A silent mutation in human alpha-A crystallin gene in patients with age-related nuclear or cortical cataract.

A cataract is a complex multifactorial disease that results from alterations in the cellular architecture, i.e. lens proteins. Genes associated with the development of lens include crystallin genes. Although crystallins are highly conserved proteins among vertebrates, a significant number of polymorphisms exist in human population. In this study, we screened for polymorphisms in crystallin alpha A (CRYAA) and alpha B (CRYAB) genes in 200 patients over 40 years of age, diagnosed with age-related cataract (ARC; nuclear and cortical cataracts). Genomic DNA was extracted from the peripheral blood. The coding regions of the CRYAA and CRYAB gene were amplified using polymerase chain reaction and subjected to restriction digestion. Restriction fragment length polymorphism (RFLP) was performed using known restriction enzymes for CRYAA and CRYAB genes. Denaturing high performance liquid chromatography and direct sequencing were performed to detect sequence variation in CRYAA gene. In silico analysis of secondary CRYAA mRNA structure was performed using CLC RNA Workbench. RFLP analysis did not show any changes in the restriction sites of CRYAA and CRYAB genes. In 6 patients (4 patients with nuclear cataract and 2 with cortical cataract), sequence analysis of the exon 1 in the CRYAA gene showed a silent single nucleotide polymorphism [D2D] (CRYAA: C to T transition). One of the patients with nuclear cataract was homozygous for this allele. The in silico analysis revealed that D2D mutation results in a compact CRYAA mRNA secondary structure, while the wild type CRYAA mRNA has a weak or loose secondary structure. D2D mutation in the CRYAA gene may be an additional risk factor for progression of ARC.