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Citation: Khan KN, El-Asrag ME, Ku CA, et al.; for NIHR BioResource-Rare Diseases and UK Inherited Retinal Disease Consortium. Specific alleles of CLN7/ MFSD8, a protein that localizes to photoreceptor synaptic terminals, cause a spectrum of nonsyndromic retinal dystrophy. Invest Ophthalmol Vis Sci. 2017;58:290658: -291458: . DOI:10.1167 PURPOSE. Recessive mutations in CLN7/MFSD8 usually cause variant late-infantile onset neuronal ceroid lipofuscinosis (vLINCL), a poorly understood neurodegenerative condition, though mutations may also cause nonsyndromic maculopathy. A series of 12 patients with nonsyndromic retinopathy due to novel CLN7/MFSD8 mutation combinations were investigated in this study. METHODS.Affected patients and their family members were recruited in ophthalmic clinics at each center where they were examined by retinal imaging and detailed electrophysiology. Whole exome or genome next generation sequencing was performed on genomic DNA from at least one affected family member. Immunofluorescence confocal microscopy of murine retina cross-sections were used to localize the protein. RESULTS.Compound heterozygous alleles were identified in six cases, one of which was always p.Glu336Gln. Such combinations resulted in isolated macular disease. Six further cases were homozygous for the variant p.Met454Thr, identified as a founder mutation of South Asian origin. Those patients had widespread generalized retinal disease, characterized by electroretinography as a rod-cone dystrophy with severe macular involvement. In addition, the photopic single flash electroretinograms demonstrated a reduced b-to a-wave amplitude ratio, suggesting dysfunction occurring after phototransduction. Immunohistology identified MFSD8 in the outer plexiform layer of the retina, a site rich in photoreceptor synapses.CONCLUSIONS. This study highlights a hierarchy of MFSD8 variant severity, predicting three consequences of mutation: (1) nonsyndromic localized maculopathy, (2) nonsyndromic widespread retinopathy, or (3) syndromic neurological disease. The data also shed light on the underlying pathogenesis by implicating the photoreceptor synaptic terminals as the major site of retinal disease.

variants also displayed a substantial reduction in sperm motility and infertility, whereas those carrying missense changes were fertile. Defects in this polyglutamylase in humans have recently been associated with cone photoreceptor dystrophy, while mouse models carrying truncating mutations in the same gene also display reduced fertility in male animals. We examined the expression levels of TTLL5 in various human tissues and determined that this gene has multiple viable isoforms, being highly expressed in testis and retina. In addition, antibodies against TTLL5 stained the basal body of photoreceptor cells in rat and the centrosome of the spermatozoon flagellum in humans, suggesting a common mechanism of action in these two cell types. Taken together, our data indicate that mutations in TTLL5 delineate a novel, allele--specific syndrome causing defects in two as yet pathogenically unrelated functions, reproduction and vision.4

Patients with DRAM2-retinopathy are typically asymptomatic in the first two decades of life and present with central visual loss and a maculopathy. A faint hyperautofluorescent ring on FAF can be a suggestive feature. The retinal periphery is frequently affected later in the disease process. Photoreceptor degeneration is likely to be the primary event and future studies on DRAM2-retinopathy are expected to provide important insights into retinal autophagy.

Inherited retinal diseases (IRDs) display an enormous genetic heterogeneity. Whole exome sequencing (WES) recently identified genes that were mutated in a small proportion of IRD cases. Consequently, finding a second case or family carrying pathogenic variants in the same candidate gene often is challenging. In this study, we searched for novel candidate IRD gene-associated variants in isolated IRD families, assessed their causality, and searched for novel genotype-phenotype correlations. Whole exome sequencing was performed in 11 probands affected with IRDs. Homozygosity mapping data was available for five cases. Variants with minor allele frequencies ≤ 0.5% in public databases were selected as candidate disease-causing variants. These variants were ranked based on their: (a) presence in a gene that was previously implicated in IRD; (b) minor allele frequency in the Exome Aggregation Consortium database (ExAC); (c) in silico pathogenicity assessment using the combined annotation dependent depletion (CADD) score; and (d) interaction of the corresponding protein with known IRD-associated proteins. Twelve unique variants were found in 11 different genes in 11 IRD probands. Novel autosomal recessive and dominant inheritance patterns were found for variants in Small Nuclear Ribonucleoprotein U5 Subunit 200 (SNRNP200) and Zinc Finger Protein 513 (ZNF513), respectively. Using our pathogenicity assessment, a variant in DEAH-Box Helicase 32 (DHX32) was the top ranked novel candidate gene to be associated with IRDs, followed by eight medium and lower ranked candidate genes. The identification of candidate disease-associated sequence variants in 11 single families underscores the notion that the previously identified IRD-associated genes collectively carry > 90% of the defects implicated in IRDs. To identify multiple patients or families with variants in the same gene and thereby provide extra proof for pathogenicity, worldwide data sharing is needed.

ardet-Biedl syndrome (BBS; OMIM 209900) is an autosomal recessive disease that is heterogeneous both clinically and genetically and is characterized by a wide and variable spectrum of clinical features that can be categorized as primary features (including retinal degeneration, polydactyly, renal and gonadal malformations, obesity, and learning disabilities) or secondary features (including speech disorders, developmental delay, ataxia, diabetes mellitus, dysmorphic features, and variable cognitive impairment). The clinical diagnosis of BBS is based on the presence of at least 4 primary features or a combination of 3 primary and 2 (or more) secondary features. 1 Bardet-Biedl syndrome is considered to be one of the ciliopathies, 2 a group of inherited diseases caused by mutations in genes encoding ciliary proteins. Owing to an overlap of clinical features with other ciliopathies, an accurate diagnosis of BBS is sometimes difficult.The prevalence of BBS was estimated to be 1 in 77 000 persons in Denmark, 3 1 in 50 000-65 000 persons in Kuwait, 4,5 and a relatively high prevalence of 1 in 3700 persons in the Faroe Islands due to a common founder mutation. 6 Bardet-Biedl syndrome is a genetically heterogeneous disorder that can be caused by mutations in at least 15 genes. The inheritance pattern is autosomal recessive, but triallelic inheritance was reported as a possible mechanism in some families. 7 The genes that are known to cause BBS when mutated encode proteins that localize to the basal body of the cilium and are associated with ciliogenesis and intraflagellar transport. Most BBS proteins form the BBSome complex, which is localized to nonmembranous centriolar satellites in the cytoplasm, but also to the membrane of the cilium, and is required for ciliogenesis and functions in ciliary membrane biogenesis. 8 IMPORTANCE A large number of genes can cause inherited retinal degenerations when mutated. It is important to identify the cause of disease for a better disease prognosis and a possible gene-specific therapeutic intervention.OBJECTIVE To identify the cause of disease in families with nonsyndromic retinitis pigmentosa. DESIGN, SETTING, AND PARTICIPANTSPatients and family members were recruited for the study and underwent clinical evaluation and genetic analyses.MAIN OUTCOMES AND MEASURES Identification of sequence variants in genes using next-generation sequencing.RESULTS We performed exome sequencing for 4 families, which was followed by Sanger sequencing of the identified mutations in 120 ethnicity-matched patients. In total, we identified 4 BBS2 missense mutations that cause nonsyndromic retinitis pigmentosa. Three siblings of Moroccan Jewish ancestry were compound heterozygotes for p.A33D and p.P134R, and 6 patients belonging to 4 families of Ashkenazi Jewish ancestry were homozygous for either p.D104A or p.R632P, or compound heterozygous for these 2 mutations. The mutations cosegregated with retinitis pigmentosa in the studied families, and the affected amino acid residues are evolutionarily conserved. C...

PurposeRetinal dystrophies are genetically heterogeneous, resulting from mutations in over 200 genes. Prior to the development of massively parallel sequencing, comprehensive genetic screening was unobtainable for most patients. Identifying the causative genetic mutation facilitates genetic counselling, carrier testing and prenatal/pre-implantation diagnosis, and often leads to a clearer prognosis. In addition, in a proportion of cases, when the mutation is known treatment can be optimised and patients are eligible for enrolment into clinical trials for gene-specific therapies.MethodsPatient genomic DNA was sheared, tagged and pooled in batches of four samples, prior to targeted capture and next generation sequencing. The enrichment reagent was designed against genes listed on the RetNet database (July 2010). Sequence data were aligned to the human genome and variants were filtered to identify potential pathogenic mutations. These were confirmed by Sanger sequencing.ResultsMolecular analysis of 20 DNAs from retinal dystrophy patients identified likely pathogenic mutations in 12 cases, many of them known and/or confirmed by segregation. These included previously described mutations in ABCA4 (c.6088C>T,p.R2030*; c.5882G>A,p.G1961E), BBS2 (c.1895G>C,p.R632P), GUCY2D (c.2512C>T,p.R838C), PROM1 (c.1117C>T,p.R373C), RDH12 (c.601T>C,p.C201R; c.506G>A,p.R169Q), RPGRIP1 (c.3565C>T,p.R1189*) and SPATA7 (c.253C>T,p.R85*) and new mutations in ABCA4 (c.3328+1G>C), CRB1 (c.2832_2842+23del), RP2 (c.884-1G>T) and USH2A (c.12874A>G,p.N4292D).ConclusionsTagging and pooling DNA prior to targeted capture of known retinal dystrophy genes identified mutations in 60% of cases. This relatively high success rate may reflect enrichment for consanguineous cases in the local Yorkshire population, and the use of multiplex families. Nevertheless this is a promising high throughput approach to retinal dystrophy diagnostics.

Primary cilia defects result in a group of related pleiotropic malformation syndromes known as ciliopathies, often characterised by cerebellar developmental and foliation defects. Here, we describe the cerebellar anatomical and signalling defects in the Tmem67 tm1 ( Dgen )/ H knockout mouse. At mid-gestation, Tmem67 mutant cerebella were hypoplastic and had aberrantly high canonical Wnt/β-catenin signalling, proliferation and apoptosis. Later in development, mutant cerebellar hemispheres had severe foliation defects and inferior lobe malformation, characterized by immature Purkinje cells (PCs). Early postnatal Tmem67 mutant cerebellum had disrupted ciliogenesis and reduced responsiveness to Shh signalling. Transcriptome profiling of Tmem67 mutant cerebella identified ectopic increased expression of homeobox-type transcription factors ( Hoxa5 , Hoxa4 , Hoxb5 and Hoxd3 ), normally required for early rostral hindbrain patterning. HOXB5 protein levels were increased in the inferior lobe, and increased canonical Wnt signalling, following loss of TMEM67, was dependent on HOXB5. HOXB5 occupancy at the β-catenin promoter was significantly increased by activation of canonical Wnt signalling in Tmem67 −/− mutant cerebellar neurones, suggesting that increased canonical Wnt signalling following mutation or loss of TMEM67 was directly dependent on HOXB5. Our results link dysregulated expression of Hox group genes with ciliary Wnt signalling defects in the developing cerebellum, providing new mechanistic insights into ciliopathy cerebellar hypoplasia phenotypes.