Pathological variants in TOP3A cause distinct disorders of mitochondrial and nuclear genome stability

Topoisomerase 3α (TOP3A) is an enzyme that removes torsional strain and interlinks between DNA molecules. TOP3A localises to both the nucleus and mitochondria, with the two isoforms playing specialised roles in DNA recombination and replication respectively. Pathogenic variants in TOP3A can cause a disorder similar to Bloom syndrome, which results from bi-allelic pathogenic variants in BLM, encoding a nuclear-binding partner of TOP3A. In this work, we describe 11 individuals from 9 families with an adult-onset mitochondrial disease resulting from bi-allelic TOP3A gene variants. The majority of patients have a consistent clinical phenotype characterised by bilateral ptosis, ophthalmoplegia, myopathy and axonal sensory-motor neuropathy. We present a comprehensive characterisation of the effect of TOP3A variants, from individuals with mitochondrial disease and Bloom-like syndrome, upon mtDNA maintenance and different aspects of enzyme function. Based on these results, we suggest a model whereby the overall severity of the TOP3A catalytic defect determines the clinical outcome, with milder variants causing adult-onset mitochondrial disease and more severe variants causing a Bloom-like syndrome with mitochondrial dysfunction in childhood

Molecular Mechanisms of Skewed X-Chromosome Inactivation in Female Hemophilia Patients—Lessons from Wide Genome Analyses

Introduction: Hemophilia A (HA) is an X-linked bleeding disorder caused by factor VIII (FVIII) deficiency or dysfunction due to F8 gene mutations. HA carriers are usually asymptomatic because their FVIII levels correspond to approximately half of the concentration found in healthy individuals. However, in rare cases, a carrier may exhibit symptoms of moderate to severe HA primarily due to skewed inactivation of her non-hemophilic X chromosome. Aim: The aim of the study was to investigate X-chromosome inactivation (XCI) patterns in HA carriers, with special emphasis on three karyotypically normal HA carriers presenting with moderate to severe HA phenotype due to skewed XCI, in an attempt to elucidate the molecular mechanism underlying skewed XCI in these symptomatic HA carriers. The study was based on the hypothesis that the presence of a pathogenic mutation on the non-hemophilic X chromosome is the cause of extreme inactivation of that X chromosome. Methods: XCI patterns were studied by PCR analysis of the CAG repeat region in the HUMARA gene. HA carriers that demonstrated skewed XCI were further studied by whole-exome sequencing (WES) followed by X chromosome-targeted bioinformatic analysis. Results: All three HA carriers presenting with the moderate to severe HA phenotype due to skewed XCI were found to carry pathogenic mutations on their non-hemophilic X chromosomes. Patient 1 was diagnosed with a frameshift mutation in the PGK1 gene that was associated with familial XCI skewing in three generations. Patient 2 was diagnosed with a missense mutation in the SYTL4 gene that was associated with familial XCI skewing in two generations. Patient 3 was diagnosed with a nonsense mutation in the NKAP gene that was associated with familial XCI skewing in two generations. Conclusion: Our results indicate that the main reason for skewed XCI in our female HA patients was negative selection against cells with a disadvantage caused by an additional deleterious mutation on the silenced X chromosome, thus complicating the phenotype of a monogenic X-linked disease. Based on our study, we are currently offering the X inactivation test to symptomatic hemophilia carriers and plan to expand this approach to symptomatic carriers of other X-linked diseases, which can be further used in pregnancy planning

Progressive Pseudorheumatoid Dysplasia resolved by whole exome sequencing: a novel mutation in WISP3 and review of the literature

Abstract Background Progressive pseudorheumatoid dysplasia (PPRD) is a rare autosomal-recessive, non-inflammatory arthropathy, shown to be caused by mutations in the WNT1-inducible signaling pathway protein 3 (WISP3) gene. Although several hundred cases were reported worldwide, the diagnosis remains challenging. Subsequently, the syndrome is often unrecognized and misdiagnosed (for instance, as Juvenile Idiopathic Arthritis), leading to unnecessary procedures and treatments. The objective of the current study was to identify the molecular basis in a family with PPRD and describe their phenotype and course of illness. Patients and methods We present here a multiply affected consanguineous family of Iraqi-Jewish descent with PPRD. The proband, a 6.5 years old girl, presented with bilateral symmetric bony enlargements of the 1st interphalangeal joints of the hands, without signs of synovitis. Molecular analysis of the family was pursued using Whole Exome Sequencing (WES) and homozygosity mapping. Results WES analysis brought to the identification of a novel homozygous missense mutation (c.257G > T, p.C86F) in the WISP3 gene. Following this diagnosis, an additional 53 years old affected family member was found to harbor the mutation. Two other individuals in the family were reported to have had similar involvement however both had died of unrelated causes. Conclusion The reported family underscores the importance of recognition of this unique skeletal dysplasia by clinicians, and especially by pediatric rheumatologists and orthopedic surgeons

Whole exome sequencing (WES) approach for diagnosing primary immunodeficiencies (PIDs) in a highly consanguineous community

Primary immunodeficiencies (PIDs) are a heterogeneous group of monogenic inborn errors of immunity. The genetic causes of these diseases can be identified using whole exome sequencing (WES). Here, DNA samples from 106 patients with a clinical suspicion of PID were subjected to WES in order to test the diagnostic yield of this test in a highly consanguineous community. A likely genetic diagnosis was achieved in 70% of patients. Several factors were considered to possibly influence the diagnostic rate of WES among our cohort including early age, presence of consanguinity, family history suggestive of PID, the number of family members who underwent WES and the clinical phenotype of the patient. The highest diagnostic rate was in patients with combined immunodeficiency or with a syndrome. Notably, WES findings altered the clinical management in 39% (41/106) of patients in our cohort. Our findings support the use of WES as an important diagnostic tool in patients with suspected PID, especially in highly consanguineous communities

Mitochondrial Complex III Deficiency Associated with a Homozygous Mutation in UQCRQ

A consanguineous Israeli Bedouin kindred presented with an autosomal-recessive nonlethal phenotype of severe psychomotor retardation and extrapyramidal signs, dystonia, athetosis and ataxia, mild axial hypotonia, and marked global dementia with defects in verbal and expressive communication skills. Metabolic workup was normal except for mildly elevated blood lactate levels. Brain magnetic resonance imaging (MRI) showed increased density in the putamen, with decreased density and size of the caudate and lentiform nuclei. Reduced activity specifically of mitochondrial complex III and variable decrease in complex I activity were evident in muscle biopsies. Homozygosity of affected individuals to UQCRB and to BCSIL, previously associated with isolated complex III deficiency, was ruled out. Genome-wide linkage analysis identified a homozygosity locus of approximately 9 cM on chromosome 5q31 that was further narrowed down to 2.14 cM, harboring 30 genes (logarithm of the odds [LOD] score 8.82 at θ = 0). All 30 genes were sequenced, revealing a single missense (p.Ser45Phe) mutation in UQCRQ (encoding ubiquinol-cytochrome c reductase, complex III subunit VII, 9.5 kDa), one of the ten nuclear genes encoding proteins of mitochondrial complex III

Maternally Inherited Birk Barel Mental Retardation Dysmorphism Syndrome Caused by a Mutation in the Genomically Imprinted Potassium Channel KCNK9

We describe a maternally transmitted genomic-imprinting syndrome of mental retardation, hypotonia, and unique dysmorphism with elongated face. We mapped the disease-associated locus to ∼7.27 Mb on chromosome 8q24 and demonstrated that the disease is caused by a missense mutation in the maternal copy of KCNK9 within this locus. KCNK9 is maternally transmitted (imprinted with paternal silencing) and encodes K2P9.1, a member of the two pore-domain potassium channel (K2P) subfamily. The mutation fully abolishes the channel's currents—both when functioning as a homodimer or as a heterodimer with K2P3.1

Bi-allelic Variants in RALGAPA1 Cause Profound Neurodevelopmental Disability, Muscular Hypotonia, Infantile Spasms, and Feeding Abnormalities

Ral (Ras-like) GTPases play an important role in the control of cell migration and have been implicated in Ras-mediated tumorigenicity. Recently, variants in RALA were also described as a cause of intellectual disability and developmental delay, indicating the relevance of this pathway to neuropediatric diseases. Here, we report the identification of bi-allelic variants in RALGAPA1 (encoding Ral GTPase activating protein catalytic alpha subunit 1) in four unrelated individuals with profound neurodevelopmental disability, muscular hypotonia, feeding abnormalities, recurrent fever episodes, and infantile spasms . Dysplasia of corpus callosum with focal thinning of the posterior part and characteristic facial features appeared to be unifying findings. RalGAPA1 was absent in the fibroblasts derived from two affected individuals suggesting a loss-of-function effect of the RALGAPA1 variants. Consequently, RalA activity was increased in these cell lines, which is in keeping with the idea that RalGAPA1 deficiency causes a constitutive activation of RalA. Additionally, levels of RalGAPB, a scaffolding subunit of the RalGAP complex, were dramatically reduced, indicating a dysfunctional RalGAP complex. Moreover, RalGAPA1 deficiency clearly increased cell-surface levels of lipid raft components in detached fibroblasts, which might indicate that anchorage-dependence of cell growth signaling is disturbed. Our findings indicate that the dysregulation of the RalA pathway has an important impact on neuronal function and brain development. In light of the partially overlapping phenotype between RALA- and RALGAPA1-associated diseases, it appears likely that dysregulation of the RalA signaling pathway leads to a distinct group of genetic syndromes that we suggest could be named RALopathies

Table1_Vici syndrome in Israel: Clinical and molecular insights.docx

Introduction: Vici Syndrome is a rare, severe, neurodevelopmental/neurodegenerative disorder with multi-systemic manifestations presenting in infancy. It is mainly characterized by global developmental delay, seizures, agenesis of the corpus callosum, hair and skin hypopigmentation, bilateral cataract, and varying degrees of immunodeficiency, among other features. Vici Syndrome is caused by biallelic pathogenic variants in EPG5, resulting in impaired autophagy. Thus far, the condition has been reported in less than a hundred individuals.Objective and Methods: We aimed to characterize the clinical and molecular findings in individuals harboring biallelic EPG5 variants, recruited from four medical centers in Israel. Furthermore, we aimed to utilize a machine learning-based tool to assess facial features of Vici syndrome.Results: Eleven cases of Vici Syndrome from five unrelated families, one of which was diagnosed prenatally with subsequent termination of pregnancy, were recruited. A total of five disease causing variants were detected in EPG5: two novel: c.2554-5A>G and c.1461delC; and 3 previously reported: c.3447G>A, c.5993C>G, and c.1007A>G, the latter previously identified in several patients of Ashkenazi-Jewish (AJ) descent. Amongst 140,491 individuals screened by the Dor Yeshorim Program, we show that the c.1007A>G variant has an overall carrier frequency of 0.45% (1 in 224) among AJ individuals. Finally, based on two-dimensional facial photographs of individuals with Vici syndrome (n = 19), a composite facial mask was created using the DeepGestalt algorithm, illustrating facial features typical of this disorder.Conclusion: We report on ten children and one fetus from five unrelated families, affected with Vici syndrome, and describe prenatal and postnatal characteristics. Our findings contribute to the current knowledge regarding the molecular basis and phenotypic features of this rare syndrome. Additionally, the deep learning-based facial gestalt adds to the clinician’s diagnostic toolbox and may aid in facilitating identification of affected individuals.</p

DataSheet1_Vici syndrome in Israel: Clinical and molecular insights.docx

Introduction: Vici Syndrome is a rare, severe, neurodevelopmental/neurodegenerative disorder with multi-systemic manifestations presenting in infancy. It is mainly characterized by global developmental delay, seizures, agenesis of the corpus callosum, hair and skin hypopigmentation, bilateral cataract, and varying degrees of immunodeficiency, among other features. Vici Syndrome is caused by biallelic pathogenic variants in EPG5, resulting in impaired autophagy. Thus far, the condition has been reported in less than a hundred individuals.Objective and Methods: We aimed to characterize the clinical and molecular findings in individuals harboring biallelic EPG5 variants, recruited from four medical centers in Israel. Furthermore, we aimed to utilize a machine learning-based tool to assess facial features of Vici syndrome.Results: Eleven cases of Vici Syndrome from five unrelated families, one of which was diagnosed prenatally with subsequent termination of pregnancy, were recruited. A total of five disease causing variants were detected in EPG5: two novel: c.2554-5A>G and c.1461delC; and 3 previously reported: c.3447G>A, c.5993C>G, and c.1007A>G, the latter previously identified in several patients of Ashkenazi-Jewish (AJ) descent. Amongst 140,491 individuals screened by the Dor Yeshorim Program, we show that the c.1007A>G variant has an overall carrier frequency of 0.45% (1 in 224) among AJ individuals. Finally, based on two-dimensional facial photographs of individuals with Vici syndrome (n = 19), a composite facial mask was created using the DeepGestalt algorithm, illustrating facial features typical of this disorder.Conclusion: We report on ten children and one fetus from five unrelated families, affected with Vici syndrome, and describe prenatal and postnatal characteristics. Our findings contribute to the current knowledge regarding the molecular basis and phenotypic features of this rare syndrome. Additionally, the deep learning-based facial gestalt adds to the clinician’s diagnostic toolbox and may aid in facilitating identification of affected individuals.</p

Image1_Vici syndrome in Israel: Clinical and molecular insights.PNG

Introduction: Vici Syndrome is a rare, severe, neurodevelopmental/neurodegenerative disorder with multi-systemic manifestations presenting in infancy. It is mainly characterized by global developmental delay, seizures, agenesis of the corpus callosum, hair and skin hypopigmentation, bilateral cataract, and varying degrees of immunodeficiency, among other features. Vici Syndrome is caused by biallelic pathogenic variants in EPG5, resulting in impaired autophagy. Thus far, the condition has been reported in less than a hundred individuals.Objective and Methods: We aimed to characterize the clinical and molecular findings in individuals harboring biallelic EPG5 variants, recruited from four medical centers in Israel. Furthermore, we aimed to utilize a machine learning-based tool to assess facial features of Vici syndrome.Results: Eleven cases of Vici Syndrome from five unrelated families, one of which was diagnosed prenatally with subsequent termination of pregnancy, were recruited. A total of five disease causing variants were detected in EPG5: two novel: c.2554-5A>G and c.1461delC; and 3 previously reported: c.3447G>A, c.5993C>G, and c.1007A>G, the latter previously identified in several patients of Ashkenazi-Jewish (AJ) descent. Amongst 140,491 individuals screened by the Dor Yeshorim Program, we show that the c.1007A>G variant has an overall carrier frequency of 0.45% (1 in 224) among AJ individuals. Finally, based on two-dimensional facial photographs of individuals with Vici syndrome (n = 19), a composite facial mask was created using the DeepGestalt algorithm, illustrating facial features typical of this disorder.Conclusion: We report on ten children and one fetus from five unrelated families, affected with Vici syndrome, and describe prenatal and postnatal characteristics. Our findings contribute to the current knowledge regarding the molecular basis and phenotypic features of this rare syndrome. Additionally, the deep learning-based facial gestalt adds to the clinician’s diagnostic toolbox and may aid in facilitating identification of affected individuals.</p