Siquan Zhu

A novel mutation in AlphaA-crystallin (CRYAA) caused autosomal dominant congenital cataract in a large Chinese family†‡

Hereditary cataract is a phenotypically and genetically heterogeneous lens disease that is responsible for a significant proportion of the visual impairment and blindness that occurs in children. In a five‐generation Chinese family with autosomal dominant inherited congenital cataract, clinical examination showed three cataract phenotypes: punctuate, nuclear, and total cataracts. Linkage analysis was performed and positive two‐point LOD scores (with maximum of 4.43 and 4.27 at θ=0) were obtained for markers D21S1411 and D21S1890 on chromosome 21q22.3, flanking the CRYAA (alphaA‐crystallin‐encoding gene) locus. Sequencing of CRYAA revealed a novel heterozygous G>A transition (c.346G>A) in exon 3 that cosegregated with the disease phenotype and results in a conservative substitution of Arg to His at codon 116 (p.R116H). To understand the molecular basis of cataract formation, mutant and wild‐type alphaA‐crystallins were expressed in E. coli. RP‐HPLC (reverse phase‐high‐performance liquid chromatography) suggested an increased hydrophobicity of the mutant recombinant protein, compared to that of wild‐type alphaA‐crystallins. Furthermore, loss of chaperone activity of the mutant was seen in DTT (DL‐dithiothreitol)‐induced insulin aggregation assay. FPLC (fast protein liquid chromatography) purification showed that the His‐116 mutant protein had increased binding affinity to lysozyme. Gain of activated lysozyme binding, elevation of hydrophobicity and loss of chaperone activity of the mutant protein may be some of the molecular mechanisms underlying cataract in this large family.

A R54L Mutation of CRYAA Associated with Autosomal Dominant Nuclear Cataracts in a Chinese Family

Abstract Purpose: To identify the genetic defect in a three-generation Chinese family with congenital cataracts. Methods: The phenotype of a three-generation Chinese family with congenital cataract was recruited. Detailed family history and clinical data of the family were recorded. Candidate genes sequencing was performed to screen out the disease-causing mutation. Bioinformatics analysis was performed to predict the function of mutant gene. Results: The phenotype of the family was identified as nuclear cataract. Direct sequencing revealed a c.161 G > T transversion in exon 1 of crystallin alpha-A (CRYAA). This mutation co-segregated with all affected individuals in the family and was not found in unaffected family members nor in the 100 unrelated controls. Bioinformatics analysis indicated that the 54th amino acid position was highly conserved and the mutation R54L caused an increase of local hydrophobicity around the substitution site. Conclusions: This study identified a novel disease-causing mutation c.161 G > T (p.R54L) in CRYAA in a Chinese family with autosomal dominant nuclear cataracts, this is the first report relating a G > T mutation in CRYAA leading to congenital nuclear cataract.

A Novel Mutation in the Connexin 50 Gene (GJA8) Associated with Autosomal Dominant Congenital Nuclear Cataract in a Chinese Family

Purpose: To identify the genetic defect in a four-generation Chinese family with autosomal dominant congenital nuclear cataract. Methods: Family history data were recorded. Clinical and ophthalmologic examinations were performed on family members. All the members were genotyped with microsatellite markers at loci associated with cataracts. Linkage analysis was performed after genotyping. Candidate genes were screened for mutation using direct sequencing. Results: Linkage analysis was obtained at markers D1S1653 (LOD score [Z] = 1.50, recombination fraction [θ] = 0.0) and D1S498 (LOD score Z = 0.90, recombination fraction [θ] = 0.0), which encompasses the connexin 50 gene (GJA8). Sequencing the coding regions of GJA8 revealed a novel, heterozygous c.773C>T transition that resulted in the substitution of a highly conserved serine by phenylalanine at codon 258 (S258F). Bioinformatics analysis showed that the mutation altered the hydrophobicity and secondary structure of the protein. This mutation co-segregated with the disease phenotype in all affected individuals and was not found in the unaffected family members or in 100 normal unrelated individuals. Conclusions: This study has identified a novel missense mutation located in the carboxyl terminus of GJA8 (S258F) associated with autosomal dominant nuclear cataract.

Autosomal Dominant Congenital Nuclear Cataracts Caused by a CRYAA Gene Mutation

Purpose: We sought to identify the genetic defect in a four-generation Chinese family with autosomal dominant congenital nuclear cataracts, examine the clinical features in detail and demonstrate the functional analysis of a candidate gene in the family. Methods: Family history data were recorded. Clinical and ophthalmological examinations were performed on affected and unaffected family members. All the members were genotyped with microsatellite markers at loci considered to be associated with cataracts. Two-point LOD scores were calculated using the LINKAGE program package after genotyping. A mutation was detected by dilff521229rect sequencing and verified by denaturing high-performance liquid chromatography (DHPLC). Wild-type and mutant proteins were analyzed with online softwares. Results: All affected members of this family had nuclear cataracts. Genetic analysis revealed a heterozygous previously described Arg116Cys mutation in the CRYAA gene in all of the affected members of the family but not in unaffected or 100 normal, unrelated individuals. Data generated with online software revealed that the different amino acid side chain, impact the aa116 interaction with other amino acids, thereby affecting the proteins secondary structure. Conclusions: This study identified a mutation in the CRYAA gene causing autosomal dominant nuclear cataracts and some patients show nystagmus or small blepharophimosis clinical features. These results provide evidence that CRYAA is a pathogenic gene for congenital cataracts, congenital cataracts are a clinically and genetically heterogeneous lens condition; at the same time, demonstrates a possible mechanism of action for the mutant gene.

A G57W Mutation of CRYGS Associated with Autosomal Dominant Pulverulent Cataracts in a Chinese Family.

Congenital cataract is the most common treatable cause of childhood blindness, which is characterized by opacification of all or part of the eye’s crystalline lens within the first year of life. The prevalence of congenital cataracts is 1 to 6 per 10,000 live births. The cataract may be isolated, may be hereditary or secondary to an intrauterine event. Approximately one-quarter to one-third of congenital cataracts are inherited. Currently, four mutations in CRYGS associated with congenital cataract have been reported. Our present study (p. G57W) is the fifth report of a mutation in CRYGS linked with congenital cataract (Table 1). A three-generation Chinese family with autosomal dominant congenital cataract from Anhui province was collected from Beijing Tongren Hospital. We also recruited 100 unrelated subjects without eye diseases except mild myopia from the Ophthalmology Clinic of Beijing Tongren Hospital as normal controls. The ethics committee of Capital Medical University approved the research. All participants from the family gave their informed consent. The study protocol followed the principles of the Declaration of Helsinki. All 13 exon and intron-exon junctions of the candidate genes were amplified by polymerase chain reaction (PCR). The sequencing results were analyzed using Chromas 2.33(http://www.technelysium.com.au/chromas.html). Ten family members of a three-generation Chinese family with a history of cataracts participated in the study (five affected and five unaffected individuals). All patients in this family had bilateral cataracts. Most patients experienced decreased visual acuity at 3 years old, and then their visual acuity decreased gradually until surgery was required. A history of cataract extraction or ophthalmologic examination was used to determine the status affected, and ten family members participated in the study. The proband, who was a 7-year-old boy experienced a decrease in vision at 3 years old and had been diagnosed with bilateral cataracts at age 3. Slit-lamp examination revealed pulverulent cataract in the center. The boy’s best corrected visual acuity was 0.2/0.3. His 31-year-old mother also had pulverulent cataract in the center with peripheral cortical opacity. Her best corrected visual acuity was 0.3/0.4. Their clinical features were similar. Through direct gene sequencing of the coding regions of the candidate genes, we identified a transversion of G4T at c. 169(p. G57W) in exon 2 of crystallin gamma S (CRYGS) in all affected individuals. However, we did not find this mutation in any unaffected family members or in the 100 unrelated controls. We found no further gene mutations in