Two nonsense mutations of PAX6 in two Japanese aniridia families: case report and review of the literature

PURPOSE. TO identify PAX6 mutations in patients from four Japanese families with aniridia. METHODS. Polymerase chain reaction (PCR)-single stand conformational polymo rphism (SSCP) analysis (SSCA) was performed in probands of the families, an d restriction analysis using MaeIII or Aval was carried out in other affect ed family members. RESULTS. PCR-SSCA demonstrated in the proband from one family an extra-band in the PCR product for PAX6 exon 8. Base sequence analysis revealed that t he patient is a heterozygote for a C to T transition mutation at codon 203. DNAs from the patient and another affected member in the same family were cut with MaeIII into two fragments, while non-affected members in the famil y showed only one MaeIII fragment, the result confirmed the mutation. In an other family, PCR-SSCA revealed an extra-band in the PCR product for exon 9 . Sequencing detected a C --> T substitution at codon 240 in the patient, t he mutation resulted in loss of an Aval site. Aval cleavage analysis confir med the mutation in the patient. The two transition mutations observed in t he two families also predict the conversion of arginine to a stop codon (R2 03X and R240X, respectively) around the homeodomain (HD), leading to the tr uncation of the PAX6 protein within ifs glycine-rich region. No abnormal SS CP bands or abnormal restriction fragments were detected in patients from t he other two families. CONCLUSIONS. The two mutations sites identified in the two families, one at codon 203 and the other at codon 240, are those most frequently observed a mong 118 previously reported PAX6 mutations. This indicates that the two mu tations are two hot-spots in the gene.