The Study On Pathogenic Gene Of Hereditary Progressive Cortical Cataract

Objective: Cataract remains the leading cause of human blindness worldwide. Congenital cataracts have an estimated frequency of 1 -6 per 10,000 live births, with one third of cases are familial. Congenital cataracts have been reported with all three types of Mendelian inheritance, autosomal dominant, autosomal recessive, and X-linked. Autosomal dominant congenital cataracts with high penetrance appear to be the most common in affected individuals. The autosomal dominant zonular pulverulent cataract (CAE1) locus was initially linked with the Duffy blood-group locus (Renwick and Lawler 1963), and, in 1968, the Duffy bloodgroup locus was assigned to chromosome 1. This mapped the CAE1 locus to human chromosome 1 (Donahue et al. 1968). To date, more than twenty autosomal dominant congenital cataract loci have been mapped to different chromosomal. Recent work in molecular genetics has identified 18 genes involved in the pathogenesis of isolated inherited cataract. However both the relation of genotype and phenotype and of hereditary and age related cataract are unclear. There is a long way to understand the mechenism and precaution of cataract. We studied a family suffering form cortical progressive cataract and. We carried out gene mapping and mutation screening and tried to identify the pathogenic gene and mechenism of cataract. Moreover cataractogenesis and its eventual therapeutic amelioration were investigated. Methods:1. We investigated a six-generation cataract family. After obtaining informed consent Clinical information and blood specimens were obtained from familymembers. Then we performed Classic genetics and cytogenetics analysis.2. Genomic DNA was extracted from peripheral blood leucocytes. Firstly, we studied 11 loci for known candidate genes using three or four fluorescent microsatellite markers per locus and no evidence of linkage was detected. Subsequently a genomewide scan consisting of 382 microsatellite markers spaced at -10 cM intervals was performed using ABI PRISM Linkage Mapping sets. At last we performed fine mapping. The markers' order and position were obtained from the Marshfield Genetic Database. Genotyping and data collection were conducted by ABI Prism GeneMapper v 3.0 software. We carried out two-point linkage analysis using the MLINK program from the LINKAGE v.5.10 software package. Pedigree and haplotype construction were performed by Cyrillic v.2.1 software.3. Candidate genes were selected in the physical map. The sequences of the genes were got from genebank database. The primer was designed by Primer 3 software. We screened the mutation of candidate genes by bidirectional sequencing PCR products.4. We perform bioinformatics analysis of mutant protein, including physical character, function, structure and et al.Results:1. We studied a Chinese six-generation cataract family composed of 119 individuals with a dominant pattern of inheritance. Clinical information and blood specimens were obtained from 64 family members, including 14 patients. The phenotype in this family was characterized by opacities in cortex. The opacities can be located in anterior cortex, posterior or peripheral cortex but no opacity was observed in the fetal nuclear region. Moreover this cataract is of a progressive nature, andcataractous changes were prominent in affected older individuals. Phenotypic variation in the size and density of opacities as well as the position was observed among the 14 affected members, even between the two lenses of an individual. From the pedigree, a autosome dominant pattern of inheritance was recognized. Chromosome2. After gene mapping and haplotype were performed, we identified a new autosomal dominant congenital cataract locus on chromosome 3q26.3-qter. The disease gene lay in the approximately 2.8 Mb physical intervals between D3S1571 and D3S3570. Linkage analysis gave a maximum two point LOD score 6.34 (9=0.00) for marker D3S1602.3. By sequencing CRYGS gene on two directions, we identified a 105G—>T (NMO17541) heterozygous mutation in exon 2 of CRYGS, resulting in a Gly-?Val substitution at codon 18 (NP060011) in 14 affected members. This mutation was not found in 50 unaffected members in this family, neither in 400 chromosomes of 200 unrelated control individuals, excluding the possibility of a rare polymorphism.4. we perform bioinformatics analysis of mutant protein, including function and structure. It has shown that the four-stranded greek key P-sheet peptide corresponding to crystallin fold forms an individual calcium-binding site, and the first calcium ligate at the residue next to the conserved aromatic amino acid of the sequence Y/E/WXXXXXXG, which is located at the end of the first p-strand. Gly is needed for forming a dihedral angle so it is irreplaceable. In this family just this Gly mutated, which may lead to structural alterations in the Ca2+-binding and -storing ability. Additionally, prediction of function sequence detect that "G[RK][RK]" is Amidation site. Gly mutatation may disturb this function, andthen effect the relation between crystallins. These might be novel mechanisms of cataract formation. Understanding non-structural properties of crystallins may be critical for understanding the malfunction in molecular cascades that lead to cataractogenesis and its eventual therapeutic amelioration. It may help to have an understanding of the disease etiology and contribute to better understanding of the function and properties of this gene. Perhaps this study of progressive phenotype and CRYGS mutation may provide insight into the cause of the more common sporadic form of age-related cataracts.Conclusion: Gamma-S crystalline gene (CRYGS) mutation causes hereditary cortical progressive cataract. This report is the first description of a mutation in CRYGS in humans and added another gene to growing list of genes involved in this heterogeneous monogenic disorder.