Mutation Screening And Pathogenic Mechanism Of ALDH3A2 Gene In Sjogren-Larsson Syndrome

Objective To detect the mutations of ALDH3A2 gene in sjogren-Larsson syndrome(Sjogren-Larsson syndrome,SLS)and functional experiments were performed on the gene.To study the type and characteristics of ALDH3A2 gene mutations in SLS patients,and to lay the foundation for further study on the pathogenesis of SLS induced by mutations in ALDH3A2 gene.Methods One case of SLS in Qingdao,Shandong Province was selected as the study object.The peripheral blood of patient and her parent was extracted and the whole exons of ALDH3A2 gene coding region were amplified by polymerase chain reaction(PCR)and then sequenced.In the second part of this study,the functional changes of the new mutation(C.723C> G)found in ALDH3A2 were studied.RNA was extracted from fresh liver tissue and subjected to reverse transcription,the corresponding primers were designed to amplify the ALDH3A2 gene whole coding region.After ALDH3A2wild-type eukaryotic expression vector was constructed,the construction of ALDH3A2 gene mutant was carried out by site-directed mutagenesis kit.The wild type expression vector and mutant expression vector were sequenced.The human skin fibroblasts were transfected with wild-type and mutant plasmids respectively.After 48 hours of transfection,cells were harvested and the total protein was extracted.The effect of mutation on the activity of FALDH was detected by a multifunctional microplate reader.The mutant activity was analyzed by comparison with wild type.Results In the ALDH3A2 gene of the subject,we found two Complex heterozygous nucleotide variation,c.723C> G(coding region No.723 nucleotides from C to G),c.1157A> G(coding region No.1157 nucleotides from A to G)(P.Cys241Trp).The above variation resulted in the change of amino acid of No.241 from Cys to Trp(p.Cys241Trp),the amino acid of No.386 was changed from Asn to Ser(p.Asn386Ser),and both of them were missense mutations.The heterozygous mutation of C> G was found at the c.723 of the subject,the father had no abnormality at that site.The heterozygous mutation of A>G was found at the parent c.1157 where the mother has nothing abnormal detected.The wild and mutant expression vectors were tested for enzymatic activity,and mutations resulted in a decrease in FALDH activity compared to wild type.Conclusion The heterozygous variants found in the ALDH3A2 gene were independently inherited from her parents,and the parents were heterozygous.The mutations were the pathogenicity of the subjects.The pathogenicity of mutation c.1157A> G has been reported in the literature and is associated with Sjoegren-Larsson syndrome.Mutation c.723C> G pathogenicity has not been reported in the literature,its functional studies confirmed that c.723C> G is the pathogenesis of the disease.