Ajoy Vincent

Mutations in the unfolded protein response regulator ATF6 cause the cone dysfunction disorder achromatopsia

Achromatopsia (ACHM) is an autosomal recessive disorder characterized by color blindness, photophobia, nystagmus and severely reduced visual acuity. Using homozygosity mapping and whole-exome and candidate gene sequencing, we identified ten families carrying six homozygous and two compound-heterozygous mutations in the ATF6 gene (encoding activating transcription factor 6A), a key regulator of the unfolded protein response (UPR) and cellular endoplasmic reticulum (ER) homeostasis. Patients had evidence of foveal hypoplasia and disruption of the cone photoreceptor layer. The ACHM-associated ATF6 mutations attenuate ATF6 transcriptional activity in response to ER stress. Atf6−/− mice have normal retinal morphology and function at a young age but develop rod and cone dysfunction with increasing age. This new ACHM-related gene suggests a crucial and unexpected role for ATF6A in human foveal development and cone function and adds to the list of genes that, despite ubiquitous expression, when mutated can result in an isolated retinal photoreceptor phenotype.

BBS genotype–phenotype assessment of a multiethnic patient cohort calls for a revision of the disease definition†

Bardet‐Biedl syndrome (BBS) is a ciliopathy characterized by retinal degeneration, obesity, polydactyly, renal abnormalities, and cognitive impairment for which 15 causative genes have been identified. Here we present the results of a mutational analysis of our multiethnic cohort of 83 families (105 cases); 75.9% of them have their mutations identified including 26 novel changes. Comprehensive phenotyping of these patients demonstrate that the spectrum of clinical features is greater than expected and overlapped with the features of other ciliopathies; specifically Alström and McKusick‐Kauffman syndromes. Hum Mutat 32:1–10, 2011.

A Phenotype-Genotype Correlation Study of X-Linked Retinoschisis

PURPOSE To compare the clinical phenotype and detailed electroretinographic parameters in X-linked retinoschisis (XLRS). DESIGN Retrospective, comparative study. PARTICIPANTS Fifty-seven patients (aged 1-67 years) with molecularly confirmed XLRS were clinically ascertained. METHODS Pattern electroretinography (PERG) and full-field electroretinography (ERG), incorporating international standard recordings, were performed in 44 cases. Thirteen patients, mostly pediatric, were tested using a simplified ERG protocol. On-Off and S-cone ERGs were performed in most adults. Fundus autofluorescence (FAF) imaging and optical coherence tomography (OCT) were available in 17 and 21 cases, respectively. MAIN OUTCOME MEASURES The clinical and electrophysiologic data associated with different types of mutation in the RS1 gene. RESULTS Forty-three patients had missense changes (group A), and 14 patients had nonsense, splice-site, or frame-shifting mutations in the RS1 gene (group B). The mean best-corrected visual acuity was better in group A than in group B (0.34 and 0.21, respectively). Fundus examination revealed foveal schisis in approximately half of both groups. The bright-flash dark-adapted (DA) ERG (11.0 candela.sec.m(-2)) waveform was electronegative in 62% of group A eyes and 100% of group B eyes. The photopic 30-Hz flicker ERG was delayed in all group B eyes and all except 6 group A eyes. On-Off ERG b-waves were subnormal in 39% of group A and 89% of group B eyes; d-waves were delayed in 14 eyes (group A = 10, group B = 4). S-cone ERGs were abnormal in 50% of both groups. The PERG was abnormal in 88% of group A and 100% of group B eyes. A spoke-wheel pattern of high and low intensity was the most common FAF abnormality observed. The OCT showed intraretinal schitic cavities in the majority of eyes. CONCLUSIONS There is profound phenotypic variability in patients with XLRS. Most patients have DA bright-flash ERGs with a low b:a ratio in keeping with inner retinal dysfunction. Generalized cone system dysfunction is common and associated with an abnormal On-response and less frequent additional Off-response involvement. Nonsense, splice-site, or frame-shifting mutations in RS1 consistently caused electronegative bright-flash ERG, delayed flicker response, and abnormal PERG; missense mutations result in a wider range of ERG abnormalities.

PATHOGNOMONIC (DIAGNOSTIC) ERGs A Review and Update

Purpose: To review three inherited retinal disorders associated with diagnostic or pathognomonic electroretinogram (ERG) abnormalities: cone dystrophy with supernormal rod ERG (KCNV2), enhanced S-cone syndrome (NR2E3), and bradyopsia (RGS9/R9AP). Methods: A review of clinical details, genetic basis, and electrophysiological features in these disorders and a brief summary of the standard and nonstandard ERG techniques required to identify the disorders. Results: The electrophysiological features in each of these three disorders are pathognomonic such that the responsible gene can be specified. The results from nonstandard electrophysiological testing in excess of international standards are necessary to describe the pathognomonic changes in cone dystrophy with supernormal rod ERG and bradyopsia. The clinical phenotype in the disorders can be variable. Mutations in NR2E3 may additionally be associated with phenotypes other than enhanced S-cone syndrome. Conclusion: Characteristic ERG changes enable the diagnosis of cone dystrophy with supernormal rod ERG, enhanced S-cone syndrome, and bradyopsia and accurate genetic screening. This review highlights the need for additional nonstandard ERGs to make the diagnosis in two of these disorders.

A detailed clinical and molecular survey of subjects with nonsyndromic USH2A retinopathy reveals an allelic hierarchy of disease-causing variants

Defects in USH2A cause both isolated retinal disease and Usher syndrome (ie, retinal disease and deafness). To gain insights into isolated/nonsyndromic USH2A retinopathy, we screened USH2A in 186 probands with recessive retinal disease and no hearing complaint in childhood (discovery cohort) and in 84 probands with recessive retinal disease (replication cohort). Detailed phenotyping, including retinal imaging and audiological assessment, was performed in individuals with two likely disease-causing USH2A variants. Further genetic testing, including screening for a deep-intronic disease-causing variant and large deletions/duplications, was performed in those with one likely disease-causing change. Overall, 23 of 186 probands (discovery cohort) were found to harbour two likely disease-causing variants in USH2A. Some of these variants were predominantly associated with nonsyndromic retinal degeneration (‘retinal disease-specific’); these included the common c.2276 G>T, p.(Cys759Phe) mutation and five additional variants: c.2802 T>G, p.(Cys934Trp); c.10073 G>A, p.(Cys3358Tyr); c.11156 G>A, p.(Arg3719His); c.12295-3 T>A; and c.12575 G>A, p.(Arg4192His). An allelic hierarchy was observed in the discovery cohort and confirmed in the replication cohort. In nonsyndromic USH2A disease, retinopathy was consistent with retinitis pigmentosa and the audiological phenotype was variable. USH2A retinopathy is a common cause of nonsyndromic recessive retinal degeneration and has a different mutational spectrum to that observed in Usher syndrome. The following model is proposed: the presence of at least one ‘retinal disease-specific’ USH2A allele in a patient with USH2A-related disease results in the preservation of normal hearing. Careful genotype–phenotype studies such as this will become increasingly important, especially now that high-throughput sequencing is widely used in the clinical setting.

Biallelic Mutations in DNAJC21 Cause Shwachman-Diamond Syndrome

Program in Genetics and Genome Biology, Research Institute, The Hospital for Sick Children, Toronto, ON, Canada; Institute of Medical Science, University of Toronto, Toronto, ON, Canada; Marrow Failure and Myelodysplasia Program, Division of Haematology/Oncology, Department of Paediatrics, The Hospital for Sick Children, Toronto, ON, Canada; Division of Haematology Oncology, Children’s Hospital of Western Ontario, London, ON, Canada; Division of Clinical and Metabolic Genetics, Department of Ophthalmology and Vision Sciences, and Division of Gastroenterology and Nutrition, The

Achromatopsia mutations target sequential steps of ATF6 activation

Significance The unfolded protein response regulator activating transcription factor 6 (ATF6) was recently identified as a novel genetic cause of the cone photoreceptor disease achromatopsia. ATF6 upregulates genes that help cells cope with endoplasmic reticulum stress. We identified the pathomechanisms of all ATF6 achromatopsia mutations. Class 1 ATF6 mutants show impaired endoplasmic reticulum (ER)-to-Golgi trafficking and diminished production of the transcriptional activator fragment. Class 2 mutants encode the intact ATF6 transcriptional activator domain with full activity. Class 3 mutants have defective basic leucine zipper (bZIP) domains with abrogated function. Patient fibroblasts show increased apoptosis after ER stress. Our findings reveal that human ATF6 mutations interrupt distinct steps of ATF6 activation. ER stress-associated damage may underlie the pathology of achromatopsia arising from ATF6. Achromatopsia is an autosomal recessive disorder characterized by cone photoreceptor dysfunction. We recently identified activating transcription factor 6 (ATF6) as a genetic cause of achromatopsia. ATF6 is a key regulator of the unfolded protein response. In response to endoplasmic reticulum (ER) stress, ATF6 migrates from the ER to Golgi to undergo regulated intramembrane proteolysis to release a cytosolic domain containing a basic leucine zipper (bZIP) transcriptional activator. The cleaved ATF6 fragment migrates to the nucleus to transcriptionally up-regulate protein-folding enzymes and chaperones. ATF6 mutations in patients with achromatopsia include missense, nonsense, splice site, and single-nucleotide deletion or duplication changes found across the entire gene. Here, we comprehensively tested the function of achromatopsia-associated ATF6 mutations and found that they group into three distinct molecular pathomechanisms: class 1 ATF6 mutants show impaired ER-to-Golgi trafficking and diminished regulated intramembrane proteolysis and transcriptional activity; class 2 ATF6 mutants bear the entire ATF6 cytosolic domain with fully intact transcriptional activity and constitutive induction of downstream target genes, even in the absence of ER stress; and class 3 ATF6 mutants have complete loss of transcriptional activity because of absent or defective bZIP domains. Primary fibroblasts from patients with class 1 or class 3 ATF6 mutations show increased cell death in response to ER stress. Our findings reveal that human ATF6 mutations interrupt distinct sequential steps of the ATF6 activation mechanism. We suggest that increased susceptibility to ER stress-induced damage during retinal development underlies the pathology of achromatopsia in patients with ATF6 mutations.

Mutational Analysis of SDCCAG8 in Bardet-Biedl Syndrome Patients with Renal Involvement and Absent Polydactyly

Purpose: To assess for SDCCAG8 mutations in Bardet-Biedl syndrome (BBS) subjects with renal involvement and no polydactyly, and to describe phenotypic characteristics of SDCCAG8-related disease. Material and Methods: Five patients (from 4 pedigrees) with clinical diagnosis of BBS, who had retinal and renal involvement and no polydactyly, were assessed. Sequence analysis of SDCCAG8 was undertaken and a detailed clinical review of an affected sibship was performed. Results: A sibship of East Indian origin who carried a putative clinical diagnosis of BBS had compound heterozygous mutations in SDCCAG8 (p.Thr482LysfsX12/p.Asp543AlafsX24). The renal involvement was early and required transplant in both cases. Both were short statured and had asthma since childhood. The younger sister also had non-alcoholic fatty liver disease. Visual acuity and central fields were preserved in the teenage years in both patients. The optical coherence tomography showed preservation of the retinal lamination at the fovea; fundus autofluorescence demonstrated a perifoveal ring of hyperfluorescence as commonly observed in other forms of retinitis pigmentosa. Full-field electroretinogram revealed rod function to be more severely affected than cone function in both cases. Conclusion: Our results and prior literature suggest that SDCCAG8 could play an important role in presumed BBS patients affected with severe kidney disease and absent polydactyly. This report enhances the phenotypic description of SDCCAG8-related disease.

Molecular modeling indicates distinct classes of missense variants with mild and severe XLRS phenotypes

X-linked retinoschisis (XLRS) is a vitreo-retinal degeneration caused by mutations in the RS1 gene which encodes the protein retinoschisin (RS1), required for the structural and functional integrity of the retina. Data are presented from a group of 38 XLRS patients from Moorfields Eye Hospital (London, UK) who had one of 18 missense mutations in RS1. Patients were grouped based on mutation severity predicted by molecular modeling: mild (class I), moderate (intermediate) and severe (class II). Most patients had an electronegative scotopic bright flash electroretinogram (ERG) (reduced b/a-wave ratio) in keeping with predominant inner retinal dysfunction. An association between the type of structural RS1 alterations and the severity of b/a-wave reduction was found in all but the oldest group of patients, significant in patients aged 15-30 years. Severe RS1 missense changes were associated with a lower ERG b/a ratio than were mild changes, suggesting that the extent of inner retinal dysfunction is influenced by the effect of the mutations on protein structure. The majority of class I mutations showed no changes involving cysteine residues. Class II mutations caused severe perturbations due to the removal or insertion of cysteine residues or due to changes in the hydrophobic core. The ERG b/a ratio in intermediate cases was abnormal but showed significant variability, possibly related to the role of proline or arginine residues. We also conducted a second study, using a completely independent cohort, to indicate a genotype-ERG phenotype correlation.

Oligocone trichromacy is part of the spectrum of CNGA3-related cone system disorders

Purpose: To report the rare observation of CNGA3 mutation as a cause of oligocone trichromacy (OT) and present phenotypic characteristics. Methods: A 20 year old male patient underwent ophthalmological evaluation including detailed color vision assessment using Ishihara pseudoisochromatic plates, American Optical Hardy Rand Rittler plates (HRR) and Mollon-Reffin Minimalist test (MRM). Optical coherence tomography (OCT), fundus autofluorescence (FAF), visual field assessment and electrophysiological testing was also performed. The patient’s DNA was sequenced for mutations in the coding sequence of CNGA3 and CNGB3 genes. Results: Best corrected visual acuity (BCVA) was 20/50 and 20/30 in the right and left eyes respectively. His color vision was normal to Ishihara, HRR and MRM tests. Fundus appearance, FAF, OCT and Goldmann visual fields (GVF) were all normal. Humphrey visual field analysis (HVF) demonstrated reduced sensitivity and paracentral scotomas (5–20°). The full-field electroretinogram (ERG) showed normal rod responses and severely reduced cone responses. The multifocal electroretinogram (mfERG) was non-recordable above noise. Compound heterozygous mutations in exon 8 of the CNGA3 coding sequence were identified; c.1070 A > G (Tyr357Cys; novel) and c.1694 C > T (Thr565Met). Allele-specific polymerase chain reaction confirmed that the mutations were located on separate alleles. No mutations were identified in CNGB3. Conclusion: This is the second reported case of CNGA3 associated OT. Mutations in CNGA3 have previously been associated with incomplete and complete achromatopsia. This report confirms that OT forms the mildest end of the spectrum of CNGA3 related diseases.