Cytological Study On The Pathogenic Mechanism Of SLC4A1 Gene Mutation R388C That Causes Primary Type Ⅰ Renal Tubular Acidosis

Background: Primary distal renal tubular acidosis(dRTA)is a rare genetic disease,characterized by distal renal tubular dysfunction leading to symptoms of metabolic acidosis and alkaline urine,about 30%-50% Of dRTA patients will have hypokalemia at the same time.However,due to the extremely low incidence of primary dRTA and the strong heterogeneity of clinical manifestations,some atypical dRTA patients may not be diagnosed until adulthood,leading to the occurrence of related complications.So,the early diagnosis and treatment of atypical dRTA is very important.So far,mutations in five genes have been found to cause dRTA in human beings: SLC4A1,ATP6V1B1,ATP6V0A4,FOXI1,and WDR72.Molecular diagnosis,especially next-generation sequencing provides the possibility of early diagnosis for primary dRTA patients with atypical clinical symptoms.Objective: This article aims to improve the diagnostic ability of patients suspected of dRTA in hypokalemia through next-generation sequencing,apply database and functional software to analyze and predict the discovered mutation sites,and study the pathogenesis of the discovered mutation sites through cell models.Methods: Screening patients with hypokalemia in the Department of Endocrinology in our hospital,and sequencing hypokalemia-related genes in patients with unclear etiology by next-generation sequencing.Further cytological research on the discovered dRTA-related gene mutations by constructing wild-type and mutant plasmids,expressing them in 293 A cells,and observing the morphology and distribution of wild-type and mutant proteins in cells.Results: A total of 13 patients with hypokalemia of unknown etiology were subjected to next-generation sequencing.Among them,5 patients were found to be diagnosed as Gitelman syndrome,1 patient was diagnosed as hypokalemic periodic paralysis,1 patient was diagnosed as Liddle syndrome and 1 patient was diagnosed as primary dRTA,the genetic diagnosis of 5 patiens was negative.Among them,patient with primary dRTA and his father both carry a heterozygous mutation(c.1162C>T,p.Arg388Cys)located in exon 11 of the SLC4A1 gene.The expressed mutant amino acids are located in the duplex helix 1(H1)region of the N-terminal of k AE1 protein and belong to highly conserved region,which may change the conformation of k AE1 and affect the function of the protein.Cytological experiments confirmed that compared with the wild-type,the mutant protein was retained in the cell and could not be transported to the cell surface to perform its normal function.It was confirmed that the R388 C mutation of the SLC4A1 gene was the cause of the primary dominant genetic type dRTA in this family.Conclusion: Next-generation sequencing has strong clinical and practical significance for the diagnosis of hypokalemia of unknown etiology.This study found a new mutation site R388 C located in the SLC4A1 gene,and proved that the R388 C mutation of the SLC4A1 gene is the cause of the primary dominant genetic type dRTA by affecting the normal transmission of the expressed protein.