Premature termination codons in PRPF31 cause retinitis pigmentosa via haploinsufficiency due to nonsense-mediated mRNA decay

Abstract

Dominant mutations in the gene encoding the mRNA splicing factor PRPF31 cause retinitis pigmentosa, a hereditary form of retinal degeneration. Most of these mutations are characterized by DNA changes that lead to premature termination codons. We investigated 6 different PRPF31 mutations, represented by single-base substitutions or microdeletions, in cell lines derived from 9 patients with dominant retinitis pigmentosa. Five of these mutations lead to premature termination codons, and 1 leads to the skipping of exon 2. Allele-specific measurement of PRPF31 transcripts revealed a strong reduction in the expression of mutant alleles. As a consequence, total PRPF31 protein abundance was decreased, and no truncated proteins were detected. Subnuclear localization of the full-length PRPF31 that was present remained unaffected. Blocking nonsense-mediated mRNA decay significantly restored the amount of mutant PRPF31 mRNA but did not restore the synthesis of mutant proteins, even in conjunction with inhibitors of protein degradation pathways. Our results indicate that most PRPF31 mutations ultimately result in null alleles through the activation of surveillance mechanisms that inactivate mutant mRNA and, possibly, proteins. Furthermore, these data provide compelling evidence that the pathogenic effect of PRPF31 mutations is likely due to haploinsufficiency rather than to gain of function.

A single-base substitution within an intronic repetitive element causes dominant retinitis pigmentosa with reduced penetrance

“…However, the vast majority of PRPF31 mutations create premature termination codon (PTC) in the mRNA transcribed from the mutant allele, which in turn is degraded by nonsense-mediated mRNA decay (NMD). In such cases, the absence of PRPF31 mRNA and protein derived from the mutant allele has been shown to be the likely cause of the disease [Rio Frio et al, 2008b]. …”

The splicing factor Prp31 is essential for photoreceptor development in Drosophila

“…The fact that down-regulating DmPrp31 gene expression leading to photoreceptor degeneration supports a haploinsufficiency model that has been suggested from studies using non-photoreceptor cell lines derived from RP11 patients (Deery et al, 2002;Wilkie et al, 2006;Rio Frio et al, 2008;Wilkie et al, 2008). This is different from the gain-of-function toxicity model suggested by our previous studies using the mammalian cell culture system.…”

“…The molecular mechanisms underlying this incomplete penetrance of Prp31 mutations in patients remain to be investigated. Possible explanations include differential expression of the wild type allele among different carriers or asymptomatic patients (Vithana et al, 2003) or the presence of a modifier gene (Rio Frio et al, 2008).…”

Suppressors of the cdc-25.1(gf)-associated intestinal hyperplasia reveal important maternal roles for prp-8 and a subset of splicing factors in C. elegans

“…5C), while loss of function in the ortholog of hPrp31, which triggers RP11 due to haploinsufficiency (Rio Frio et al 2008) similarly suppresses the intestinal hyperplasia in our RNAi assay (Table 2). hPrp3 triggers RP most probably in a dominantnegative manner (Gonzalez-Santos et al 2008), and lossof-function in its C. elegans ortholog did not suppress cdc-25.1(rr31).…”