RPGR ORF15 isoform co-localizes with RPGRIP1 at centrioles and basal bodies and interacts with nucleophosmin

110

108

RPGR-interacting protein 1 (RPGRIP1) is a key component of cone and rod photoreceptor cells, where it interacts with RPGR (retinitis pigmentosa GTPase regulator). Mutations in RPGRIP1lead to autosomal recessive congenital blindness [Leber congenital amaurosis (LCA)]. Most LCA-associated missense mutations in RPGRIP1 are located in a segment that encodes two C2 domains. Based on the C2 domain of novel protein kinase C (PKC ), we built a 3D-homology model for the C-terminal C2 domain of RPGRIP1. This model revealed a potential Ca 2؉ -binding site that was predicted to be disrupted by a missense mutation in RPGRIP1, which was previously identified in an LCA patient. Through yeast two-hybrid screening of a retinal cDNA library, we found this C2 domain to specifically bind to nephrocystin-4, encoded by NPHP4. Mutations in NPHP4 are associated with nephronophthisis and a combination of nephronophthisis and retinitis pigmentosa called Senior-Løken syndrome (SLSN). We show that RPGRIP1 and nephrocystin-4 interact strongly in vitro and in vivo, and that they colocalize in the retina, matching the panretinal localization pattern of specific RPGRIP1 isoforms. Their interaction is disrupted by either mutations in RPGRIP1, found in patients with LCA, or by mutations in NPHP4, found in patients with nephronophthisis or SLSN. Thus, we provide evidence for the involvement of this disrupted interaction in the retinal dystrophy of both SLSN and LCA patients.T he X-linked gene RPGR (retinitis pigmentosa GTPase regulator) is mutated in patients with retinitis pigmentosa type 3 (RP3) (1, 2), cone or cone-rod dystrophy (COD1) (3, 4), atrophic macular degeneration (5), and RP in combination with impaired hearing and sinorespiratory infections (6). All RP3-associated missense mutations in RPGR have been identified in the N-terminal RCC1-homologous domain, and they disrupt the interaction with the C-terminal domain of RPGR-interacting protein 1 (RPGRIP1) (7,8). Mutations in RPGRIP1 lead to Leber congenital amaurosis (LCA), a genetically heterogeneous recessive disorder that is regarded to be the earliest and most severe form of all retinal dystrophies (9, 10). LCA accounts for at least 5% of all retinal dystrophies and is one of the main causes for blindness in children (11). RPGR and RPGRIP1 isoforms have been found to colocalize at the connecting cilia as well as the outer segments of rod and cone photoreceptors (12, 13). Differential localization among species has also been reported (12,14), and different isoforms of RPGRIP1 have been described resulting from splicing variation (7,14,15). The distinct partitioning of a subset of RPGRIP1 isoforms between the nuclear and cytoplasmic compartments combined with the differential and limited proteolysis of RPGRIP1 among retinal neurons, and the impact of LCA-linked mutations in RPGRIP1 in these processes, support the involvement of nucleocytoplasmatic signaling processes mediated by RPGRIP1 and its interacting partners in the pathogenesis of LCA, RP3, and allied diseases (15, 16). The pres...

mentioning

confidence: 71%

Interaction of nephrocystin-4 and RPGRIP1 is disrupted by nephronophthisis or Leber congenital amaurosis-associated mutations

Roepman

Letteboer²,

Arts³

et al. 2005

110

108

“…RPGR has a complex splicing pattern with multiple tissue-and cell-specific isoforms (52), is known to interact with a number of ciliary proteins (53,54), acts as a gunanine nucleotide exchange factor for small GTPase RAB8A (55), and may have a role in vertebrate development (56). Such complexity may account partially for the variability in disease phenotype.…”

Section: Discussionmentioning

confidence: 99%

Gene therapy rescues photoreceptor blindness in dogs and paves the way for treating human X-linked retinitis pigmentosa

Beltran

Cideciyan

Lewin

et al. 2012

236

260

Hereditary retinal blindness is caused by mutations in genes expressed in photoreceptors or retinal pigment epithelium. Gene therapy in mouse and dog models of a primary retinal pigment epithelium disease has already been translated to human clinical trials with encouraging results. Treatment for common primary photoreceptor blindness, however, has not yet moved from proof of concept to the clinic. We evaluated gene augmentation therapy in two blinding canine photoreceptor diseases that model the common X-linked form of retinitis pigmentosa caused by mutations in the retinitis pigmentosa GTPase regulator (RPGR) gene, which encodes a photoreceptor ciliary protein, and provide evidence that the therapy is effective. After subretinal injections of adeno-associated virus-2/5-vectored human RPGR with human IRBP or GRK1 promoters, in vivo imaging showed preserved photoreceptor nuclei and inner/ outer segments that were limited to treated areas. Both rod and cone photoreceptor function were greater in treated (three of four) than in control eyes. Histopathology indicated normal photoreceptor structure and reversal of opsin mislocalization in treated areas expressing human RPGR protein in rods and cones. Postreceptoral remodeling was also corrected: there was reversal of bipolar cell dendrite retraction evident with bipolar cell markers and preservation of outer plexiform layer thickness. Efficacy of gene therapy in these large animal models of X-linked retinitis pigmentosa provides a path for translation to human treatment.retina | retinal degeneration P hotoreceptors function cooperatively with the retinal pigment epithelium (RPE) to optimize photon catch and generate signals that are transmitted to higher vision centers and perceived as a visual image. Disruption of the visual process in the retinal photoreceptors can result in blindness. Genetic defects in the retina cause substantial numbers of sight-impairing disorders by a multitude of mechanisms (1, 2). These genetic diseases were classically considered incurable, but the past few years have witnessed a new era of retinal therapeutics in which successful gene therapy of an animal model of one blinding human disease (3) was followed by stepwise translation to the clinic. The RPE65 form of Leber congenital amaurosis, due to a biochemical blockade of the retinoid cycle in the RPE, was the first and remains the only blinding genetic disease to be successfully treated in humans (reviewed in ref. 4).The next level of challenge is to initiate treatment for the majority of blinding retinal disorders in which the genetic flaws are primarily in the photoreceptors. Successful targeting of therapeutic vectors to mutant photoreceptors would be required to restore function and preserve structure. Among photoreceptor dystrophies, the X-linked forms of retinitis pigmentosa (XLRP) are one of the most common causes of severe vision loss (5). More than 25 y ago, the genetic loci were identified (6), and discovery of the underlying gene defects followed (7,8). Mutations in the reti...

“…Selective entry through the nuclear pore requires cargo binding to a GTPbinding protein and guanine-nucleotide exchange involving a GEF to release cargo into the nucleus (27). We showed that RPGR ORF15 , a putative GEF (28), is a cilia/centrosomal protein that may regulate selective transport of cargo proteins from basal bodies distally along the ciliary axoneme in the photoreceptor connecting cilium (14,29). We now show that RPGR ORF15 is in complex with CEP290 in OSNs of both WT and rd16 mice, suggesting a possible role in regulating protein entry into the olfactory cilium.…”

Section: Discussionmentioning

confidence: 99%

Hypomorphic CEP290/NPHP6 mutations result in anosmia caused by the selective loss of G proteins in cilia of olfactory sensory neurons

McEwen¹,

Koenekoop

Khanna

et al. 2007

146

156

Cilia regulate diverse functions such as motility, fluid balance, and sensory perception. The cilia of olfactory sensory neurons (OSNs) compartmentalize the signaling proteins necessary for odor detection; however, little is known regarding the mechanisms of protein sorting/entry into olfactory cilia. Nephrocystins are a family of ciliary proteins likely involved in cargo sorting during transport from the basal body to the ciliary axoneme. In humans, loss-offunction of the cilia-centrosomal protein CEP290/NPHP6 is associated with Joubert and Meckel syndromes, whereas hypomorphic mutations result in Leber congenital amaurosis (LCA), a form of early-onset retinal dystrophy. Here, we report that CEP290 -LCA patients exhibit severely abnormal olfactory function. In a mouse model with hypomorphic mutations in CEP290 [retinal dystrophy-16 mice (rd16)], electro-olfactogram recordings revealed an anosmic phenotype analogous to that of CEP290 -LCA patients. Despite the loss of olfactory function, cilia of OSNs remained intact in the rd16 mice. As in wild type, CEP290 localized to dendritic knobs of rd16 OSNs, where it was in complex with ciliary transport proteins and the olfactory G proteins G olf and G␥13. Interestingly, we observed defective ciliary localization of G olf and G␥13 but not of G protein-coupled odorant receptors or other components of the odorant signaling pathway in the rd16 OSNs. Our data implicate distinct mechanisms for ciliary transport of olfactory signaling proteins, with CEP290 being a key mediator involved in G protein trafficking. The assessment of olfactory function can, therefore, serve as a useful diagnostic tool for genetic screening of certain syndromic ciliary diseases.nephrocystin ͉ olfaction