Mutations in the X-linked RP2 gene cause intracellular misrouting and lossof the protein

Mutations in RP2 cause the second most frequent form of X-linked retinitis pigmentosa, a severe retinal degeneration that leads to loss of visual acui ty and blindness, The RP2 gene encodes a protein with homology to cofactor C, a tubulin-folding chaperone, By searching protein sequence databases, we identified a whole set of similar molecules from diverse organisms. Protei n sequence alignments show that RP2 and cofactor C represent members of two distinct orthologous groups. All previously identified missense mutations affect amino acid residues which are conserved in all RP2 orthologues or bo th orthologous groups. Intracellular localization of the wild-type protein and mutated variants was determined by fluorescence microscopy of cells exp ressing RP2 with a green fluorescent protein tag. A mutation in the N-termi nus of RP2 abolishes localization to the plasma membrane in HeLa cells. C-t erminal protein truncation mutations, which account for 2/3 of the pathogen ic RP2 variants, lead to scattered fluorescent foci in the cytoplasm of COS -7 and HeLa cells, Analysis of protein extracts from the respective cells w ith anti-RP2 antibodies identified truncated proteins of expected size in a low-speed centrifugation pellet while the wild-type protein appeared in th e supernatant, Moreover, no protein was detected in immortalized cell lines from patients with protein truncation mutations while mRNA was still prese nt. Thus, loss of the protein and/or aberrant intracellular distribution mi ght be the basis for the photoreceptor cell degeneration in most RP2 cases.