Phenotype And Genotype Analysis Of USH2A Gene Related Hereditary Retinal Diseases

Background :The variation of USH2 A gene is related to Usher syndrome type 2 and non-syndromic retinitis pigmentosa with high genetic heterogeneity.At present,896 variants of USH2 A can be retrieved by HGMD.These variants are located in the exon region and the flanking of the intron region.The variant types include nonsense variant,missense variant,splicing variant,chromosome replication and so on.Usherin protein encoded by USH2 A gene plays an important role in retina and inner ear.Because of the large structure of usherin protein,its specific function is not clear.At present,clinical trials of gene replacement therapy have acquired good effects in patients with retinitis pigmentosa caused by some genes.However,due to the excessive size of USH2 A gene exceeds the capacity of the vector commonly used at present,the research is primarily concentrated on animal experiments,and there is no breakthrough in Usher syndrome type 2 and non-syndromic retinitis pigmentosa caused by this gene variants.Purpose:After the study of this subject,we can further understand the association between phenotype and genotype of USH2 A gene-related diseases.On the basis of all-sided analysis genes and clinical diagnosis to further determine the occurrence and seriousness of the disease.Method:The patients who suffered from retinitis pigmentosa and Usher syndrome were collected from September 2017 to January 2019 in Henan Eye institute.The subjects were examined by detailed ophthalmology and peripheral venous blood of patients and their families were collected.The second generation sequencing analysis based on targeted capture was performed on some family members,and Sanger sequencing and family co-segregation were verified.According the sequencing results,the families whose pathogenic gene was USH2 A were screened out.The genotype and phenotype association of the found variants,and the conservatism and pathogenicity of the novel variants were analyzed.Result:In this subject,three pedigrees with pathogenic genes of USH2 A were collected.In the F1 family,In addition to the clinical phenotype of RP,the proband was also complicated with sensorineural deafness.Sequencing revealed two novel frameshift variants: c.13877-13880 del AGAC(p.Q4626Pfs)in exon 64 and c.798 del T(p.F266Lfs)in exon 5 of USH2 A.The homology analysis of the two variants among multiple mammals revealed a high degree of conservation.The variants lead to premature termination of polypeptide chain synthesis,abnormal amino acid translation,and loss of protein function.The function of the novel variations were predicted by online tools SIFT,Polyphen2 and Mutation Taster,and they were predicted to be pernicious.In the F2 family,both proband and his brother showed RP signs without deafness.A novel heterozygous variants c.6986C>A(p.P2329H)in exon 37 and two known heterozygous variants c.15178T>C(p.S5060 P)in exon 70 and c.5836C>T(p.R1946X)in exon 29 of USH2 A were identified.The homology analysis of the novel variant among multiple mammals revealed a high degree of conservation.c.6986C>A may lead to changes in amino acid sequence impacting protein function.c.15178T>C may affect the protein function,and c.5836C>T is a stop pathogenic variant which prematurely terminates protein synthesis.In the F3 family,the proband showed RP signs without deafness.A novel heterozygous missense variant c.14951C>T(p.P4984L)in exon 68 and a known variant c.11156G>A(p.R3719H)in exon 57 of USH2 A were found.The homology analysis of c.14951C>T among multiple mammals revealed a high degree of conservation and the mutation may lead to changes in amino acid sequence affecting protein function.Conclusion:Seven heterozygous variants were found in USH2 A gene in three families,and four of them were novel.The variants of USH2 A gene are the main reason of Usher syndrome type 2 and non-syndromic RP.Different variants affect protein translation and synthesis with varying degrees,consequently causing different clinical phenotypes.