Pathogenicity Of A New Site Mutation Of DNAAF4 Gene (c.1118G>A) On Primary Ciliary Dyskinesia

Objective: To screen the pathogenic gene of a patient with PCD and explore its pathogenic mechanism.Methods: In the course of clinical diagnosis and treatment,our research group collected a female patient with bronchiectasis,sinusitis and total visceral translocation as the main clinical manifestations.The ciliary ultrastructure was observed by transmission electron microscope and immunofluorescence microscope,and the whole exome sequencing was carried out to screen the pathogenic genes.The screened pathogenic genes were sequenced in their family members by Sanger sequencing to analyze the genetic mode.Further,HEK293 T tool cells were used as vectors to construct wild-type DNAAF4 plasmid,mutant DNAAF4 plasmid and the plasmid of its downstream binding protein DNAAF2.The effects of DNAAF4 gene mutation on DNAAF4 protein expression,stability and binding ability with downstream proteins were explored by cell transfection,cycloheximide experiment and immunocoprecipitation technology.So as to reveal the pathogenic mechanism of pathogenic genes.Results:1.In the patients with recurrent cough and expectoration,bronchiectasis,chronic sinusitis and total visceral transposition found in this study,cilia transmission electron microscopy and immunofluorescence staining showed partial loss of internal and external power arms,and the diagnosis of Kartagener syndrome was clear.The mutation of DNAAF4(c.1118G>A)was found by full exon sequencing,and the parents and sisters were carriers by Sanger sequencing.It can be seen that the mutation is consistent with autosomal recessive inheritance,which is consistent with the genetic mode of PCD.2.Using cell transfection,cycloheximide experiment and immunocoprecipitation technology to explore the effect of new site mutation of DNAAF4 gene on the expression and stability of DNAAF4 protein and the binding ability with downstream proteins.It was found that DNAAF4 gene mutation did not affect the expression of DNAAF4 protein in cells or the binding ability between DNAAF4 protein and downstream DNAAF2 protein,but played a pathogenic role by reducing the stability of DNAAF4 protein..Conclusion: the new site mutation of DNAAF4 gene(c.1118G>A)plays a pathogenic role in PCD by reducing the stability of DNAAF4 protein.