Purpose: Mutations in RP1 are a common cause of dominant retinitis pigmentosa (RP), but the mechanism by which the identified mutations lead to photoreceptor cell death and blindness has not been determined. To investigate the function of the RP1 protein in photoreceptors and gain insight into the mechanism of disease, gene-targeting techniques were used to produce mice with a mutant Rp1 allele that mimics the truncation alleles found to cause disease.
Methods: RT-PCR was used to amplify illegitimate RP1 transcripts from lymphoblasts. Gene targeting was used to create mice with a mutant Rp1-myc allele. Confocal immunofluorescence microscopy was used to identify the location of the mutant Rp1-myc protein in photoreceptors. The structure of the photoreceptors in the resultant Rp1-myc mice was studied by light and electron microscopy. The retinal function of the mutant mice was investigated using analysis of full-field ERGs.
Results: Wild-type and mutant RP1 mRNA were both detected in lymphoblasts from patients with RP1 disease. Rp1-myc mice produced a truncated version of the Rp1 protein, containing the N-terminal 662 amino acids, which localized correctly to the axoneme of the photoreceptor outer segments. Mice homozygous for the mutant Rp1-myc allele underwent a rapid-onset retinal degeneration characterized by incorrectly oriented outer segment discs that failed to stack properly into outer segments. In contrast, the photoreceptors of heterozygous mice remained relatively healthy.
Conclusions: The presence of mutant RP1 mRNA in lymphoblasts from patients with RP1 disease implies that the mutant message can escape nonsense-mediated mRNA decay and that a truncated RP1 protein may be produced in the retina. The truncated Rp1-myc protein appears to be nonfunctional, and not to exert a dominant negative effect in the photoreceptors of heterozygous mice. Results from homozygous Rp1-myc mice indicate that RP1 is required for the correct orientation and higher order stacking of outer segment discs.