, and a positive family history. To date, the pathogenic gene underlying PDFSH remains unidentified. The aim of this study was to identify the underlying gene and the pathogenesis of three familial cases with autosomal dominant PDFSH. Whole-exome sequencing was performed in two unrelated families of autosomal dominant PDFSH. The identified candidate gene was further screened for variants in an additional case using Sanger sequencing. The ultrastructure of sebaceous glands was analyzed by transmission electron microscopy (TEM). Immunofluorescence staining was performed to assess lipid peroxidation levels in sebaceous glands. Functional analyses included quantification of NADPH/NADP⁺ ratio, glutathione (GSH) levels, and reactive oxygen species (ROS) levels. Flow cytometry with C11-BODIPY and propidium iodide (PI) staining was performed to assess lipid peroxidation and cell viability, respectively. We identified a missense variant c.2063T>G (p.Leu688Trp) in NNT in all affected members across the three PDFSH families. Both patient-derived keratinocytes and NNT-knockdown SZ95 sebocytes overexpressing the mutant nicotinamide nucleotide transhydrogenase (NNT) exhibited enhanced antioxidant capacity, evidenced by elevated NADPH/NADP⁺ ratio, increased GSH levels, and reduced ROS production compared to controls. Ultrastructural analysis revealed a decreased proportion of mitochondria with cristae disorganization, and immunofluorescence staining showed reduced levels of lipid peroxidation in the patient's sebaceous glands, suggesting decreased ferroptosis susceptibility. In vitro experiments confirmed that the NNT c.2063T>G variant protects SZ95 sebocytes from ferroptosis through oxidative stress mitigation. We identified a gain-of-function variant c.2063T>G (p.Leu688Trp) in NNT underlying PDFSH. This genetic variant enhances antioxidant capacity of NNT while attenuating intracellularly accumulated ROS levels, and reduces sebaceous glands' susceptibility to ferroptosis.