A photic visual cycle of rhodopsin regeneration is dependent on Rgr.

During visual excitation, rhodopsin undergoes photoactivation and bleaches to opsin and all-trans-retinal. To regenerate rhodopsin and maintain normal visual sensitivity, the all-trans isomer must be metabolized and reisomerized to produce the chromophore 11-cis-retinal in biochemical steps that constitute the visual cycle and involve the retinal pigment epithelium (RPE; refs. 3-8). A key step in the visual cycle is isomerization of an all-trans retinoid to 11-cis-retinol in the RPE (refs. 9-11). It could be that the retinochrome-like opsins, peropsin, or the retinal G protein-coupled receptor (RGR) opsin12-16 are isomerases in the RPE. In contrast to visual pigments, RGR is bound predominantly to endogenous all-trans-retinal, and irradiation of RGR in vitro results in stereospecific conversion of the bound all-trans isomer to 11-cis-retinal. Here we show that RGR is involved in the formation of 11-cis-retinal in mice and functions in a light-dependent pathway of the rod visual cycle. Mutations in the human gene encoding RGR are associated with retinitis pigmentosa.     

Mutations in NYX, encoding the leucine-rich proteoglycan nyctalopin, cause X-linked complete congenital stationary night blindness

During development, visual photoreceptors, bipolar cells and other neurons establish connections within the retina enabling the eye to process visual images over approximately 7 log units of illumination. Within the retina, cells that respond to light increment and light decrement are separated into ON- and OFF-pathways. Hereditary diseases are known to disturb these retinal pathways, causing either progressive degeneration or stationary deficits. Congenital stationary night blindness (CSNB) is a group of stable retinal disorders that are characterized by abnormal night vision. Genetic subtypes of CSNB have been defined and different disease actions have been postulated. The molecular bases have been elucidated in several subtypes, providing a better understanding of the disease mechanisms and developmental retinal neurobiology. Here we have studied 22 families with 'complete' X-linked CSNB (CSNB1; MIM 310500; ref. 4) in which affected males have night blindness, some photopic vision loss and a defect of the ON-pathway. We have found 14 different mutations, including 1 founder mutation in 7 families from the United States, in a novel candidate gene, NYX. NYX, which encodes a glycosylphosphatidyl (GPI)-anchored protein called nyctalopin, is a new and unique member of the small leucine-rich proteoglycan (SLRP) family. The role of other SLRP proteins suggests that mutant nyctalopin disrupts developing retinal interconnections involving the ON-bipolar cells, leading to the visual losses seen in patients with complete CSNB.     

Mutations in RDH12 encoding a photoreceptor cell retinol dehydrogenase cause childhood-onset severe retinal dystrophy

We identified three consanguineous Austrian kindreds with 15 members affected by autosomal recessive childhood-onset severe retinal dystrophy, a genetically heterogeneous group of disorders characterized by degeneration of the photoreceptor cells. A whole-genome scan by microarray analysis of single-nucleotide polymorphisms (ref. 2) identified a founder haplotype and defined a critical interval of 1.53 cM on chromosome 14q23.3-q24.1 that contains the gene associated with this form of retinal dystrophy. RDH12 maps in this region and encodes a retinol dehydrogenase proposed to function in the visual cycle. A homozygous 677A-->G transition (resulting in Y226C) in RDH12 was present in all affected family members studied, as well as in two Austrian individuals with sporadic retinal dystrophy. We identified additional mutations in RDH12 in 3 of 89 non-Austrian individuals with retinal dystrophy: a 5-nucleotide deletion (806delCCCTG) and the transition 565C-->T (resulting in Q189X), each in the homozygous state, and 146C-->T (resulting in T49M) and 184C-->T (resulting in R62X) in compound heterozygosity. When expressed in COS-7 cells, Cys226 and Met49 variants had diminished and aberrant activity, respectively, in interconverting isomers of retinol and retinal. The severe visual impairment of individuals with mutations in RDH12 is in marked contrast to the mild visual deficiency in individuals with fundus albipunctatus caused by mutations in RDH5, encoding another retinal dehydrogenase. Our studies show that RDH12 is associated with retinal dystrophy and encodes an enzyme with a unique, nonredundant role in the photoreceptor cells.     

Rescue of embryonic lethality in Mdm4-null mice by loss of Trp53 suggests a nonoverlapping pathway with MDM2 to regulate p53

The relationship between the neurosensory photoreceptors and the adjacent retinal pigment epithelium (RPE) controls not only normal retinal function, but also the pathogenesis of hereditary retinal degenerations. The molecular bases for both primary photoreceptor and RPE diseases that cause blindness have been identified. Gene therapy has been used successfully to slow degeneration in rodent models of primary photoreceptor diseases, but efficacy of gene therapy directed at photoreceptors and RPE in a large-animal model of human disease has not been reported. Here we study one of the most clinically severe retinal degenerations, Leber congenital amaurosis (LCA). LCA causes near total blindness in infancy and can result from mutations in RPE65 (LCA, type II; MIM 180069 and 204100). A naturally occurring animal model, the RPE65-/- dog, suffers from early and severe visual impairment similar to that seen in human LCA. We used a recombinant adeno-associated virus (AAV) carrying wild-type RPE65 (AAV-RPE65) to test the efficacy of gene therapy in this model. Our results indicate that visual function was restored in this large animal model of childhood blindness.     

Mutations in a novel retina-specific gene cause autosomal dominant retinitis pigmentosa.

Inherited retinal diseases are a common cause of visual impairment in children and young adults, often resulting in severe loss of vision in later life. The most frequent form of inherited retinopathy is retinitis pigmentosa (RP), with an approximate incidence of 1 in 3,500 individuals worldwide. RP is characterized by night blindness and progressive degeneration of the midperipheral retina, accompanied by bone spicule-like pigmentary deposits and a reduced or absent electroretinogram (ERG). The disease process culminates in severe reduction of visual fields or blindness. RP is genetically heterogeneous, with autosomal dominant, autosomal recessive and X-linked forms. Here we have identified two mutations in a novel retina-specific gene from chromosome 8q that cause the RP1 form of autosomal dominant RP in three unrelated families. The protein encoded by this gene is 2,156 amino acids and its function is currently unknown, although the amino terminus has similarity to that of the doublecortin protein, whose gene (DCX) has been implicated in lissencephaly in humans. Two families have a nonsense mutation in codon 677 of this gene (Arg677stop), whereas the third family has a nonsense mutation in codon 679 (Gln679stop). In one family, two individuals homozygous for the mutant gene have more severe retinal disease compared with heterozygotes.     

New views on RPE65 deficiency: the rod system is the source of vision in a mouse model of Leber congenital amaurosis

Leber congenital amaurosis (LCA) is the most serious form of the autosomal recessive childhood-onset retinal dystrophies. Mutations in the gene encoding RPE65, a protein vital for regeneration of the visual pigment rhodopsin in the retinal pigment epithelium, account for 10-15% of LCA cases. Whereas previous studies of RPE65 deficiency in both animal models and patients attributed remaining visual function to cones, we show here that light-evoked retinal responses in fact originate from rods. For this purpose, we selectively impaired either rod or cone function in Rpe65-/- mice by generating double- mutant mice with models of pure cone function (rhodopsin-deficient mice; Rho-/-) and pure rod function (cyclic nucleotide-gated channel alpha3-deficient mice; Cnga3-/-). The electroretinograms (ERGs) of Rpe65-/- and Rpe65-/-Cnga3-/- mice were almost identical, whereas there was no assessable response in Rpe65-/-Rho-/- mice. Thus, we conclude that the rod system is the source of vision in RPE65 deficiency. Furthermore, we found that lack of RPE65 enables rods to mimic cone function by responding under normally cone-isolating lighting conditions. We propose as a mechanism decreased rod sensitivity due to a reduction in rhodopsin content to less than 1%. In general, the dissection of pathophysiological processes in animal models through the introduction of additional, selective mutations is a promising concept in functional genetics.     

Mutations in ABCC6 cause pseudoxanthoma elasticum

Pseudoxanthoma elasticum (PXE) is a heritable disorder of the connective tissue. PXE patients frequently experience visual field loss and skin lesions, and occasionally cardiovascular complications. Histopathological findings reveal calcification of the elastic fibres and abnormalities of the collagen fibrils. Most PXE patients are sporadic, but autosomal recessive and dominant inheritance are also observed. We previously localized the PXE gene to chromosome 16p13.1 (refs 8,9) and constructed a physical map. Here we describe homozygosity mapping in five PXE families and the detection of deletions or mutations in ABCC6 (formerly MRP6) associated with all genetic forms of PXE in seven patients or families.     

Nuclear gene OPA1, encoding a mitochondrial dynamin-related protein, is mutated in dominant optic atrophy

Optic atrophy type 1 (OPA1, MIM 165500) is a dominantly inherited optic neuropathy occurring in 1 in 50,000 individuals that features progressive loss in visual acuity leading, in many cases, to legal blindness. Phenotypic variations and loss of retinal ganglion cells, as found in Leber hereditary optic neuropathy (LHON), have suggested possible mitochondrial impairment. The OPA1 gene has been localized to 3q28-q29 (refs 13-19). We describe here a nuclear gene, OPA1, that maps within the candidate region and encodes a dynamin-related protein localized to mitochondria. We found four different OPA1 mutations, including frameshift and missense mutations, to segregate with the disease, demonstrating a role for mitochondria in retinal ganglion cell pathophysiology.     

A 5-bp deletion in ELOVL4 is associated with two related forms of autosomal dominant macular dystrophy

Stargardt-like macular dystrophy (STGD3, MIM 600110) and autosomal dominant macular dystrophy (adMD) are inherited forms of macular degeneration characterized by decreased visual acuity, macular atrophy and extensive fundus flecks. Genetic mapping data suggest that mutations in a single gene may be responsible for both conditions, already known to bear clinical resemblance. Here we limit the minimum genetic region for STGD3 and adMD to a 0.6-cM interval by recombination breakpoint mapping and identify a single 5-bp deletion within the protein-coding region of a new retinal photoreceptor-specific gene, ELOVL4, in all affected members of STGD3 and adMD families. Bioinformatic analysis of ELOVL4 revealed that it has homology to a group of yeast proteins that function in the biosynthesis of very long chain fatty acids. Our results are therefore the first to implicate the biosynthesis of fatty acids in the pathogenesis of inherited macular degeneration.     

Blindness and auditory impairment caused by loss of the sodium bicarbonate cotransporter NBC3

Normal sensory transduction requires the efficient disposal of acid (H+) generated by neuronal and sensory receptor activity. Multiple highly sensitive transport mechanisms have evolved in prokaryotic and eukaryotic organisms to maintain acidity within strict limits. It is currently assumed that the multiplicity of these processes provides a biological robustness. Here we report that the visual and auditory systems have a specific requirement for H+ disposal mediated by the sodium bicarbonate cotransporter NBC3 (refs. 7,8). Mice lacking NBC3 develop blindness and auditory impairment because of degeneration of sensory receptors in the eye and inner ear as in Usher syndrome. Our results indicate that in certain sensory organs, in which the requirement to transduce specific environmental signals with speed, sensitivity and reliability is paramount, the choice of the H+ disposal mechanism used is limited.     

Mutations in the gene encoding lecithin retinol acyltransferase are associated with early-onset severe retinal dystrophy

The chromophore of the visual pigments, 11-cis retinal, is derived from vitamin A (all-trans retinol) through a series of reactions that take place in retinal pigment epithelium (RPE); (ref. 1). The first of these reactions is catalyzed by lecithin retinol acyltransferase (LRAT); (ref. 2). We screened 267 retinal dystrophy patients for mutations in LRAT and identified disease-associated mutations (S175R and 396delAA) in three individuals with severe, early-onset disease. We showed that the S175R mutant has no acyltransferase activity in transfected COS-7 cells. Our findings highlight the importance of genetic defects in vitamin A metabolism as causes of retinal dystrophies and extend prospects for retinoid replacement therapy in this group of diseases.     

Haploinsufficiency of ATP1A2 encoding the Na+/K+ pump alpha2 subunit associated with familial hemiplegic migraine type 2

Headache attacks and autonomic dysfunctions characterize migraine, a very common, disabling disorder with a prevalence of 12% in the general population of Western countries. About 20% of individuals affected with migraine experience aura, a visual or sensory-motor neurological dysfunction that usually precedes or accompanies the headache. Although the mode of transmission is controversial, population-based and twin studies have implicated genetic factors, especially in migraine with aura. Familial hemiplegic migraine is a hereditary form of migraine characterized by aura and some hemiparesis. Here we show that mutations in the gene ATP1A2 that encodes the alpha2 subunit of the Na+/K+ pump are associated with familial hemiplegic migraine type 2 (FHM2) linked to chromosome 1q23 (OMIM 602481). Functional data indicate that the putative pathogenetic mechanism is triggered by a loss of function of a single allele of ATP1A2. This is the first report associating mutations of Na+K+ pump subunits to genetic diseases.     

Identification and characterization of rod-derived cone viability factor

Retinitis pigmentosa is an untreatable, inherited retinal disease that leads to blindness. The disease initiates with the loss of night vision due to rod photoreceptor degeneration, followed by irreversible, progressive loss of cone photoreceptor. Cone loss is responsible for the main visual handicap, as cones are essential for day and high-acuity vision. Their loss is indirect, as most genes associated with retinitis pigmentosa are not expressed by these cells. We previously showed that factors secreted from rods are essential for cone viability. Here we identified one such trophic factor by expression cloning and named it rod-derived cone viability factor (RdCVF). RdCVF is a truncated thioredoxin-like protein specifically expressed by photoreceptors. The identification of this protein offers new treatment possibilities for retinitis pigmentosa.     

A genetic approach to understanding auditory function

Little is known of the molecular basis of normal auditory function. In contrast to the visual or olfactory senses, in which reasonable amounts of sensory tissue can be gathered, the auditory system has proven difficult to access through biochemical routes, mainly because such small amounts of tissue are available for analysis. Key molecules, such as the transduction channel, may be present in only a few tens of copies per sensory hair cell, compounding the difficulty. Moreover, fundamental differences in the mechanism of stimulation and, most importantly, the speed of response of audition compared with other senses means that we have no well-understood models to provide good candidate molecules for investigation. For these reasons, a genetic approach is useful for identifying the key components of auditory transduction, as it makes no assumptions about the nature or expression level of molecules essential for hearing. We review here some of the major advances in our understanding of auditory function resulting from the recent rapid progress in identification of genes involved in deafness.     

A common CFH haplotype, with deletion of CFHR1 and CFHR3, is associated with lower risk of age-related macular degeneration

Age-related macular degeneration (AMD; OMIM #603075) is the most frequent cause of visual impairment in the elderly population, with severe disease affecting nearly 10% of individuals of European descent over the age of 75 years. It is a complex disease in which genetic and environmental factors contribute to susceptibility. Complement factor H (CFH) has recently been identified as a major AMD susceptibility gene, and the Y402H polymorphism has been proposed as the likely causative factor. We genotyped polymorphisms spanning the cluster of CFH and five CFH-related genes on chromosome 1q23 in 173 individuals with severe neovascular AMD and 170 elderly controls with no signs of AMD. Detailed analysis showed a common haplotype associated with decreased risk of AMD that was present on 20% of chromosomes of controls and 8% of chromosomes of individuals with AMD. We found that this haplotype carried a deletion of CFHR1 and CFHR3, and the proteins encoded by these genes were absent in serum of homozygotes. The protective effect of the deletion haplotype cannot be attributed to linkage disequilibrium with Y402H and was replicated in an independent sample.     

Genome-wide association study identifies susceptibility loci for open angle glaucoma at TMCO1 and CDKN2B-AS1

We report a genome-wide association study for open-angle glaucoma (OAG) blindness using a discovery cohort of 590 individuals with severe visual field loss (cases) and 3,956 controls. We identified associated loci at TMCO1 (rs4656461[G] odds ratio (OR) = 1.68, P = 6.1 × 10(-10)) and CDKN2B-AS1 (rs4977756[A] OR = 1.50, P = 4.7 × 10(-9)). We replicated these associations in an independent cohort of cases with advanced OAG (rs4656461 P = 0.010; rs4977756 P = 0.042) and two additional cohorts of less severe OAG (rs4656461 combined discovery and replication P = 6.00 × 10(-14), OR = 1.51, 95% CI 1.35-1.68; rs4977756 combined P = 1.35 × 10(-14), OR = 1.39, 95% CI 1.28-1.51). We show retinal expression of genes at both loci in human ocular tissues. We also show that CDKN2A and CDKN2B are upregulated in the retina of a rat model of glaucoma.     

Genetic basis of total colourblindness among the Pingelapese islanders

Complete achromatopsia is a rare, autosomal recessive disorder characterized by photophobia, low visual acuity, nystagmus and a total inability to distinguish colours. In this disease, cone photoreceptors, the retinal sensory neurons mediating colour vision, seem viable but fail to generate an electrical response to light. Achromatopsia, or rod monochromatism, was first mapped to 2p11-2q12 (MIM 216900; ref. 3), where it is associated with missense mutations in CNGA3 (ref. 4). CNGA3 encodes the alpha-subunit of the cone cyclic nucleotide-gated cation channel, which generates the light-evoked electrical responses of cone photoreceptors. A second locus at 8q21-q22 has been identified among the Pingelapese islanders of Micronesia, who have a high incidence of recessive achromatopsia (MIM 262300). Here we narrow the achromatopsia locus to 1.4 cM and show that Pingelapese achromatopsia segregates with a missense mutation at a highly conserved site in CNGB3, a new gene that encodes the beta-subunit of the cone cyclic nucleotide-gated cation channel. Two independent frameshift deletions establish that achromatopsia is the null phenotype of CNGB3. Combined with earlier findings, our results demonstrate that both alpha- and beta-subunits of the cGMP-gated channel are essential for phototransduction in all three classes of cones.