Expression of Precursor RNA Processing Factor 31 (PRPF31) mRNA in Patients with Autosomal Dominant Retinitis Pigmentosa: A Molecular `Clue' for Incomplete Penetrance?

Abstract

Abstract: : Purpose:To identify the molecular basis of incomplete penetrance phenotype characteristic of adRP families linked to chromosome 19q13.4 (RP11) with mutations in PRPF31 gene. Analysis of single nucleotide polymorphisms (SNPs) within the PRPF31 gene in sib-ships in RP11 linked families demonstrated that asymptomatic individuals consistently inherited a different wild type allele to the one inherited by their symptomatic siblings. The existence of differentially expressed wild type alleles has been suggested to explain the incomplete penetrance phenotype at this locus. In order to test this hypothesis, we measured wild type mRNA levels in symptomatic and asymptomatic individuals carrying a PRPF31 mutation.Methods:We performed real time quantitative RT-PCR on RNA from lymphoblastoid cell lines derived from a large adRP family (RP856/AD5). All disease gene carriers in this family have an 11bp deletion in exon 11 of PRPF31. In order to quantify only the mRNA levels due to the wild type allele of PRPF31, the probe was designed across the deletion. To test the validity of using RNA from lymphoblastoid cell lines, PRPF31 expression in nucleated blood cells from control individuals and donor retinas was compared with expression in lymphoblastoid cells-showing no significant difference. Mann-Whitney U test was used to compare the median copy number in pairs of the three groups namely the symptomatic and asymptomatic carriers of the mutant PRPF31 allele and non-carrier individuals. The PRPF31 protein levels from symptomatic and asymptomatic individuals were also assayed by western blot analysis using an antibody specific to the wild type PRPF31 protein.Results:We observed a significant difference in wild type PRPF31 mRNA levels between symptomatic and asymptomatic individuals (p<0.001) supported by western blot analysis of the of the PRPF31 protein.Conclusions:We have shown that in our largest adRP pedigree with a deletion mutation in PRPF31, the clinical manifestation of RP could well be due to the co-inheritance of a PRPF31 gene defect and a low expressed wild type allele. This study has revealed a potential avenue for future therapy, as it appears that moderate over expression of wild type PRPF31 may prevent clinical manifestation of the disease.

Keywords: gene/expression • molecular biology • proteins encoded by disease genes