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Xinxin Zhang, Yupu Liu, Fei Liu, Tingyu Yan, Youjin Wang, Yanyan He, Changyang Liu, qian liu, shizhen lei, Xiaochen Wang, Jun Kong; Molecular studies of a novel RPGR gene mutation in a Chinese family with X-linked retinitis pigmentosa. Invest. Ophthalmol. Vis. Sci. 2019;60(9):4479.
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Retinitis pigmentosa (RP) is a group of common genetic ocular fundus diseases that cause nyctalopia and loss of peripheral visual field due to the degeneration of photoreceptor cells and pigment epithelial cells in the retina. X-linked RP is one of the most severe forms of RP, 70-80 percent of which is caused by RPGR gene mutations.This study screened gene mutations in a Chinese family with XLRP to identify the etiology and genotypes of the family members and to explore the mechanisms by which the gene causes RP.
Methods: Peripheral blood of the proband and the family members was gathered to construct genomic library. Target gene sequencing hybrid capture technique was used to detect 1651 gene sequences and to screen target genes. The results were further verified by Sanger sequencing. PyMOL software was used for predicting the three-dimensional structure of wild type and mutants of RPGR. The vectors of wild and mutant type of RPGR gene were constructed and were transfected into Y-79 cells line. Expression of RPGR, rhodopsin and long wavelength opsin was detected using real-time PCR and Western Blot. RNA expression of Gelsolin, which is an important actin regulator can interact with RPGR and has been shown to regulate cilia formation was also detected.
Frameshift mutation c. 2405-2408del of RPGR gene in the X chromosome of proband was discovered by target gene sequencing, which is carried by the mother without RP phenotype and has not been reported to be pathogenic in HGMD database. The mutation was found to be in the α-helix that disappears resulting from premature termination codon by predicting protein structure. Compared with the wild type, mRNA expression level of RPGR, rhodopsin and opsin in cells with mutant genotype decreased significantly (t = 4.836, P < 0.05; t = 4.213, P < 0.05; t = 8.242, P < 0.05, respectively). Western Blot results showed a decrease in RPGR protein expression in cells with mutant genotype. The mRNA expression of Gelsolin (t=3.603, P<0.05) increased after transfection of mutant genotype compared with the wild type.
This novel mutation in RPGR causes RP phenotype in males and no RP phenotype in female carriers. The frameshift mutation of RPGR gene can cause decrease in expression of rhodopsin, opsin and, the visual function proteins, which may be involved in the pathological changes of RP.
This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.
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