Studies On Phenotype And Pathogenic Gene Of Four Ocular Albinism Type 1 Families

ObjectiveOcular Albinism Type 1（OA1, OMIM 300500） is the most common ocular albinism with the prevalence of 1,50000 from foreign report. Affected patients mainly present with congenital nystagmus, poor visual acuity, iris and fundus hypopigmentation and macular hypoplasia. This disease follows X-linked inheritance pattern and GPR143 gene was found to be the only disease-causative gene. In this study, we describe the clinical features of the four ocular albinism type 1 families（OA1-1―4） and identify the molecular defect. Molecular genetics studies were carried out, including directed sequencing of GPR143 gene to screen the pathogenic mutations and further analyzed the effect of the mutation on the protein function.Methods1. We collect four ocular albinism type 1 families referred to the Department of Pediatric ophthalmology and Strabismus of Tianjin Eye Hospital for congenital nystagmus from October 2014 to October 2015 and a comprehensive eye examination was performed for the affected males and female carriers including best vision acuity, refractive error, slit-lamp microscope, direct ophthalmoscope, fundus photography and optical coherence tomography（OCT） examination, ERG and VEP.2. Human genomic DNA was extracted from 5ml peripheral blood of the family members. The whole nine coding exons and exon-intron boundaries of GPR143 gene were amplified by polymerase chain reaction（PCR）. The PCR products were bi-directionally sequenced on ABI 3130-avant Genetic Analyzer after purification. The DNAstar software was used to analyze data to screen the pathogenic mutations. The pathogenic mutation detected was confirmed by direct sequencing method for available other family members and 100 normal controls.3. Biomedical prediction softwares were used to analyze the influence of the GPR143 gene mutation on OA1 protein. Polyphen 2 and SIFT softwares were used to analyze the screened GPR143 missense mutations, and the Human Splicing Finder-version 3.0 software was used for forecasting the influence on OA1 protein made by splicing mutations.Results1. All affected males in the four ocular albinism type 1 families showed clinical features of congenital nystagmus and poor vision acuity. With exception of one patient with typical albinotic fundus: retina hypopigmentation diffusely to see choroid blood vessels and macular hypoplasia, other patients showed unconspicuous pigmentary change of iris and fundus from mild hypopigmentation to normal. However, the results of the fundus and OCT examination of all affected males showed macular hypoplasia.2. Four mutations were identified in the four OA1 families after directed sequencing of the GPR143 gene including two novel mutations and two known mutations. The two novel mutations included one splicing mutation c.659-2A>C in intron5-6, and a cross deletion g.4572₅239del688 which comprised almost the whole exon 2. One of the two known mutations was a missense mutation c.251G>A（p.G84D） in exon 2 and the other was a nonsense mutation c.733C>T（p.R245X） in exon 6.3. Polyphen 2 and SIFT softwares were used to analyse the mutation c.251G>A（p.G84D） with the obtained value 0.980 and 0.01 respectively, indicating the mutation had a destructive effect on OA1 protein structure. The Human Splicing Finder-version 3.0 was used to analyze the splicing mutation c.659-2A>C and showed the Conserved Value（CV） score of WT was 88.93 and the CV score of mutant was 59.98 and the score variation（between WT and Mutant） was-32.55%. This result showed the splicing mutation broken the WT GPR143 gene splicing site and changed the exon splicing site to create new amino acid sequence and the OA1 protein structure and function was damaged.Conclusion1. The Chinese OA1 patients went to hospital with congenital nystagmus, and other clinical symptoms, especially the iris and fundus hypopigmentation were not obvious so that these patients could be mistaken for idiopathic nystagmus. OCT examination for macular hypoplasia and molecular genetic research were helpful to the diagnosis and differential diagnosis of OA1.2. The four GPR143 mutations c.251G>A 、 c.733C>T 、 c.659-2A>C g.4572₅239del688 were the disease-causative molecular basis for the four OA1 families respectively and the two novel extend the spectrum of GPR143 mutations.