Relatively mild blue cone monochromacy phenotype caused by various haplotypes in the L- and M-cone opsin genes.

Purpose: Blue cone monochromacy (BCM) is an X-linked retinopathy caused by mutations in the red and green cone opsin genes. The aim of this study was to establish the clinical, genetic, and electrophysiological characteristics of a specific form of BCM. Methods: Patients harboring mutations in the OPN1LW/OPN1MW genes underwent a full clinical examination, including ocular examination, color vision, full-field electroretinography, color fundus and autofluorescence photography, and optical coherence tomography. Genetic analysis was performed using whole-exome sequencing, duplex PCR, PCR/restriction fragment length polymorphism, and Sanger sequencing. IBM SPSS Statistics v. 21.0 was used for the data analysis. Results: Twenty-five patients harboring various haplotypes in exon 3 of the OPN1LW/OPN1MW genes were recruited. They showed a milder incomplete phenotype of BCM than the typical BCM control group. They presented significantly better visual acuity (logarithm of the minimum angle of resolution [logMAR] 0.48 ± 0.26 vs. 1.10 ± 0.54; p < 0.0001) and a highly myopic refraction (-7.81 ± 5.81 D vs. -4.78 ± 5.27 D; p = 0.0222) compared with the BCM control group. The study group had higher 30-Hz cone flicker responses (28.60 ± 15.02 µv; n = 24), whereas the BCM group had none (0.66 ± 2.12 µv; n = 21; p < 0.0001). The Lanthony 15-HUE desaturated test was variable for the exon 3 haplotype group, with a tendency toward the deutan-protan axis. Conclusions: The present study included genetic and clinical data from the largest cohort of patients with exon 3 haplotypes that were previously shown to cause missplicing of the OPN1LW and OPN1MW genes. Analysis of the clinical data revealed better best-corrected visual acuity, more severe myopia, and higher 30-Hz cone flicker responses in the patients with exon 3 haplotypes than in those with typical BCM.

Rare germline heterozygous missense variants in BRCA1-associated protein 1, BAP1, cause a syndromic neurodevelopmental disorder.

Nuclear deubiquitinase BAP1 (BRCA1-associated protein 1) is a core component of multiprotein complexes that promote transcription by reversing the ubiquitination of histone 2A (H2A). BAP1 is a tumor suppressor whose germline loss-of-function variants predispose to cancer. To our knowledge, there are very rare examples of different germline variants in the same gene causing either a neurodevelopmental disorder (NDD) or a tumor predisposition syndrome. Here, we report a series of 11 de novo germline heterozygous missense BAP1 variants associated with a rare syndromic NDD. Functional analysis showed that most of the variants cannot rescue the consequences of BAP1 inactivation, suggesting a loss-of-function mechanism. In T cells isolated from two affected children, H2A deubiquitination was impaired. In matching peripheral blood mononuclear cells, histone H3 K27 acetylation ChIP-seq indicated that these BAP1 variants induced genome-wide chromatin state alterations, with enrichment for regulatory regions surrounding genes of the ubiquitin-proteasome system (UPS). Altogether, these results define a clinical syndrome caused by rare germline missense BAP1 variants that alter chromatin remodeling through abnormal histone ubiquitination and lead to transcriptional dysregulation of developmental genes.

Loss-of-function variants in DNM1 cause a specific form of developmental and epileptic encephalopathy only in biallelic state.

BACKGROUND: Developmental and epileptic encephalopathies (DEEs) represent a group of severe neurological disorders characterised by an onset of refractory seizures during infancy or early childhood accompanied by psychomotor developmental delay or regression. DEEs are genetically heterogeneous with, to date, more than 80 different genetic subtypes including DEE31 caused by heterozygous missense variants in DNM1. METHODS: We performed a detailed clinical characterisation of two unrelated patients with DEE and used whole-exome sequencing to identify causative variants in these individuals. The identified variants were tested for cosegregation in the respective families. RESULTS: We excluded pathogenic variants in known, DEE-associated genes. We identified homozygous nonsense variants, c.97C>T; p.(Gln33*) in family 1 and c.850C>T; p.(Gln284*) in family 2, in the DNM1 gene, indicating that biallelic, loss-of-function pathogenic variants in DNM1 cause DEE. CONCLUSION: Our finding that homozygous, loss-of-function variants in DNM1 cause DEE expands the spectrum of pathogenic variants in DNM1. All parents who were heterozygous carriers of the identified loss-of-function variants were healthy and did not show any clinical symptoms, indicating that the type of mutation in DNM1 determines the pattern of inheritance.

Recurrent de novo missense variants in GNB2 can cause syndromic intellectual disability.

PURPOSE: Binding proteins (G-proteins) mediate signalling pathways involved in diverse cellular functions and comprise Gα and Gßγ units. Human diseases have been reported for all five Gß proteins. A de novo missense variant in GNB2 was recently reported in one individual with developmental delay/intellectual disability (DD/ID) and dysmorphism. We aim to confirm GNB2 as a neurodevelopmental disease gene, and elucidate the GNB2-associated neurodevelopmental phenotype in a patient cohort. METHODS: We discovered a GNB2 variant in the index case via exome sequencing and sought individuals with GNB2 variants via international data-sharing initiatives. In silico modelling of the variants was assessed, along with multiple lines of evidence in keeping with American College of Medical Genetics and Genomics guidelines for interpretation of sequence variants. RESULTS: We identified 12 unrelated individuals with five de novo missense variants in GNB2, four of which are recurrent: p.(Ala73Thr), p.(Gly77Arg), p.(Lys89Glu) and p.(Lys89Thr). All individuals have DD/ID with variable dysmorphism and extraneurologic features. The variants are located at the universally conserved shared interface with the Gα subunit, which modelling suggests weaken this interaction. CONCLUSION: Missense variants in GNB2 cause a congenital neurodevelopmental disorder with variable syndromic features, broadening the spectrum of multisystem phenotypes associated with variants in genes encoding G-proteins.

Bezafibrate Improves Mitochondrial Fission and Function in DNM1L-Deficient Patient Cells.

Mitochondria are involved in many cellular processes and their main role is cellular energy production. They constantly undergo fission and fusion, and these counteracting processes are under strict balance. The cytosolic dynamin-related protein 1, Drp1, or dynamin-1-like protein (DNM1L) mediates mitochondrial and peroxisomal division. Defects in the DNM1L gene result in a complex neurodevelopmental disorder with heterogeneous symptoms affecting multiple organ systems. Currently there is no curative treatment available for this condition. We have previously described a patient with a de novo heterozygous c.1084G>A (p.G362S) DNM1L mutation and studied the effects of a small molecule, bezafibrate, on mitochondrial functions in this patient's fibroblasts compared to controls. Bezafibrate normalized growth on glucose-free medium, as well as ATP production and oxygen consumption. It improved mitochondrial morphology in the patient's fibroblasts, although causing a mild increase in ROS production at the same time. A human foreskin fibroblast cell line overexpressing the p.G362S mutation showed aberrant mitochondrial morphology, which normalized in the presence of bezafibrate. Further studies would be needed to show the consistency of the response to bezafibrate, possibly using fibroblasts from patients with different mutations in DNM1L, and this treatment should be confirmed in clinical trials. However, taking into account the favorable effects in our study, we suggest that bezafibrate could be offered as a treatment option for patients with certain DNM1L mutations.

De novo pathogenic DNM1L variant in a patient diagnosed with atypical hereditary sensory and autonomic neuropathy.

BACKGROUND: Profiling the entire genome at base pair resolution in a single test offers novel insights into disease by means of dissection of genetic contributors to phenotypic features. METHODS: We performed genome sequencing for a patient who presented with atypical hereditary sensory and autonomic neuropathy, severe epileptic encephalopathy, global developmental delay, and growth hormone deficiency. RESULTS: Assessment of the variants detected by mapped sequencing reads followed by Sanger confirmation revealed that the proband is a compound heterozygote for rare variants within RETREG1 (FAM134B), a gene associated with a recessive form of hereditary sensory and autonomic neuropathy, but not with epileptic encephalopathy or global developmental delay. Further analysis of the data also revealed a heterozygous missense variant in DNM1L, a gene previously implicated in an autosomal dominant encephalopathy, epilepsy, and global developmental delay and confirmed by Sanger sequencing to be a de novo variant not present in parental genomes. CONCLUSIONS: Our findings emphasize the importance of genome-wide sequencing in patients with a well-characterized genetic disease with atypical presentation. This approach reduces the potential for misdiagnoses.

TRPM1 Mutations are the Most Common Cause of Autosomal Recessive Congenital Stationary Night Blindness (CSNB) in the Palestinian and Israeli Populations.

Precise genetic and phenotypic characterization of congenital stationary night blindness (CSNB) patients is needed for future therapeutic interventions. The aim of this study was to estimate the prevalence of CSNB in our populations and to study clinical and genetic aspects of the autosomal recessive (AR) form of CSNB. This is a retrospective cohort study of Palestinian and Israeli CSNB patients harboring mutations in TRPM1 underwent comprehensive ocular examination. Genetic analysis was performed using homozygosity mapping and sequencing. 161 patients (from 76 families) were recruited for this study, leading to a prevalence of 1:6210 in the vicinity of Jerusalem, much higher than the worldwide prevalence. 61% of the families were consanguineous with AR inheritance pattern. Biallelic pathogenic TRPM1 mutations were identified in 36 families (72 patients). Two founder mutations explain the vast majority of cases: a nonsense mutation c.880A>T (p.Lys294*) identified in 22 Palestinian families and a large genomic deletion (36,445 bp) encompassing exons 2-7 of TRPM1 present in 13 Ashkenazi Jewish families. Most patients were myopic (with mean BCVA of 0.40 LogMAR) and all had absent rod responses in full field electroretinography. To the best of our knowledge, this is the largest report of a clinical and genetic analysis of patients affected with CSNB due to TRPM1 mutations.

Biallelic variants in AGTPBP1, involved in tubulin deglutamylation, are associated with cerebellar degeneration and motor neuropathy.

The ATP/GTP-Binding Protein 1 (AGTPBP1) gene (OMIM *606830) catalyzes deglutamylation of polyglutamylated proteins, and its deficiency manifests by cerebellar ataxia and peripheral neuropathy in mice and lower motor neuron-like disease in sheep. In the mutant mice, cerebellar atrophy due to Purkinje cell degeneration is observed, likely due to increased tubulin polyglutamylation in affected brain areas. We report two unrelated individuals who presented with early onset cerebellar atrophy, developmental arrest with progressive muscle weakness, and feeding and respiratory difficulties, accompanied by severe motor neuronopathy. Whole exome sequencing followed by segregation analysis in the families and cDNA studies revealed deleterious biallelic variants in the AGTPBP1 gene. We conclude that complete loss-of-function of AGTPBP1 in humans, just like in mice and sheep, is associated with cerebellar and motor neuron disease, reminiscent of Pontocerebellar Hypoplasia Type 1 (PCH1).

Role of a conserved glutamine in the function of voltage-gated Ca2+ channels revealed by a mutation in human CACNA1D.

Voltage-gated Cav Ca2+ channels play crucial roles in regulating gene transcription, neuronal excitability, and synaptic transmission. Natural or pathological variations in Cav channels have yielded rich insights into the molecular determinants controlling channel function. Here, we report the consequences of a natural, putatively disease-associated mutation in the CACNA1D gene encoding the pore-forming Cav1.3 α1 subunit. The mutation causes a substitution of a glutamine residue that is highly conserved in the extracellular S1-S2 loop of domain II in all Cav channels with a histidine and was identified by whole-exome sequencing of an individual with moderate hearing impairment, developmental delay, and epilepsy. When introduced into the rat Cav1.3 cDNA, Q558H significantly decreased the density of Ca2+ currents in transfected HEK293T cells. Gating current analyses and cell-surface biotinylation experiments suggested that the smaller current amplitudes caused by Q558H were because of decreased numbers of functional Cav1.3 channels at the cell surface. The substitution also produced more sustained Ca2+ currents by weakening voltage-dependent inactivation. When inserted into the corresponding locus of Cav2.1, the substitution had similar effects as in Cav1.3. However, the substitution introduced in Cav3.1 reduced current density, but had no effects on voltage-dependent inactivation. Our results reveal a critical extracellular determinant of current density for all Cav family members and of voltage-dependent inactivation of Cav1.3 and Cav2.1 channels.

Postnatal microcephaly and pain insensitivity due to a de novo heterozygous DNM1L mutation causing impaired mitochondrial fission and function.

An emerging class of mitochondrial disorders is caused by mutations in nuclear genes affecting mitochondrial dynamics and function. One of these is the DNM1L gene encoding the dynamin-related protein 1 (DRP1), which is pivotal in the mitochondrial fission process. Here, we describe a patient with a novel dominant-negative, de novo DNM1L mutation, which expands the clinical spectrum. The patient reported here exhibits a chronic neurological disorder, characterized by postnatal microcephaly, developmental delay, and pain insensitivity. Muscle biopsy disclosed decreased respiratory chain complex IV activity. Exome sequencing showed a de novo heterozygous c.1084G>A (p.G362S) mutation. Subsequent studies of patient skin fibroblasts showed markedly impaired mitochondrial fission and a partial respiratory chain defect while peroxisomal morphology remained intact. Human foreskin fibroblasts over-expressing the mutant DNM1L gene displayed aberrant mitochondrial morphology. © 2016 Wiley Periodicals, Inc.

Fe/S protein assembly gene IBA57 mutation causes hereditary spastic paraplegia.

OBJECTIVE: To present the clinical, molecular, and cell biological findings in a family with an autosomal recessive form of hereditary spastic paraplegia characterized by a combination of spastic paraplegia, optic atrophy, and peripheral neuropathy (SPOAN). METHODS: We used a combination of whole-genome linkage analysis and exome sequencing to map the disease locus and to identify the responsible gene. To analyze the physiologic consequences of the disease, we used biochemical and cell biological methods. RESULTS: Ten members of a highly consanguineous family manifested a childhood-onset SPOAN-like phenotype with slow progression into late adulthood. We mapped this disorder to a locus on chromosome 1q and identified a homozygous donor splice-site mutation in the IBA57 gene, previously implicated in 2 infants with lethal perinatal encephalomyopathy. This gene encodes the mitochondrial iron-sulfur (Fe/S) protein assembly factor IBA57. In addition to a severely decreased amount of normal IBA57 messenger RNA, a patient's cells expressed an aberrantly spliced messenger RNA with a premature stop codon. Lymphoblasts contained 10-fold-lower levels of wild-type, but no signs of truncated IBA57 protein. The decrease in functional IBA57 resulted in reduced levels and activities of several mitochondrial [4Fe-4S] proteins, including complexes I and II, while mitochondrial [2Fe-2S] proteins remained normal. CONCLUSIONS: Our findings reinforce the suggested specific function of IBA57 in mitochondrial [4Fe-4S] protein maturation and provide additional evidence for its role in human disease. The less decreased IBA57 protein level in this family explains phenotypic differences compared with the previously described lethal encephalomyopathy with no functional IBA57.

Hindbrain malformation and myoclonic seizures associated with a deleterious mutation in the INPP4A gene.

Hindbrain malformations with predominant cerebellar involvement have many causes including chromosomal disorders, specific genetic syndromes, and prenatal disruptions. The combination of a hindbrain malformation and myoclonic epilepsy is rare. Using exome sequencing in a consanguineous family, we identified a homozygous genomic deletion of 1770 bp within the INPP4A gene in a patient with myoclonic epilepsy, microcephaly, and atrophy of the inferior vermis and cerebellum. INPP4A participates in the excitatory glutamate signaling pathway and is essential for the degradation of phosphatidylinositol (3,4)-bisphosphate. Glutamatergic signaling is important for hindbrain development and is implicated in the pathogenesis of epilepsy, as well as excitotoxic cell death. Indeed, excessive glutamatergic stimulation was previously reported in INPP4A knockout mice. Our data adds a new etiology to the spectrum of hindbrain malformations in human, and when presented with myoclonic epilepsy may lead to the clinical suspicion of INPP4A defect. The present report further underscores the importance of phosphoinositides for the development of the inferior cerebellum and vermis.

KIF1A missense mutations in SPG30, an autosomal recessive spastic paraplegia: distinct phenotypes according to the nature of the mutations.

The hereditary spastic paraplegias (HSPs) are a clinically and genetically heterogeneous group of neurodegenerative diseases characterised by progressive spasticity in the lower limbs. The nosology of autosomal recessive forms is complex as most mapped loci have been identified in only one or a few families and account for only a small percentage of patients. We used next-generation sequencing focused on the SPG30 chromosomal region on chromosome 2q37.3 in two patients from the original linked family. In addition, wide genome scan and candidate gene analysis were performed in a second family of Palestinian origin. We identified a single homozygous mutation, p.R350G, that was found to cosegregate with the disease in the SPG30 kindred and was absent in 970 control chromosomes while affecting a strongly conserved amino acid at the end of the motor domain of KIF1A. Homozygosity and linkage mapping followed by mutation screening of KIF1A allowed us to identify a second mutation, p.A255V, in the second family. Comparison of the clinical features with the nature of the mutations of all reported KIF1A families, including those reported recently with hereditary sensory and autonomic neuropathy, suggests phenotype-genotype correlations that may help to understand the mechanisms involved in motor neuron degeneration. We have shown that mutations in the KIF1A gene are responsible for SPG30 in two autosomal recessive HSP families. In published families, the nature of the KIF1A mutations seems to be of good predictor of the underlying phenotype and vice versa.

Early-onset sensorineural hearing loss is a prominent feature of H syndrome.

This case report describes two patients with H syndrome, a multisystemic autosomal recessive disorder, caused by mutations in the SLC29A3 gene. It is characterized by cutaneous hyperpigmentation, camptodactyly or flexion contractures and other features, among them hearing loss. The two patients had hearing loss as their presenting symptom, and had mutations in SLC29A3, one of them a novel mutation. The aim of this paper is to increase awareness to this recently described disorder, and to emphasize that H syndrome should be included in the differential diagnosis of congenital or acquired syndromic hearing loss in children.

Increased risk for developmental delay in Saethre-Chotzen syndrome is associated with TWIST deletions: an improved strategy for TWIST mutation screening.

The majority of patients with Saethre-Chotzen syndrome have mutations in the TWIST gene, which codes for a basic helix-loop-helix transcription factor. Of the genetic alterations identified in TWIST, nonsense mutations, frameshifts secondary to small deletions or insertions, and large deletions implicate haploinsufficiency as the pathogenic mechanism. We identified three novel intragenic mutations and six deletions in our patients by using a new strategy to screen for TWIST mutations. We used polymerase chain reaction (PCR) amplification with subsequent sequencing to identify point mutations and small insertions or deletions in the coding region, and real-time PCR-based gene dosage analysis to identify large deletions encompassing the gene, with confirmation by microsatellite and fluorescence in situ hybridization (FISH) analyses. The size of the deletions can also be analyzed by using the gene dosage assay with "PCR walking" across the critical region. In 55 patients with features of Saethre-Chotzen syndrome, 11% were detected to have deletions by real-time gene dosage analysis. Two patients had a translocation or inversion at least 260 kb 3' of the gene, suggesting they had position-effect mutations. Of the 37 patients with classic features of Saethre-Chotzen syndrome, the overall detection rate for TWIST mutations was 68%. The risk for developmental delay in patients with deletions involving the TWIST gene is approximately 90% or eight times more common than in patients with intragenic mutations.