SAT-450 Chronic kidney disease and End-stage Kidney Disease: Major Comorbid Conditions at Initial Presentations of Systemic Lupus Erythematosus population

Abstract

Introduction: Hereditary podocytopathies are genetically highly heterogeneous. Extra-renal symptoms, when associated, are specific for the causal gene. Two unrelated pedigrees presented with a novel infantileonset severe disorder characterized by steroid-resistant nephrotic syndrome (SRNS), cataracts, sensorineural deafness and enterocolitis. In the first four-generation pedigree (FamA), the disorder segregated in an X-linked pattern and in an autosomal recessive mode in the second (FamB). Since the disorder did not correspond to any of the known podocytopathies, we aimed to identify the causal genes. Methods: We performed linkage analysis and multiple parallel sequencing to identify the causal mutations. Flow cytometry, Southern blot and MM-qPCR to measure telomere length. X-inactivation was assessed by qPCR. Molecular dynamics simulations, co-immunoprecipitation and pressure tuning fluorescence spectroscopy were used to assess the complex formation capacity of the mutant proteins. Zebrafish dkc1 mutants were generated using CRISPR/Cas9. Pseudouridylation was assessed In patients and zebrafish by immuno-Northern and HPLC-MS. Whole transcriptome analysis was performed in zebrafish mutants. The localisation of the mutant protein was determined by immunocytochemistry. Results: Linkage analysis in Family A idenified a 5 Mb region in Xq28 and a homozygous region of 2.6Mb in 15q14 in FamB. We found a DKC1 mutation (c.616 G>A, p.Glu206Lys) in FamA which appeared de novo in the second generation in parallel with the disorder and a homozygous NOP10 mutation (c.47C>T, p.Thr16Met) in FamB. Both mutations affected evolutionary conserved amino acids, were predicted to be pathogenic and were absent from population databases. The affected proteins, dyskerin and NOP10, are core members of the H/ACA snoRNP complexes, implicated in the pseudouridylation of RNAs and in telomere maintenance. Based on molecular dynamics simulations, the two affected amino acids interact with each other in the wild type H/ACA snoRNP complex. We found both of their mutations to impair the dyskerin-NOP10 interaction as reflected in an altered dissociation constant and to also disrupt the pseudouridylation catalytic site. We found both telomere attrition and a pseudouridylation defect in the patients. Despite telomere attrition, no mucocutaneous features were suggestive of dyskeratosis congenita. Zebrafish dkc1 null mutants recapitulated the human phenotype, which was rescued by the wild type, but not by the p.Glu206Lys human dyskerin. Mutant larvae showed reduced 18S pseudouridylation and low amount of 18S rRNA, but no telomere attrition. Transcriptomic analysis of the mutant larvae also suggested ribosome biogenesis defect. We found wild type embryos to develop a similar phenotype after cycloheximide treatment. Conclusions: We described a novel SRNS phenotype secondary to DKC1 and NOP10 mutations and propose that it represents the consequences of defective pseudouridylation and ribosomal dysfunction.Introduction: The systemic lupus erythematosus (SLE)...