Biallelic MAD2L1BP (p31comet) mutation is associated with mosaic aneuploidy and juvenile granulosa cell tumors.

MAD2L1BP-encoded p31comet mediates Trip13-dependent disassembly of Mad2- and Rev7-containing complexes and, through this antagonism, promotes timely spindle assembly checkpoint (SAC) silencing, faithful chromosome segregation, insulin signaling, and homology-directed repair (HDR) of DNA double-strand breaks. We identified a homozygous MAD2L1BP nonsense variant, R253*, in 2 siblings with microcephaly, epileptic encephalopathy, and juvenile granulosa cell tumors of ovary and testis. Patient-derived cells exhibited high-grade mosaic variegated aneuploidy, slowed-down proliferation, and instability of truncated p31comet mRNA and protein. Corresponding recombinant p31comet was defective in Trip13, Mad2, and Rev7 binding and unable to support SAC silencing or HDR. Furthermore, C-terminal truncation abrogated an identified interaction of p31comet with tp53. Another homozygous truncation, R227*, detected in an early-deceased patient with low-level aneuploidy, severe epileptic encephalopathy, and frequent blood glucose elevations, likely corresponds to complete loss of function, as in Mad2l1bp-/- mice. Thus, human mutations of p31comet are linked to aneuploidy and tumor predisposition.

The tectonic complex regulates membrane protein composition in the photoreceptor cilium.

The primary cilium is a signaling organelle with a unique membrane composition maintained by a diffusional barrier residing at the transition zone. Many transition zone proteins, such as the tectonic complex, are linked to preserving ciliary composition but the mechanism remains unknown. To understand tectonic's role, we generate a photoreceptor-specific Tctn1 knockout mouse. Loss of Tctn1 results in the absence of the entire tectonic complex and associated MKS proteins yet has minimal effects on the transition zone structure of rod photoreceptors. We find that the protein composition of the photoreceptor cilium is disrupted as non-resident membrane proteins accumulate in the cilium over time, ultimately resulting in photoreceptor degeneration. We further show that fluorescent rhodopsin moves faster through the transition zone in photoreceptors lacking tectonic, which suggests that the tectonic complex acts as a physical barrier to slow down membrane protein diffusion in the photoreceptor transition zone to ensure proper removal of non-resident membrane proteins.

[Inherited retinal diseases in Germany-Challenges in health care supply structure and diagnostics]. / Erbliche Netzhautdystrophien in Deutschland ­ Versorgungsstrukturelle und diagnostische Herausforderungen.

BACKGROUND: Inherited retinal diseases (IRD) are rare eye diseases and pose high diagnostic challenges. A care structure with few highly specialized centers in Germany, misdiagnosis due to the lack of molecular genetic testing, and a lack of a central registry lead to a lack of reliable information on the prevalence and distribution of IRDs in Germany. METHODS: Based on clinical data from an ophthalmological center and molecular data from a genetic center as well as a nationwide health insurance data query, we estimated the prevalence of IRDs in Germany in addition to collecting information on their phenotypic and genotypic distribution. RESULTS: The median travelling distance to the ophthalmological center was 60 km. The most frequent diagnoses were retinitis pigmentosa, macular dystrophy and general retinal dystrophy. Molecular genetic testing was performed in 87% of patients with clinical suspicion of IRD, with marked differences in frequencies among age cohorts. The molecular genetic detection rate in the genetic center was 51%. The prevalence of inherited retinal dystrophy in Germany determined by health insurance data retrieval was approximately 1:1150. CONCLUSION: Many patients must travel long distances to visit specialized clinics for IRDs with access to genetic testing. To obtain more reliable numbers on the prevalence in Germany, routine molecular genetic testing, and a national registry for IRD detection are needed.

Expanding the phenotypic spectrum and clinical severity associated with WLS gene.

WLS (Wnt ligand secretion mediator or Wntless) orchestrates the secretion of all Wnt proteins, a family of evolutionary conserved proteins, involved in Wnt signaling pathway that has many essential biological functions including the regulation of development, cell proliferation, migration and apoptosis. Biallelic variants in WLS have recently been described in 10 patients with pleiotropic multiple congenital anomalies (MCA) known as Zaki syndrome. We identified a likely disease-causing variant in WLS (c.1579G>A, p.Gly527Arg) in a boy presented with a broad range of MCA including microcephaly, facial dysmorphism, alopecia, ophthalmologic anomalies, and complete soft tissue syndactyly. These features were reminiscent of Zaki syndrome although variable clinical severity was observed. In a detailed clinical assessment, our patient also displayed microphthalmia, dental anomalies, skeletal dysplasia with spontaneous fractures and Dandy-Walker malformation. As such, we extend the phenotype linked to Zaki syndrome. This study further highlights the importance of a thorough clinical evaluation to delineate the phenotypic spectrum associated with WLS variants and suggests that genotype-phenotype correlations due to variant localization seems likely. However, future work on additional patients and more functional studies may give further insights into genotype-phenotype correlations and the complex function of WLS.

CEP162 deficiency causes human retinal degeneration and reveals a dual role in ciliogenesis and neurogenesis.

Defects in primary or motile cilia result in a variety of human pathologies, and retinal degeneration is frequently associated with these so-called ciliopathies. We found that homozygosity for a truncating variant in CEP162, a centrosome and microtubule-associated protein required for transition zone assembly during ciliogenesis and neuronal differentiation in the retina, caused late-onset retinitis pigmentosa in 2 unrelated families. The mutant CEP162-E646R*5 protein was expressed and properly localized to the mitotic spindle, but it was missing from the basal body in primary and photoreceptor cilia. This impaired recruitment of transition zone components to the basal body and corresponded to complete loss of CEP162 function at the ciliary compartment, reflected by delayed formation of dysmorphic cilia. In contrast, shRNA knockdown of Cep162 in the developing mouse retina increased cell death, which was rescued by expression of CEP162-E646R*5, indicating that the mutant retains its role for retinal neurogenesis. Human retinal degeneration thus resulted from specific loss of the ciliary function of CEP162.

A novel large in-frame FBN1 deletion causes neonatal Marfan syndrome.

Neonatal Marfan syndrome (nMFS) is a rare and severe form of Marfan syndrome (MFS) with a poor prognosis, that presents with a highly variable phenotype, particularly regarding skeletal, ocular, and cardiovascular manifestations. Mutations in the fibrillin-1 (FBN1) gene are known as the principal cause of MFS and MFS-related syndromes. Here, we report on a full-term female neonate with postnatal characteristics suggestive of nMFS, including severe cardiovascular disease resulting in cardiorespiratory failure and death by 4 mo of age. We identified a novel large genomic in-frame deletion of FBN1 exons 42-45, c.(5065 + 1_5066 - 1)_(5545 + 1_5546 - 1)del. Large FBN1 in-frame deletions between exons 24 and 53 have been associated with severe MFS. The deletion in our patient differs from the FBN1 region associated with the majority of nMFS cases, exons 24-32.

KCND2 variants associated with global developmental delay differentially impair Kv4.2 channel gating.

Here, we report on six unrelated individuals, all presenting with early-onset global developmental delay, associated with impaired motor, speech and cognitive development, partly with developmental epileptic encephalopathy and physical dysmorphisms. All individuals carry heterozygous missense variants of KCND2, which encodes the voltage-gated potassium (Kv) channel α-subunit Kv4.2. The amino acid substitutions associated with the variants, p.(Glu323Lys) (E323K), p.(Pro403Ala) (P403A), p.(Val404Leu) (V404L) and p.(Val404Met) (V404M), affect sites known to be critical for channel gating. To unravel their likely pathogenicity, recombinant mutant channels were studied in the absence and presence of auxiliary ß-subunits under two-electrode voltage clamp in Xenopus oocytes. All channel mutants exhibited slowed and incomplete macroscopic inactivation, and the P403A variant in addition slowed activation. Co-expression of KChIP2 or DPP6 augmented the functional expression of both wild-type and mutant channels; however, the auxiliary ß-subunit-mediated gating modifications differed from wild type and among mutants. To simulate the putative setting in the affected individuals, heteromeric Kv4.2 channels (wild type + mutant) were studied as ternary complexes (containing both KChIP2 and DPP6). In the heteromeric ternary configuration, the E323K variant exhibited only marginal functional alterations compared to homomeric wild-type ternary, compatible with mild loss-of-function. By contrast, the P403A, V404L and V404M variants displayed strong gating impairment in the heteromeric ternary configuration, compatible with loss-of-function or gain-of-function. Our results support the etiological involvement of Kv4.2 channel gating impairment in early-onset monogenic global developmental delay. In addition, they suggest that gain-of-function mechanisms associated with a substitution of V404 increase epileptic seizure susceptibility.

NPHP1 gene-associated nephronophthisis is associated with an occult retinopathy.

Biallelic deletions in the NPHP1 gene are the most frequent molecular defect of nephronophthisis, a kidney ciliopathy and leading cause of hereditary end-stage kidney disease. Nephrocystin 1, the gene product of NPHP1, is also expressed in photoreceptors where it plays an important role in intra-flagellar transport between the inner and outer segments. However, the human retinal phenotype has never been investigated in detail. Here, we characterized retinal features of 16 patients with homozygous deletions of the entire NPHP1 gene. Retinal assessment included multimodal imaging (optical coherence tomography, fundus autofluorescence) and visual function testing (visual acuity, full-field electroretinography, color vision, visual field). Fifteen patients had a mild retinal phenotype that predominantly affected cones, but with relative sparing of the fovea. Despite a predominant cone dysfunction, night vision problems were an early symptom in some cases. The consistent retinal phenotype on optical coherence tomography images included reduced reflectivity and often a granular appearance of the ellipsoid zone, fading or loss of the interdigitation zone, and mild outer retinal thinning. However, there were usually no obvious structural changes visible upon clinical examination and fundus autofluorescence imaging (occult retinopathy). More advanced retinal degeneration might occur with ageing. An identified additional CEP290 variant in one patient with a more severe retinal degeneration may indicate a potential role for genetic modifiers, although this requires further investigation. Thus, diagnostic awareness about this distinct retinal phenotype has implications for the differential diagnosis of nephronophthisis and for individual prognosis of visual function.

Peripapillary Sparing in Autosomal Recessive Bestrophinopathy.

PURPOSE: To demonstrate that peripapillary sparing on autofluorescence images is a characteristic feature of autosomal recessive bestrophinopathy (ARB). DESIGN: Retrospective, cross-sectional case series and review of previous published cases. PARTICIPANTS: Twelve patients with ARB. METHODS: Ophthalmic assessment included best-corrected visual acuity testing, electrophysiologic examinations, and multimodal retinal imaging. Retinal imaging included OCT, blue-light autofluorescence imaging, fundus photography, and widefield pseudocolor and autofluorescence fundus imaging. MAIN OUTCOME MEASURES: Presence of peripapillary sparing on fundus autofluorescence images. RESULTS: Relatively normal-appearing peripapillary autofluorescence was identified in all patients, independent of the disease stage or presence of widespread changes on autofluorescence widefield images. OCT images of the peripapillary region revealed mild structural abnormalities, including a thinned outer nuclear layer and intraretinal or subretinal fluid. A review of previously published cases confirmed peripapillary sparing as consistent feature on fundus autofluorescence images. Genetic analysis revealed 10 previously reported mutations, 1 novel missense (c.83T>A; p.Ile28Asn) and 2 novel truncating (c.658C>T; p.Gln220* and c.1370C>G; p.Ser457*) variants in BEST1. CONCLUSIONS: In ARB patients, peripapillary sparing is a consistent feature on fundus autofluorescence images, whereas the same region is less preserved on OCT images.

Comprehensive Geno- and Phenotyping in a Complex Pedigree Including Four Different Inherited Retinal Dystrophies.

Inherited retinal dystrophies (IRDs) are characterized by high clinical and genetic heterogeneity. A precise characterization is desirable for diagnosis and has impact on prognosis, patient counseling, and potential therapeutic options. Here, we demonstrate the effectiveness of the combination of in-depth retinal phenotyping and molecular genetic testing in complex pedigrees with different IRDs. Four affected Caucasians and two unaffected relatives were characterized including multimodal retinal imaging, functional testing, and targeted next-generation sequencing. A considerable intrafamilial phenotypic and genotypic heterogeneity was identified. While the parents of the index family presented with rod-cone dystrophy and ABCA4-related retinopathy, their two sons revealed characteristics in the spectrum of incomplete congenital stationary night blindness and ocular albinism, respectively. Molecular testing revealed previously described variants in RHO, ABCA4, and MITF as well as a novel variant in CACNA1F. Identified variants were verified by intrafamilial co-segregation, bioinformatic annotations, and in silico analysis. The coexistence of four independent IRDs caused by distinct mutations and inheritance modes in one pedigree is demonstrated. These findings highlight the complexity of IRDs and underscore the need for the combination of extensive molecular genetic testing and clinical characterization. In addition, a novel variant in the CACNA1F gene is reported associated with incomplete congenital stationary night blindness.

First confirmatory study on PTPRQ as an autosomal dominant non-syndromic hearing loss gene.

BACKGROUND: Biallelic PTPRQ pathogenic variants have been previously reported as causative for autosomal recessive non-syndromic hearing loss. In 2018 the first heterozygous PTPRQ variant has been implicated in the development of autosomal dominant non-syndromic hearing loss (ADNSHL) in a German family. The study presented the only, so far known, PTPRQ pathogenic variant (c.6881G>A) in ADNSHL. It is located in the last PTPRQ coding exon and introduces a premature stop codon (p.Trp2294*). METHODS: A five-generation Polish family with ADNSHL was recruited for the study (n = 14). Thorough audiological, neurotological and imaging studies were carried out to precisely define the phenotype. Genomic DNA was isolated from peripheral blood samples or buccal swabs of available family members. Clinical exome sequencing was conducted for the proband. Family segregation analysis of the identified variants was performed using Sanger sequencing. Single nucleotide polymorphism array on DNA samples from the Polish and the original German family was used for genome-wide linkage analysis. RESULTS: Combining clinical exome sequencing and family segregation analysis, we have identified the same (NM_001145026.2:c.6881G>A, NP_001138498.1:p.Trp2294*) PTPRQ alteration in the Polish ADNSHL family. Using genome-wide linkage analysis, we found that the studied family and the original German family derive from a common ancestor. Deep phenotyping of the affected individuals showed that in contrast to the recessive form, the PTPRQ-related ADNSHL is not associated with vestibular dysfunction. In both families ADNSHL was progressive, affected mainly high frequencies and had a variable age of onset. CONCLUSION: Our data provide the first confirmation of PTPRQ involvement in ADNSHL. The finding strongly reinforces the inclusion of PTPRQ to the small set of genes leading to both autosomal recessive and dominant hearing loss.

A Specific Macula-Predominant Retinal Phenotype Is Associated With the CDHR1 Variant c.783G>A, a Silent Mutation Leading to In-Frame Exon Skipping.

Purpose: To report the clinical and molecular findings in patients with retinal dystrophy associated with the c.783G>A variant in CDHR1. Methods: The retinal phenotype of 10 patients with CDHR1-related retinopathy was characterized by multimodal imaging including color fundus photography, optical coherence tomography (OCT), and blue- and near-infrared fundus autofluorescence imaging. Functional testing included electroretinography, visual acuity, and visual field testing. Results: Six patients homozygous for the c.783G>A variant in CDHR1 showed a retinal phenotype resembling central areolar choroidal dystrophy (CACD) on multimodal imaging. Retinal function outside an area of slowly progressive macular atrophy remained relatively preserved. In contrast, biallelic severe/truncating CDHR1 mutations result in retina-wide retinal degeneration in addition to macular atrophy, with overall severely reduced retinal function. Patients compound heterozygous for the c.783G>A mutation and a truncating mutation in CDHR1 showed an intermediate phenotype. All patients except one with biallelic severe CDHR1 mutations were asymptomatic in the first four decades of life, irrespective of their individual CDHR1 mutations. Analysis of blood RNA from patients with the c.783G>A variant revealed in-frame skipping of exon 8 in vivo, predicting a partial deletion of CDHR1 ectodomains 2 and 3. Conclusions: Patients with biallelic c.783G>A CDHR1 mutations demonstrate a retinal phenotype consistent with autosomal recessive CACD. The apparently silent dbSNP-annotated c.783G>A CDHR1 variant (rs147346345) has a relatively high minor allele frequency (0.31%), with homozygous individuals annotated in the general population, and it may therefore have been disregarded in many next-generation sequencing (NGS)-based studies. The differential diagnosis includes PRPH2-associated CACD and age-related macular degeneration.

Biallelic mutation of human SLC6A6 encoding the taurine transporter TAUT is linked to early retinal degeneration.

We previously reported that inactivation of the transmembrane taurine transporter (TauT or solute carrier 6a6) causes early retinal degeneration in mice. Compatible with taurine's indispensability for cell volume homeostasis, protein stabilization, cytoprotection, antioxidation, and immuno- and neuromodulation, mice develop multisystemic dysfunctions (hearing loss; liver fibrosis; and behavioral, heart, and skeletal muscle abnormalities) later on. Here, by genetic, cell biologic, in vivo1H-magnetic resonance spectroscopy and molecular dynamics simulation studies, we conducted in-depth characterization of a novel disorder: human TAUT deficiency. Loss of TAUT function due to a homozygous missense mutation caused panretinal degeneration in 2 brothers. TAUTp.A78E still localized in the plasma membrane but is predicted to impact structural stabilization. 3H-taurine uptake by peripheral blood mononuclear cells was reduced by 95%, and taurine levels were severely reduced in plasma, skeletal muscle, and brain. Extraocular dysfunctions were not yet detected, but significantly increased urinary excretion of 8-oxo-7,8-dihydroguanosine indicated generally enhanced (yet clinically unapparent) oxidative stress and RNA oxidation, warranting continuous broad surveillance.-Preising, M. N., Görg, B., Friedburg, C., Qvartskhava, N., Budde, B. S., Bonus, M., Toliat, M. R., Pfleger, C., Altmüller, J., Herebian, D., Beyer, M., Zöllner, H. J., Wittsack, H.-J., Schaper, J., Klee, D., Zechner, U., Nürnberg, P., Schipper, J., Schnitzler, A., Gohlke, H., Lorenz, B., Häussinger, D., Bolz, H. J. Biallelic mutation of human SLC6A6 encoding the taurine transporter TAUT is linked to early retinal degeneration.

Genetic testing in patients with retinitis pigmentosa: Features of unsolved cases.

IMPORTANCE: Uncommon characteristics in genetically unsolved retinitis pigmentosa (RP) patients may indicate an incorrect clinical diagnosis or as yet unknown genetic causes resulting in specific retinal phenotypes. The diagnostic yield of targeted next-generation sequencing may be increased by a reasonable preselection of RP-patients. BACKGROUND: To systematically evaluate and compare features of genetically solved and unsolved RP-patients. DESIGN: Retrospective, observational study. PARTICIPANTS: One-hundred and twelve consecutive RP-patients who underwent extensive molecular genetic analysis. METHODS: Characterization of patients based on multimodal imaging and medical history. MAIN OUTCOME MEASURES: Differences between genetically solved and unsolved RP-patients. RESULTS: Compared to genetically solved patients (n = 77), genetically unsolved patients (n = 35) more frequently had an age of disease-onset above 30 years (60% vs 8%; P < 0.0001), showed atypical fundus features (49% vs 8%; P < 0. 0001) and indicators for phenocopies (eg, autoimmune diseases) (17% vs 0%; P < 0. 001). Evidence for a particular inheritance pattern was less common (20% vs 49%; P < 0. 01). The diagnostic yield was 84% (71/85) in patients with first symptoms below 30 years-of-age, compared to 69% (77/112) in the overall cohort. The other selection criteria alone or in combination resulted in limited further increase of the diagnostic yield (up to 89%) while excluding considerably more patients (up to 56%) from genetic testing. CONCLUSIONS AND RELEVANCE: The medical history and retinal phenotype differ between genetically solved and a subgroup of unsolved RP-patients, which may reflect undetected genotypes or retinal conditions mimicking RP. Patient stratification may inform on the individual likelihood of identifying disease-causing mutations and may impact patient counselling.

Next-generation sequencing identifies unexpected genotype-phenotype correlations in patients with retinitis pigmentosa.

Retinitis pigmentosa (RP) is an inherited degenerative disease causing severe retinal dystrophy and visual impairment mainly with onset in infancy or adolescence. Targeted next-generation sequencing (NGS) has become an efficient tool to encounter the enormous genetic heterogeneity of diverse retinal dystrophies, including RP. To identify disease-causing mutations in unselected, consecutive RP patients, we conducted Sanger sequencing of genes commonly involved in the suspected genetic RP subtype, followed by targeted large-panel NGS if no mutation was identified, or NGS as primary analysis. A high (70%) detection rate of disease-causing mutations was achieved in a large cohort of 116 unrelated patients. About half (48%) of the solved RP cases were explained by mutations in four genes: RPGR, EYS, PRPF31 and USH2A. Overall, 110 different mutations distributed across 30 different genes were detected, and 46 of these mutations were novel. A molecular diagnosis was achieved in the majority (82-100%) of patients if the family history was suggestive for a particular mode of inheritance, but only in 60% in cases of sporadic RP. The diagnostic potential of extensive molecular analysis in a routine setting is also illustrated by the identification of unexpected genotype-phenotype correlations for RP patients with mutations in CRX, CEP290, RPGRIP1, MFSD8. Furthermore, we identified numerous mutations in autosomal dominant (PRPF31, PRPH2, CRX) and X-linked (RPGR) RP genes in patients with sporadic RP. Variants in RP2 and RPGR were also found in female RP patients with apparently sporadic or dominant disease. In summary, this study demonstrates that massively parallel sequencing of all known retinal dystrophy genes is a valuable diagnostic approach for RP patients.

Olfactory Dysfunction in Patients With CNGB1-Associated Retinitis Pigmentosa.

Importance: Co-occurrence of retinitis pigmentosa (RP) and olfactory dysfunction may have a common genetic cause. Objective: To report olfactory function and the retinal phenotype in patients with biallelic mutations in CNGB1, a gene coding for a signal transduction channel subunit expressed in rod photoreceptors and olfactory sensory neurons. Design, Setting, and Participants: This case series was conducted from August 2015 through July 2017. The setting was a multicenter study involving 4 tertiary referral centers for inherited retinal dystrophies. Participants were 9 patients with CNGB1-associated RP. Main Outcomes and Measures: Results of olfactory testing, ocular phenotyping, and molecular genetic testing using targeted next-generation sequencing. Results: Nine patients were included in the study, 3 of whom were female. Their ages ranged between 34 and 79 years. All patients had an early onset of night blindness but were usually not diagnosed as having RP before the fourth decade because of slow retinal degeneration. Retinal features were characteristic of a rod-cone dystrophy. Olfactory testing revealed reduced or absent olfactory function, with all except one patient scoring in the lowest quartile in relation to age-related norms. Brain magnetic resonance imaging and electroencephalography measurements in response to olfactory stimulation were available for 1 patient and revealed no visible olfactory bulbs and reduced responses to odor, respectively. Molecular genetic testing identified 5 novel (c.1312C>T, c.2210G>A, c.2492+1G>A, c.2763C>G, and c.3044_3050delGGAAATC) and 5 previously reported mutations in CNGB1. Conclusions and Relevance: Mutations in CNGB1 may cause an autosomal recessive RP-olfactory dysfunction syndrome characterized by a slow progression of retinal degeneration and variable anosmia or hyposmia.

Clinical and genetic characteristics of 251 consecutive patients with macular and cone/cone-rod dystrophy.

Macular and cone/cone-rod dystrophies (MD/CCRD) demonstrate a broad genetic and phenotypic heterogeneity, with retinal alterations solely or predominantly involving the central retina. Targeted next-generation sequencing (NGS) is an efficient diagnostic tool for identifying mutations in patient with retinitis pigmentosa, which shows similar genetic heterogeneity. To detect the genetic causes of disease in patients with MD/CCRD, we implemented a two-tier procedure consisting of Sanger sequencing and targeted NGS including genes associated with clinically overlapping conditions. Disease-causing mutations were identified in 74% of 251 consecutive MD/CCRD patients (33% of the variants were novel). Mutations in ABCA4, PRPH2 and BEST1 accounted for 57% of disease cases. Further mutations were identified in CDHR1, GUCY2D, PROM1, CRX, GUCA1A, CERKL, MT-TL1, KIF11, RP1L1, MERTK, RDH5, CDH3, C1QTNF5, CRB1, JAG1, DRAM2, POC1B, NPHP1 and RPGR. We provide detailed illustrations of rare phenotypes, including autofluorescence and optical coherence tomography imaging. Targeted NGS also identified six potential novel genotype-phenotype correlations for FAM161A, INPP5E, MERTK, FBLN5, SEMA4A and IMPDH1. Clinical reassessment of genetically unsolved patients revealed subgroups with similar retinal phenotype, indicating a common molecular disease cause in each subgroup.

Recurrent and Prolonged Infections in a Child with a Homozygous IFIH1 Nonsense Mutation.

In an Egyptian girl born to consanguineous parents, whole-exome sequencing (WES) identified a homozygous mutation in PHGDH, c.1273G>A (p.Val425Met), indicating 3-phosphoglycerate dehydrogenase deficiency. This diagnosis was compatible with the patient's microcephaly, severe psychomotor retardation, seizures and cataracts. However, she additionally suffered from recurrent (at least monthly) episodes of prolonged and severe chest infections requiring hospitalization, suggesting a secondary, predisposing and potentially Mendelian, condition. A local reactivation of an EBV infection in the respiratory tract was detected after a recent chest infection, likely representing an opportunistic infection based on a compromised immune system. Further inspection of WES data revealed a homozygous nonsense mutation, c.2665A>T (p.Lys889∗), in IFIH1, encoding MDA5. MDA5 detects long viral double-stranded RNA that is generated during replication of picorna viruses, and thereby activates the type I interferon signaling pathway. The results of Western blot analysis of protein from cultured fibroblasts of the patient indicates absence of wild type MDA5/IFIH1, compatible with NMD. We propose that, analogous to the severe course of primary influenza infection due to biallelic deficiency of a downstream effector, IRF7, homozygous loss of IFIH1 defines a novel Mendelian immunodeficiency disorder that increases susceptibility to severe viral infections. This is contrasted to heterozygous gain-of-function IFIH1 mutations in autoimmune diseases. Our findings highlight the potential of comprehensive genomic investigations in patients from consanguineous families to identify monogenic predispositions to severe infections.

Next-generation sequencing reveals the mutational landscape of clinically diagnosed Usher syndrome: copy number variations, phenocopies, a predominant target for translational read-through, and PEX26 mutated in Heimler syndrome.

BACKGROUND: Combined retinal degeneration and sensorineural hearing impairment is mostly due to autosomal recessive Usher syndrome (USH1: congenital deafness, early retinitis pigmentosa (RP); USH2: progressive hearing impairment, RP). METHODS: Sanger sequencing and NGS of 112 genes (Usher syndrome, nonsyndromic deafness, overlapping conditions), MLPA, and array-CGH were conducted in 138 patients clinically diagnosed with Usher syndrome. RESULTS: A molecular diagnosis was achieved in 97% of both USH1 and USH2 patients, with biallelic mutations in 97% (USH1) and 90% (USH2), respectively. Quantitative readout reliably detected CNVs (confirmed by MLPA or array-CGH), qualifying targeted NGS as one tool for detecting point mutations and CNVs. CNVs accounted for 10% of identified USH2A alleles, often in trans to seemingly monoallelic point mutations. We demonstrate PTC124-induced read-through of the common p.Trp3955* nonsense mutation (13% of detected USH2A alleles), a potential therapy target. Usher gene mutations were found in most patients with atypical Usher syndrome, but the diagnosis was adjusted in case of double homozygosity for mutations in OTOA and NR2E3, genes implicated in isolated deafness and RP. Two patients with additional enamel dysplasia had biallelic PEX26 mutations, for the first time linking this gene to Heimler syndrome. CONCLUSION: Targeted NGS not restricted to Usher genes proved beneficial in uncovering conditions mimicking Usher syndrome.