Dominant Congenital Stationary Night Blindness Research Articles

Identification of a novel large multigene deletion and a frameshift indel in PDE6B as the underlying cause of early-onset recessive rod-cone degeneration.

A family, with two affected identical twins with early-onset recessive inherited retinal degeneration, was analyzed to determine the underlying genetic cause of pathology. Exome sequencing revealed a rare and previously reported causative variant (c.1923_1969delinsTCTGGG; p.Asn643Glyfs*29) in the PDE6B gene in the affected twins and their unaffected father. Further investigation, using genome sequencing, identified a novel ∼7.5-kb deletion (Chr 4:670,405-677,862del) encompassing the ATP5ME gene, part of the 5' UTR of MYL5, and a 378-bp (Chr 4:670,405-670,782) region from the 3' UTR of PDE6B in the affected twins and their unaffected mother. Both variants segregated with disease in the family. Analysis of the relative expression of PDE6B, in peripheral blood cells, also revealed a significantly lower level of PDE6B transcript in affected siblings compared to a normal control. PDE6B is associated with recessive rod-cone degeneration and autosomal dominant congenital stationary night blindness. Ophthalmic evaluation of these patients showed night blindness, fundus abnormalities, and peripheral vision loss, which are consistent with PDE6B-associated recessive retinal degeneration. These findings suggest that the loss of PDE6B transcript resulting from the compound heterozygous pathogenic variants is the underlying cause of recessive rod-cone degeneration in the study family.

Coexistence of GNAT1 and ABCA4 variants associated with Nougaret-type congenital stationary night blindness and childhood-onset cone-rod dystrophy.

A single variant (p.G38D) in the GNAT1 gene, encoding the rod-specific transducin α-subunit in phototransduction, has been reported only in one French family with Nougaret-type autosomal dominant congenital stationary night blindness (CSNB). We identified a Japanese family with Nougaret-type CSNB and cone-rod dystrophy (CORD). Five patients with CSNB and two patients with childhood-onset CORD were recruited. We performed a comprehensive ophthalmic examination including electroretinography (ERG). Disease-causing variants were identified by whole exome sequencing, with candidates confirmed by Sanger sequencing in nine family members. The GNAT1 variant (p.G38D) was identified in all four CSNB patients, whereas the two CORD patients carried biallelic truncated known ABCA4 variants as well as the GNAT1 variant. Clinically, no remarkable findings were observed in fuduscopy, fundus autofluorescence, or optical coherence tomography images from the CSNB patients. No response was detectable by rod ERG. The a-waves of standard and bright flash ERG were delayed and broadened rather than biphasic, and b/a-wave amplitude ratio was negative. Cone and 30-Hz flicker responses were normal, and overall, the ERG findings were compatible with previous descriptions of Nougaret-type CSNB. ERG of the CORD patients with macular atrophy showed non-recordable rod response and severely decreased standard flash, cone and 30-Hz flicker responses. This is the second report of a Nougaret-type CSNB family with the GNAT1 variant. Our novel findings suggest that coexistence of the GNAT1 and biallelic ABCA4 variants is associated with an overlapping phenotype with both Nougaret-type CSNB and CORD.

The molecular and cellular basis of rhodopsin retinitis pigmentosa reveals potential strategies for therapy.

Inherited mutations in the rod visual pigment, rhodopsin, cause the degenerative blinding condition, retinitis pigmentosa (RP). Over 150 different mutations in rhodopsin have been identified and, collectively, they are the most common cause of autosomal dominant RP (adRP). Mutations in rhodopsin are also associated with dominant congenital stationary night blindness (adCSNB) and, less frequently, recessive RP (arRP). Recessive RP is usually associated with loss of rhodopsin function, whereas the dominant conditions are a consequence of gain of function and/or dominant negative activity. The in-depth characterisation of many rhodopsin mutations has revealed that there are distinct consequences on the protein structure and function associated with different mutations. Here we categorise rhodopsin mutations into seven discrete classes; with defects ranging from misfolding and disruption of proteostasis, through mislocalisation and disrupted intracellular traffic to instability and altered function. Rhodopsin adRP offers a unique paradigm to understand how disturbances in photoreceptor homeostasis can lead to neuronal cell death. Furthermore, a wide range of therapies have been tested in rhodopsin RP, from gene therapy and gene editing to pharmacological interventions. The understanding of the disease mechanisms associated with rhodopsin RP and the development of targeted therapies offer the potential of treatment for this currently untreatable neurodegeneration.

Mechanisms of mutant PDE6 proteins underlying retinal diseases

Mutations in PDE6 genes encoding the effector enzymes in rods and cones underlie severe retinal diseases including retinitis pigmentosa (RP), autosomal dominant congenital stationary night blindness (adCSNB), and achromatopsia (ACHM). Here we examined a spectrum of pathogenic missense mutations in PDE6 using the system based on co-expression of cone PDE6C with its specialized chaperone AIPL1 and the regulatory Pγ subunit as a potent co-chaperone. We uncovered two mechanisms of PDE6C mutations underlying ACHM: (a) folding defects leading to expression of catalytically inactive proteins and (b) markedly diminished ability of Pγ to co-chaperone mutant PDE6C proteins thereby dramatically reducing the levels of functional enzyme. The mechanism of the Rambusch adCSNB associated with the H258N substitution in PDE6B was probed through the analysis of the model mutant PDE6C-H262N. We identified two interrelated deficits of PDE6C-H262N: disruption of the inhibitory interaction of Pγ with mutant PDE6C that markedly reduced the ability of Pγ to augment the enzyme folding. Thus, we conclude that the Rambusch adCSNB is triggered by low levels of the constitutively active PDE6. Finally, we examined PDE6C-L858V, which models PDE6B-L854V, an RP-linked mutation that alters the protein isoprenyl modification. This analysis suggests that the type of prenyl modifications does not impact the folding of PDE6, but it modulates the enzyme affinity for its trafficking partner PDE6D. Hence, the pathogenicity of PDE6B-L854V likely arises from its trafficking deficiency. Taken together, our results demonstrate the effectiveness of the PDE6C expression system to evaluate pathogenicity and elucidate the mechanisms of PDE6 mutations in retinal diseases.

Characterization of Ribozymes Targeting a Congenital Night Blindness Mutation in Rhodopsin Mutation.

The G90D mutation in the rhodopsin gene leads to autosomal dominant congenital stationary night blindness (CSNB) in patients. This occurs because the G90D mutant protein cannot efficiently bind chromophore and is constitutively active. To combat this mutation, we designed and characterized two different hammerhead ribozymes to cleave G90D transcript. In vitro testing showed that the G90D1 ribozyme efficiently and specifically cleaved the mutant transcript while G90D2 cleaved both WT and mutant transcript. AAV-mediated delivery of G90D1 under the control of the mouse opsin promoter (MOP500) to G90D transgenic eyes showed that the ribozyme partially retarded the functional degeneration (as measured by electroretinography [ERG]) associated with this mutation. These results suggest that with additional optimization, ribozymes may be a useful part of the gene therapy knockdown strategy for dominant retinal disease.

A truncated form of rod photoreceptor PDE6 β-subunit causes autosomal dominant congenital stationary night blindness by interfering with the inhibitory activity of the γ-subunit.

Autosomal dominant congenital stationary night blindness (adCSNB) is caused by mutations in three genes of the rod phototransduction cascade, rhodopsin (RHO), transducin α-subunit (GNAT1), and cGMP phosphodiesterase type 6 β-subunit (PDE6B). In most cases, the constitutive activation of the phototransduction cascade is a prerequisite to cause adCSNB. The unique adCSNB-associated PDE6B mutation found in the Rambusch pedigree, the substitution p.His258Asn, leads to rod photoreceptors desensitization. Here, we report a three-generation French family with adCSNB harboring a novel PDE6B mutation, the duplication, c.928-9_940dup resulting in a tyrosine to cysteine substitution at codon 314, a frameshift, and a premature termination (p.Tyr314Cysfs*50). To understand the mechanism of the PDE6β1-314fs*50 mutant, we examined the properties of its PDE6-specific portion, PDE6β1-313. We found that PDE6β1-313 maintains the ability to bind noncatalytic cGMP and the inhibitory γ-subunit (Pγ), and interferes with the inhibition of normal PDE6αβ catalytic subunits by Pγ. Moreover, both truncated forms of the PDE6β protein, PDE6β1-313 and PDE6β1-314fs*50 expressed in rods of transgenic X. laevis are targeted to the phototransduction compartment. We hypothesize that in affected family members the p.Tyr314Cysfs*50 change results in the production of the truncated protein, which binds Pγ and causes constitutive activation of the phototransduction thus leading to the absence of rod adaptation.

Mutation analysis of pathogenic genes in a Henan family affected with congenital stationary night blindness

To detect genetic mutations associated with autosomal dominant congenital stationary night blindness (ADCSNB) in a family from Henan province. Genomic DNA was extracted from peripheral blood samples of 14 family members. Based on 3 genes reported previously, PCR primers were designed and corresponding exons containing the mutation sites were amplified with PCR. PCR products were purified and directly sequenced. A c.281C>T heterozygous missense mutation was detected in RHO gene in all of the patients. This mutation can cause a change of the protein structure (p.Thr94Ile). The same mutation was not detected in normal individuals from the family and 50 normal controls. A c.281C>T mutation in RHO gene is responsible for the onset of ADCSNB in this Chinese family and results in symptoms of night blindness.

Molecular Mechanisms of Disease for Mutations at Gly-90 in Rhodopsin

Two different mutations at Gly-90 in the second transmembrane helix of the photoreceptor protein rhodopsin have been proposed to lead to different phenotypes. G90D has been classically associated with congenital night blindness, whereas the newly reported G90V substitution was linked to a retinitis pigmentosa phenotype. Here, we used Val/Asp replacements of the native Gly at position 90 to unravel the structure/function divergences caused by these mutations and the potential molecular mechanisms of inherited retinal disease. The G90V and G90D mutants have a similar conformation around the Schiff base linkage region in the dark state and same regeneration kinetics with 11-cis-retinal, but G90V has dramatically reduced thermal stability when compared with the G90D mutant rhodopsin. The G90V mutant also shows, like G90D, an altered photobleaching pattern and capacity to activate Gt in the opsin state. Furthermore, the regeneration of the G90V mutant with 9-cis-retinal was improved, achieving the same A(280)/A(500) as wild type isorhodopsin. Hydroxylamine resistance was also recovered, indicating a compact structure around the Schiff base linkage, and the thermal stability was substantially improved when compared with the 11-cis-regenerated mutant. These results support the role of thermal instability and/or abnormal photoproduct formation in eliciting a retinitis pigmentosa phenotype. The improved stability and more compact structure of the G90V mutant when it was regenerated with 9-cis-retinal brings about the possibility that this isomer or other modified retinoid analogues might be used in potential treatment strategies for mutants showing the same structural features.

Identification and Functional Characterization of a Novel Rhodopsin Mutation Associated with Autosomal Dominant CSNB

Mutations in RHO, PDE6B, and GNAT1 can lead to autosomal dominant congenital stationary night blindness (adCSNB). The study was conducted to identify the genetic defect in a large Swiss family affected with adCSNB and to investigate the pathogenic mechanism of the mutation. Two affected cousins of a large Swiss family were examined clinically by standard methods: funduscopy, EOG, ERG, and dark adaptometry. Twelve family members were screened for mutations in RHO. The ability of mutant rhodopsin to activate transducin constitutively was monitored by measuring the catalytic exchange of bound GDP for radiolabeled [(35)S]GTPgammaS in transducin. A novel mutation was identified in RHO (c.884C>T, p.Ala295Val) in patients with adCSNB. They had full vision under photopic conditions, showed no fundus abnormalities, revealed EOG results in the normal range, but presented night blindness with an altered scotopic ERG. In the presence of 11-cis retinal, the mutant rhodopsin is inactive, similar to wild-type, responding only when exposed to light. However, in the absence of 11-cis-retinal, unlike wild-type opsin, the mutant opsin constitutively activates transducin. The study adds a fourth rhodopsin mutation associated with CSNB. Although the phenotype of autosomal dominant CSNB may vary slightly in patients showing mutations in RHO, PDE6B, or GNAT1, the disease course seems to be stationary with only scotopic vision being affected. The data indicate that the mutant opsin activates transducin constitutively, which is a consistent and common feature of all four CSNB-associated rhodopsin mutations reported to date.

P.Gln200Glu, a putative constitutively active mutant of rod α-transducin (GNAT1) in autosomal dominant congenital stationary night blindness

Congenital stationary night blindness (CSNB) is a non-progressive Mendelian condition resulting from a functional defect in rod photoreceptors. A small number of unique missense mutations in the genes encoding various members of the rod phototransduction cascade, e.g. rhodopsin (RHO), cGMP phosphodiesterase beta-subunit (PDE6B), and transducin alpha-subunit (GNAT1) have been reported to cause autosomal dominant (ad) CSNB. While the RHO and PDE6B mutations result in constitutively active proteins, the only known adCSNB-associated GNAT1 change (p.Gly38Asp) produces an alpha-transducin that is unable to activate its downstream effector molecule in vitro. In a multigeneration Danish family with adCSNB, we identified a novel heterozygous C to G transversion (c.598C>G) in exon 6 of GNAT1 that should result in a p.Gln200Glu substitution in the evolutionarily highly conserved Switch 2 region of alpha-transducin, a domain that has an important role in binding and hydrolyzing GTP. Computer modeling based on the known crystal structure of transducin suggests that the p.Gln200Glu mutant exhibits impaired GTPase activity, and thereby leads to constitutive activation of phototransduction. This assumption is in line with our results of trypsin protection assays as well as previously published biochemical data on mutants of this glutamine in the GTPase active site of alpha-transducin following in vitro expression, and observations that inappropriately activating mutants of various members of the rod phototransduction cascade represent one of the major molecular causes of adCSNB.

Exclusion of the association of five known mutations with congenital stationary nyctalopia in a large Chinese family

To detect gene mutations associated with autosomal dominant congenital stationary night blindness(ADCSNB) in a large Chinese family. Genomic DNAs were extracted from peripheral blood samples of 16 affected and 14 unaffected family members. According to 5 missense mutations in 3 genes reported previously, 4 pairs of primers were designed and corresponding exons containing the five mutation sites were amplified by polymerase chain reaction. Amplified products were purified and sequenced by MegaBACE1000 capillary array electrophoresis DNA sequencer. Full field electroretinogram (ERG, ISCEV) of patients was recorded and analyzed by Roland Consult System. Dark-adapted ERG showed a-wave was normal, but b-wave of the patients was markedly decreased. None of the five missense mutations were detected in 16 affected and 14 unaffected family members. The molecular pathogenesis of ADCSNB in this family does not involve point mutations or deletions of these five sites, which indicates the heterogeneity of ADCSNB.

A novel mutation within the rhodopsin gene (Thr-94-Ile) causing autosomal dominant congenital stationary night blindness.

More than 100 mutations within the rhodopsin gene have been found to be responsible for some forms of retinitis pigmentosa, a progressive retinal degeneration characterized by night blindness and subsequent disturbance of day vision that may eventually result in total blindness. Congenital stationary night blindness (CSNB) is an uncommon inherited retinal dysfunction in which patients complain of night vision difficulties of a nonprogressive nature only and in which generally there is no involvement of day vision. We report the results of molecular genetic analysis of an Irish family segregating an autosomal dominant form of CSNB in which a previously unreported threonine-to-isoleucine substitution at codon 94 in the rhodopsin gene was found to segregate with the disease. Computer modeling suggests that constitutive activation of transducin by the altered rhodopsin protein may be a mechanism for disease causation in this family. Only two mutations within the rhodopsin gene have been previously reported in patients with congenital stationary night blindness, constitutive activation also having been proposed as a possible disease mechanism.

Correlation of phenotype with genotype in inherited retinal degeneration

AbstractDiseases causing inherited retinal degeneration in humans, such as retinitis pigmentosa and macular dystrophy, are genetically heterogeneous and clinically diverse. More than 40 genes causing retinal degeneration have been mapped to specific chromosomal sites; of these, at least 10 have been cloned and characterized. Mutations in two proteins, rhodopsin and peripherin/RDS, account for approximately 35% of all cases of autosomal dominant retinitis pigmentosa and a lesser fraction of other retinal conditions. This target article reviews the genes and mutations causing retinal degeneration and proposes mechanisms whereby specific mutations lead to particular clinical consequences, that is, the relationship between genotype and phenotype. Retinitis pigmentosa and macular dystrophy are genetically heterogeneous diseases that cause retinal degeneration in humans and often result in severe visual impairment or blindness. Although many of the genes causing these diseases have not been identified, three photoreceptor-specific proteins have been implicated: rhodopsin, peripherin/RDS, and the P-subunit of rod phosphodiesterase. Mutations in the genes for these three proteins can cause either dominant retinitis pigmentosa, recessive retinitis pigmentosa, dominant congenital stationary night blindness, or dominant macular degeneration. Why this multiplicity of clinical phenotypes? Our target article summarizes the genetic and biochemical background to this question and proposes a number of possible explanations. Discussion focuses mainly on 73 distinct disease-causing mutations of rhodopsin. We feel that rhodopsin and other photoreceptor proteins can serve as model systems for unraveling the connection between genotype and phenotype, not only for inherited retinal diseases but for other degenerative disorders as well.

Heterozygous missense mutation in the rod cGMP phosphodiesterase β–subunit gene in autosomal dominant stationary night blindness

The locus for autosomal dominant congenital stationary night blindness (adCSNB) has recently been assigned to distal chromosome 4p by linkage analysis in a large Danish family. Within the candidate gene encoding the beta-subunit of rod photoreceptor cGMP-specific phosphodiesterase (beta PDE), we have identified a heterozygous C to A transversion in exon 4, predicting a His258Asp change in the polypeptide. We found a perfect cosegregation (Zmax = 22.6 at theta = 0.00) of this mutation with the disease phenotype suggesting that this missense mutation is responsible for the disease in this pedigree. Homozygous nonsense mutations in the beta PDE gene have been found recently in patients with autosomal recessive retinitis pigmentosa, a common hereditary photoreceptor dystrophy.

Gene for autosomal dominant congenital stationary night blindness maps to the same region as the gene for the beta-subunit of the rod photoreceptor cGMP phosphodiesterase (PDEB) in chromosome 4p16.3.

We studied a large multigeneration Danish family with autosomal dominant congenital stationary night blindness. Both electrophysiological and psychophysical findings in affected family members were identical to those reported in patients from the 'Nougaret family'. The disease locus in the Danish family has now been mapped by demonstrating close linkage without recombination (Q = 0.00 at Zmax = 14.4) to the locus for alpha-L-iduronidase assigned to chromosome 4p16.3. Interestingly the gene for the beta-subunit of the rod photoreceptor cGMP-specific phosphodiesterase maps to the very same chromosomal region.

Autosomal Dominant Congenital Stationary Night Blindness and Normal Fundus With an Electronegative Electroretinogram

We studied three members of three successive generations of a family with autosomal dominant congenital stationary night blindness and normal fundi. Psychophysical studies on two members showed normal final cone thresholds and mildly increased rod thresholds. Full-field electroretinograms on all three members showed normal photopic b-wave amplitudes and implicit times. Under scotopic conditions, the rod response was absent, and with a bright flash stimulus, there was a normal a-wave with no b-wave. This electronegative dark-adapted electroretinogram resembled the Schubert-Bornschein type seen in congenital stationary blindness, which has been seen only in autosomal and X-linked recessive pedigrees.