Genetic Verification Of The Pathogenic Gene Of A New Mutation In ADAR1 In Hereditary Dyschromes Symmetries

Objective: Dyschromatosis symmetrica hereditaria(DSH)is a clinically heterogeneous pigmentary genetic disorder caused by a mutation in adenosine deaminase acting on RNA1(ADAR1)that is inherited in an autosomal dominant manner.This study aimed to analyze the clinical manifestations of a DSH family to investigate the mechanisms underlying pigmentary hyperpigmentation and hypopigmentation on the distal limbs,dorsum of the hands and feet,and face and neck,validate a novel mutation site,explore its pathogenic mechanism,and its relationship with skin lesions.Methods: A patient with suspected DSH was diagnosed clinically,and peripheral blood was collected from the patient,the patient’s father and son with the same skin lesion,family members without skin lesions,and healthy unrelated individuals as controls.Sanger sequencing and whole-exome sequencing were used to validate the ADAR1 mutation site,with preliminary validation indicating a missense mutation c.2668G>C/p.G890 R.Bioinformatics software(SIFT,Poly Phen-2,Mutation Taster,and gnom AD)was used to predict the pathogenicity of the mutation site.Functional experiments were conducted on2668 ADAR1 mutations,including constructing wild-type and mutant plasmids,transfecting HEK293 T cells,measuring m RNA expression using q PCR,measuring protein expression using Western blotting,and measuring fluorescence enzyme activity using the luciferase assay after IFN-γ induction.Statistical analysis software,Image J and Graph Pad,were used to analyze the data,with P<0.05 indicating statistical significance.Results: Both Sanger and whole-exome sequencing validated the presence of a missense mutation(c.2668G>C/p.G890R)in the patient and his father and son that affected the exon encoding the transmembrane protein(Exon8),with the 2668 th base G changing to C,altering the structure and function of the 890 th amino acid from glycine to arginine.The mutation was not detected in family members without skin lesions or healthy unrelated individuals.Multiple-sequence homology analysis revealed its position in a highly conserved region of the protein,and bioinformatics software predicted the mutation to be deleterious.Functional experiments indicated that the m RNA and protein expression of 2668 ADAR1 mutants decreased significantly(P<0.05)compared to wild-type and empty plasmid controls.In addition,the fluorescence enzyme activity of mutant ADAR1 was significantly increased(P<0.05)compared to wild-type ADAR1 after IFN-γ induction.Conclusion: This study identified a novel missense mutation site(c.2668G>C/p.G890R)in ADAR1 that had not been previously reported in HGMD,enriched the mutation spectrum of ADAR1,revealed the related mechanisms underlying DSH pathogenesis by analyzing the functional studies,and confirmed the relationship between the mutation and the phenotype of DSH in the family.This study provides a molecular biology basis for early diagnosis and treatment of DSH and suggests new avenues for exploring the pathogenesis and treatment of DSH.