NPHS2 Revisited Through 208 Cases and Podocin Complex Modeling

IntroductionSteroid-resistant nephrotic syndrome (SRNS) is the leading cause of chronic glomerular disease in individuals under 25 years of age.Biallelic variants in NPHS2, encoding podocin, are the most common monogenic etiology.Podocin homo-oligomerizes to a critical slit diaphragm complex.NPHS2 genotype-phenotype relationships and podocin's assembly architecture remain incompletely defined. MethodsAnalysis of 208 individuals with pathogenic NPHS2 variants, the largest reported single cohort.Genotype-phenotype correlations were tested using clinical data, in silico pathogenicity tools (REVEL, EVE), and high-order structural modeling (AlphaFold3) based on published experimental data. ResultsA broad allelic spectrum with variable disease onset (median 2.75 years; range 0-30) was identified.While most cases presented early, later onset was associated with compound heterozygous variants with p.R229Q (including in trans with novel variant p.E281K), the homozygous missense variant p.V180M, or select compound heterozygous variants with p.R138Q (in trans with p.V180M or p.V290M).REVEL and EVE scores reliably distinguished pathogenic from benign missense variants.A novel AIgenerated architecture of podocin higher-order assemblies, featuring a chalice-like J o u r n a l P r e -p r o o f structure with a central hydrophobic core and a single transmembrane plane, is proposed, resembling the solved structures of multiple paralogous SPFH family proteins.Pathogenic missense variants in the proposed model cluster at oligomer interfaces, with distinct physicochemical alterations, and variants associated with earlier onset appear to localize closer to the transmembrane plane. ConclusionThis study delineates genotype-phenotype relationships in NPHS2-associated SRNS and proposes high-order podocin assemblies.Combined genetic and structural insights enhance variant assessment, though experimental validation remains essential.