RESUMEN
Non-centrosomal microtubules are essential cytoskeletal filaments that are important for neurite formation, axonal transport, and neuronal migration. They require stabilization by microtubule minus-end-targeting proteins including the CAMSAP family of molecules. Using exome sequencing on samples from five unrelated families, we show that bi-allelic CAMSAP1 loss-of-function variants cause a clinically recognizable, syndromic neuronal migration disorder. The cardinal clinical features of the syndrome include a characteristic craniofacial appearance, primary microcephaly, severe neurodevelopmental delay, cortical visual impairment, and seizures. The neuroradiological phenotype comprises a highly recognizable combination of classic lissencephaly with a posterior more severe than anterior gradient similar to PAFAH1B1(LIS1)-related lissencephaly and severe hypoplasia or absence of the corpus callosum; dysplasia of the basal ganglia, hippocampus, and midbrain; and cerebellar hypodysplasia, similar to the tubulinopathies, a group of monogenic tubulin-associated disorders of cortical dysgenesis. Neural cell rosette lineages derived from affected individuals displayed findings consistent with these phenotypes, including abnormal morphology, decreased cell proliferation, and neuronal differentiation. Camsap1-null mice displayed increased perinatal mortality, and RNAScope studies identified high expression levels in the brain throughout neurogenesis and in facial structures, consistent with the mouse and human neurodevelopmental and craniofacial phenotypes. Together our findings confirm a fundamental role of CAMSAP1 in neuronal migration and brain development and define bi-allelic variants as a cause of a clinically distinct neurodevelopmental disorder in humans and mice.
Asunto(s)
Lisencefalias Clásicas y Heterotopias Subcorticales en Banda , Lisencefalia , Malformaciones del Sistema Nervioso , Humanos , Animales , Ratones , Lisencefalia/genética , Alelos , Tubulina (Proteína)/genética , Fenotipo , Malformaciones del Sistema Nervioso/genética , Lisencefalias Clásicas y Heterotopias Subcorticales en Banda/genética , Ratones Noqueados , Proteínas Asociadas a Microtúbulos/genéticaRESUMEN
The highly evolutionarily conserved transport protein particle (TRAPP) complexes (TRAPP II and III) perform fundamental roles in subcellular trafficking pathways. Here we identified biallelic variants in TRAPPC10, a component of the TRAPP II complex, in individuals with a severe microcephalic neurodevelopmental disorder. Molecular studies revealed a weakened interaction between mutant TRAPPC10 and its putative adaptor protein TRAPPC2L. Studies of patient lymphoblastoid cells revealed an absence of TRAPPC10 alongside a concomitant absence of TRAPPC9, another key TRAPP II complex component associated with a clinically overlapping neurodevelopmental disorder. The TRAPPC9/10 reduction phenotype was recapitulated in TRAPPC10-/- knockout cells, which also displayed a membrane trafficking defect. Notably, both the reduction in TRAPPC9 levels and the trafficking defect in these cells could be rescued by wild type but not mutant TRAPPC10 gene constructs. Moreover, studies of Trappc10-/- knockout mice revealed neuroanatomical brain defects and microcephaly, paralleling findings seen in the human condition as well as in a Trappc9-/- mouse model. Together these studies confirm autosomal recessive TRAPPC10 variants as a cause of human disease and define TRAPP-mediated pathomolecular outcomes of importance to TRAPPC9 and TRAPPC10 mediated neurodevelopmental disorders in humans and mice.
Asunto(s)
Microcefalia , Trastornos del Neurodesarrollo , Animales , Humanos , Ratones , Microcefalia/genética , Trastornos del Neurodesarrollo/genética , FenotipoRESUMEN
Due to the majority of currently available genome data deriving from individuals of European ancestry, the clinical interpretation of genomic variants in individuals from diverse ethnic backgrounds remains a major diagnostic challenge. Here, we investigated the genetic cause of a complex neurodevelopmental phenotype in two Palestinian siblings. Whole exome sequencing identified a homozygous missense TECPR2 variant (Chr14(GRCh38):g.102425085G>A; NM_014844.5:c.745G>A, p.(Gly249Arg)) absent in gnomAD, segregating appropriately with the inheritance pattern in the family. Variant assessment with in silico pathogenicity prediction and protein modeling tools alongside population database frequencies led to classification as a variant of uncertain significance. As pathogenic TECPR2 variants are associated with hereditary sensory and autonomic neuropathy with intellectual disability, we reviewed previously published candidate TECPR2 missense variants to clarify clinical outcomes and variant classification using current approved guidelines, classifying a number of published variants as of uncertain significance. This work highlights genomic healthcare inequalities and the challenges in interpreting rare genetic variants in populations underrepresented in genomic databases. It also improves understanding of the clinical and genetic spectrum of TECPR2-related neuropathy and contributes to addressing genomic data disparity and inequalities of the genomic architecture in Palestinian populations.
Asunto(s)
Secuenciación del Exoma , Neuropatías Hereditarias Sensoriales y Autónomas , Mutación Missense , Linaje , Hermanos , Humanos , Masculino , Femenino , Mutación Missense/genética , Neuropatías Hereditarias Sensoriales y Autónomas/genética , Neuropatías Hereditarias Sensoriales y Autónomas/patología , Neuropatías Hereditarias Sensoriales y Autónomas/diagnóstico , Fenotipo , Niño , Árabes/genética , Predisposición Genética a la Enfermedad , HomocigotoRESUMEN
SLC4A10 is a plasma-membrane bound transporter that utilizes the Na+ gradient to drive cellular HCO3- uptake, thus mediating acid extrusion. In the mammalian brain, SLC4A10 is expressed in principal neurons and interneurons, as well as in epithelial cells of the choroid plexus, the organ regulating the production of CSF. Using next generation sequencing on samples from five unrelated families encompassing nine affected individuals, we show that biallelic SLC4A10 loss-of-function variants cause a clinically recognizable neurodevelopmental disorder in humans. The cardinal clinical features of the condition include hypotonia in infancy, delayed psychomotor development across all domains and intellectual impairment. Affected individuals commonly display traits associated with autistic spectrum disorder including anxiety, hyperactivity and stereotyped movements. In two cases isolated episodes of seizures were reported in the first few years of life, and a further affected child displayed bitemporal epileptogenic discharges on EEG without overt clinical seizures. While occipitofrontal circumference was reported to be normal at birth, progressive postnatal microcephaly evolved in 7 out of 10 affected individuals. Neuroradiological features included a relative preservation of brain volume compared to occipitofrontal circumference, characteristic narrow sometimes 'slit-like' lateral ventricles and corpus callosum abnormalities. Slc4a10 -/- mice, deficient for SLC4A10, also display small lateral brain ventricles and mild behavioural abnormalities including delayed habituation and alterations in the two-object novel object recognition task. Collapsed brain ventricles in both Slc4a10-/- mice and affected individuals suggest an important role of SLC4A10 in the production of the CSF. However, it is notable that despite diverse roles of the CSF in the developing and adult brain, the cortex of Slc4a10-/- mice appears grossly intact. Co-staining with synaptic markers revealed that in neurons, SLC4A10 localizes to inhibitory, but not excitatory, presynapses. These findings are supported by our functional studies, which show the release of the inhibitory neurotransmitter GABA is compromised in Slc4a10-/- mice, while the release of the excitatory neurotransmitter glutamate is preserved. Manipulation of intracellular pH partially rescues GABA release. Together our studies define a novel neurodevelopmental disorder associated with biallelic pathogenic variants in SLC4A10 and highlight the importance of further analyses of the consequences of SLC4A10 loss-of-function for brain development, synaptic transmission and network properties.
Asunto(s)
Convulsiones , Simportadores de Sodio-Bicarbonato , Niño , Ratones , Humanos , Animales , Simportadores de Sodio-Bicarbonato/genética , Simportadores de Sodio-Bicarbonato/metabolismo , Convulsiones/genética , Mutación/genética , Neurotransmisores , Ácido gamma-Aminobutírico/genética , Mamíferos/metabolismo , Antiportadores de Cloruro-Bicarbonato/genética , Antiportadores de Cloruro-Bicarbonato/metabolismoRESUMEN
KPTN-related disorder is an autosomal recessive disorder associated with germline variants in KPTN (previously known as kaptin), a component of the mTOR regulatory complex KICSTOR. To gain further insights into the pathogenesis of KPTN-related disorder, we analysed mouse knockout and human stem cell KPTN loss-of-function models. Kptn -/- mice display many of the key KPTN-related disorder phenotypes, including brain overgrowth, behavioural abnormalities, and cognitive deficits. By assessment of affected individuals, we have identified widespread cognitive deficits (n = 6) and postnatal onset of brain overgrowth (n = 19). By analysing head size data from their parents (n = 24), we have identified a previously unrecognized KPTN dosage-sensitivity, resulting in increased head circumference in heterozygous carriers of pathogenic KPTN variants. Molecular and structural analysis of Kptn-/- mice revealed pathological changes, including differences in brain size, shape and cell numbers primarily due to abnormal postnatal brain development. Both the mouse and differentiated induced pluripotent stem cell models of the disorder display transcriptional and biochemical evidence for altered mTOR pathway signalling, supporting the role of KPTN in regulating mTORC1. By treatment in our KPTN mouse model, we found that the increased mTOR signalling downstream of KPTN is rapamycin sensitive, highlighting possible therapeutic avenues with currently available mTOR inhibitors. These findings place KPTN-related disorder in the broader group of mTORC1-related disorders affecting brain structure, cognitive function and network integrity.
Asunto(s)
Transducción de Señal , Serina-Treonina Quinasas TOR , Humanos , Animales , Ratones , Transducción de Señal/genética , Serina-Treonina Quinasas TOR/metabolismo , Encéfalo/metabolismo , Diana Mecanicista del Complejo 1 de la Rapamicina/metabolismo , Cognición , Proteínas de Microfilamentos/genéticaRESUMEN
OBJECTIVES: In October 2020, rapid prenatal exome sequencing (pES) was introduced into routine National Health Service (NHS) care in England. This study aimed to explore parent experiences and their information and support needs from the perspective of parents offered pES and of health professionals involved in its delivery. METHODS: In this qualitative study, semi-structured interviews were conducted with 42 women and 6 male partners and 63 fetal medicine and genetic health professionals. Interviews were transcribed verbatim and analysed using thematic analysis. RESULTS: Overall views about pES were positive and parents were grateful to be offered the test. Highlighted benefits of pES included the value of the additional information for pregnancy management and planning for future pregnancies. An anxious wait for results was common, often associated with the need to make decisions near to 24 weeks in pregnancy when there are legal restrictions for late termination. Descriptions of dealing with uncertainty were also common, even when results had been returned. Many parents described pES results as informing decision-making around whether or not to terminate pregnancy. Some professionals were concerned that a non-informative result could be overly reassuring and highlighted that careful counselling was needed to ensure parents have a good understanding of what the result means for their pregnancy. Emotional support from professionals was valued; however, some parents felt that post-test support was lacking. CONCLUSION: Parents and professionals welcomed the introduction of pES. Results inform parents' decision-making around the termination of pregnancy. When there are no diagnostic findings or uncertain findings from pES, personalised counselling that considers scans and other tests are crucial. Directing parents to reliable online sources of information and providing emotional support throughout could improve their experiences of care.
Asunto(s)
Padres , Medicina Estatal , Embarazo , Humanos , Masculino , Femenino , Secuenciación del Exoma , Padres/psicología , Inglaterra , Consejo , Investigación CualitativaRESUMEN
BACKGROUND: Oculocutaneous albinism (OCA) is a genetically heterogeneous condition that is associated with reduced or absent melanin pigment in the skin, hair, and eyes, resulting in reduced vision, high sensitivity to light, and rapid and uncontrolled eye movements. To date, seventeen genes have been associated with OCA including syndromic and non-syndromic forms of the condition. METHODS: Whole exome sequencing (WES) was performed to identify pathogenic variants in nine Pakistani families with OCA, with validation and segregation of candidate variants performed using Sanger sequencing. Furthermore, the pathogenicity of the identified variants was assessed using various in-silico tools and 3D protein structural analysis software. RESULTS: WES identified biallelic variants in three genes explaining the OCA in these families, including four variants in TYR, three in OCA2, and two in HPS1, including two novel variants c.667C > T: p.(Gln223*) in TYR, and c.2009 T > C: p.(Leu670Pro) in HPS1. CONCLUSIONS: Overall, this study adds further knowledge of the genetic basis of OCA in Pakistani communities and facilitates improved management and counselling services for families suffering from severe genetic diseases in Pakistan.
Asunto(s)
Albinismo Oculocutáneo , Secuenciación del Exoma , Síndrome de Hermanski-Pudlak , Mutación , Linaje , Humanos , Albinismo Oculocutáneo/genética , Pakistán , Femenino , Masculino , Síndrome de Hermanski-Pudlak/genética , Niño , Adulto , Adolescente , Análisis Mutacional de ADN , Monofenol Monooxigenasa/genética , Preescolar , Proteínas de Transporte de Membrana/genética , Adulto Joven , Proteínas de la MembranaRESUMEN
SNIP1 (Smad nuclear interacting protein 1) is a widely expressed transcriptional suppressor of the TGF-ß signal-transduction pathway which plays a key role in human spliceosome function. Here, we describe extensive genetic studies and clinical findings of a complex inherited neurodevelopmental disorder in 35 individuals associated with a SNIP1 NM_024700.4:c.1097A>G, p.(Glu366Gly) variant, present at high frequency in the Amish community. The cardinal clinical features of the condition include hypotonia, global developmental delay, intellectual disability, seizures, and a characteristic craniofacial appearance. Our gene transcript studies in affected individuals define altered gene expression profiles of a number of molecules with well-defined neurodevelopmental and neuropathological roles, potentially explaining clinical outcomes. Together these data confirm this SNIP1 gene variant as a cause of an autosomal recessive complex neurodevelopmental disorder and provide important insight into the molecular roles of SNIP1, which likely explain the cardinal clinical outcomes in affected individuals, defining potential therapeutic avenues for future research.
Asunto(s)
Alelos , Amish/genética , Trastornos del Neurodesarrollo/genética , Proteínas de Unión al ARN/genética , Expresión Génica/genética , Genes Recesivos , HumanosRESUMEN
The hereditary spastic paraplegias (HSP) are among the most genetically diverse of all Mendelian disorders. They comprise a large group of neurodegenerative diseases that may be divided into 'pure HSP' in forms of the disease primarily entailing progressive lower-limb weakness and spasticity, and 'complex HSP' when these features are accompanied by other neurological (or non-neurological) clinical signs. Here, we identified biallelic variants in the transmembrane protein 63C (TMEM63C) gene, encoding a predicted osmosensitive calcium-permeable cation channel, in individuals with hereditary spastic paraplegias associated with mild intellectual disability in some, but not all cases. Biochemical and microscopy analyses revealed that TMEM63C is an endoplasmic reticulum-localized protein, which is particularly enriched at mitochondria-endoplasmic reticulum contact sites. Functional in cellula studies indicate a role for TMEM63C in regulating both endoplasmic reticulum and mitochondrial morphologies. Together, these findings identify autosomal recessive TMEM63C variants as a cause of pure and complex HSP and add to the growing evidence of a fundamental pathomolecular role of perturbed mitochondrial-endoplasmic reticulum dynamics in motor neurone degenerative diseases.
Asunto(s)
Canales de Calcio , Mitocondrias , Paraplejía Espástica Hereditaria , Canales de Calcio/genética , Retículo Endoplásmico/genética , Humanos , Mitocondrias/patología , Mutación , Paraplejía Espástica Hereditaria/genéticaRESUMEN
HERC2 gene encodes an E3 ubiquitin ligase involved in several cellular processes by regulating the ubiquitylation of different protein substrates. Biallelic pathogenic sequence variants in the HERC2 gene are associated with HERC2 Angelman-like syndrome. In pathogenic HERC2 variants, complete absence or marked reduction in HERC2 protein levels are observed. The most common pathological variant, c.1781C > T (p.Pro594Leu), encodes an unstable HERC2 protein. A better understanding of how pathologic HERC2 variants affect intracellular signalling may aid definition of potential new therapies for these disorders. For this purpose, we studied patient-derived cells with the HERC2 Pro594Leu variant. We observed alteration of mitogen-activated protein kinase signalling pathways, reflected by increased levels of C-RAF protein and p38 phosphorylation. HERC2 knockdown experiments reproduced the same effects in other human and mouse cells. Moreover, we demonstrated that HERC2 and RAF proteins form molecular complexes, pull-down and proteomic experiments showed that HERC2 regulates C-RAF ubiquitylation and we found out that the p38 activation due to HERC2 depletion occurs in a RAF/MKK3-dependent manner. The displayed cellular response was that patient-derived and other human cells with HERC2 deficiency showed higher resistance to oxidative stress with an increase in the master regulator of the antioxidant response NRF2 and its target genes. This resistance was independent of p53 and abolished by RAF or p38 inhibitors. Altogether, these findings identify the activation of C-RAF/MKK3/p38 signalling pathway in HERC2 Angelman-like syndrome and highlight the inhibition of RAF activity as a potential therapeutic option for individuals affected with these rare diseases.
Asunto(s)
Proteínas Proto-Oncogénicas c-raf , Proteína p53 Supresora de Tumor , Animales , Antioxidantes/metabolismo , Factores de Intercambio de Guanina Nucleótido/genética , Factores de Intercambio de Guanina Nucleótido/metabolismo , Humanos , Ratones , Proteínas Quinasas Activadas por Mitógenos/metabolismo , Factor 2 Relacionado con NF-E2/genética , Factor 2 Relacionado con NF-E2/metabolismo , Estrés Oxidativo , Proteínas Serina-Treonina Quinasas , Proteómica , Proteínas Proto-Oncogénicas/metabolismo , Proteínas Proto-Oncogénicas c-raf/metabolismo , Proteína p53 Supresora de Tumor/metabolismo , Ubiquitina-Proteína Ligasas/genética , Ubiquitina-Proteína Ligasas/metabolismo , Proteínas Quinasas p38 Activadas por Mitógenos/genética , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismoRESUMEN
BACKGROUND: Retinitis Pigmentosa (RP) is a clinically and genetically progressive retinal dystrophy associated with severe visual impairments and sometimes blindness, the most common syndromic form of which is Usher syndrome (USH). This study aimed to further increase understanding of the spectrum of RP in the Khyber Pakhtunkhwa region of Pakistan. METHODOLOGY: Four consanguineous families of Pashtun ethnic group were investigated which were referred by the local collaborating ophthalmologists. In total 42 individuals in four families were recruited and investigated using whole exome and dideoxy sequencing. Among them, 20 were affected individuals including 6 in both family 1 and 2, 5 in family 3 and 3 in family 4. RESULT: Pathogenic gene variants were identified in all four families, including two in cone dystrophy and RP genes in the same family (PDE6C; c.480delG, p.Asn161ThrfsTer33 and TULP1; c.238 C > T, p.Gln80Ter) with double-homozygous individuals presenting with more severe disease. Other pathogenic variants were identified in MERTK (c.2194C > T, p.Arg732Ter), RHO (c.448G > A, p.Glu150Lys) associated with non-syndromic RP, and MYO7A (c.487G > A, p.Gly163Arg) associated with USH. In addition, the reported variants were of clinical significance as the PDE6C variant was detected novel, whereas TULP1, MERTK, and MYO7A variants were detected rare and first time found segregating with retinal dystrophies in Pakistani consanguineous families. CONCLUSIONS: This study increases knowledge of the genetic basis of retinal dystrophies in families from Pakistan providing information important for genetic testing and diagnostic provision particularly from the Khyber Pakhtunkhwa region.
Asunto(s)
Distrofias Retinianas , Retinitis Pigmentosa , Humanos , Consanguinidad , Pakistán , Tirosina Quinasa c-Mer/genética , Mutación , Distrofias Retinianas/genética , Retinitis Pigmentosa/diagnóstico , Linaje , Análisis Mutacional de ADNRESUMEN
Many studies have demonstrated the clinical utility and importance of epilepsy gene panel testing to confirm the specific aetiology of disease, enable appropriate therapeutic interventions, and inform accurate family counselling. Previously, SCN9A gene variants, in particular a c.1921A>T p.(Asn641Tyr) substitution, have been identified as a likely autosomal dominant cause of febrile seizures/febrile seizures plus and other monogenic seizure phenotypes indistinguishable from those associated with SCN1A, leading to inclusion of SCN9A on epilepsy gene testing panels. Here we present serendipitous findings of genetic studies that identify the SCN9A c.1921A>T p.(Asn641Tyr) variant at high frequency in the Amish community in the absence of such seizure phenotypes. Together with findings in UK Biobank these data refute an association of SCN9A with epilepsy, which has important clinical diagnostic implications.
Asunto(s)
Errores Diagnósticos/prevención & control , Epilepsia/diagnóstico , Pruebas Genéticas/métodos , Canal de Sodio Activado por Voltaje NAV1.7/genética , Sustitución de Aminoácidos , Amish/genética , Niño , Preescolar , Epilepsia/genética , Femenino , Estudios de Seguimiento , Frecuencia de los Genes , Predisposición Genética a la Enfermedad , Heterocigoto , Humanos , Lactante , Masculino , Mutación , Linaje , Polimorfismo de Nucleótido Simple , Secuenciación del Exoma , WisconsinRESUMEN
Germline mutations in fundamental epigenetic regulatory molecules including DNA methyltransferase 3 alpha (DNMT3A) are commonly associated with growth disorders, whereas somatic mutations are often associated with malignancy. We profiled genome-wide DNA methylation patterns in DNMT3A c.2312G > A; p.(Arg771Gln) carriers in a large Amish sibship with Tatton-Brown-Rahman syndrome (TBRS), their mosaic father, and 15 TBRS patients with distinct pathogenic de novo DNMT3A variants. This defined widespread DNA hypomethylation at specific genomic sites enriched at locations annotated as genes involved in morphogenesis, development, differentiation, and malignancy predisposition pathways. TBRS patients also displayed highly accelerated DNA methylation aging. These findings were most marked in a carrier of the AML-associated driver mutation p.Arg882Cys. Our studies additionally defined phenotype-related accelerated and decelerated epigenetic aging in two histone methyltransferase disorders: NSD1 Sotos syndrome overgrowth disorder and KMT2D Kabuki syndrome growth impairment. Together, our findings provide fundamental new insights into aberrant epigenetic mechanisms, the role of epigenetic machinery maintenance, and determinants of biological aging in these growth disorders.
Asunto(s)
Envejecimiento/genética , ADN (Citosina-5-)-Metiltransferasas/genética , Epigénesis Genética , Trastornos del Crecimiento/genética , Mutación , Anomalías Múltiples/genética , Adolescente , Adulto , Amish/genética , Niño , Metilación de ADN , ADN Metiltransferasa 3A , Cara/anomalías , Enfermedades Hematológicas/genética , Humanos , Discapacidad Intelectual/genética , Leucemia Mieloide Aguda/genética , Masculino , Metiltransferasas , Morfogénesis/genética , Síndrome , Enfermedades Vestibulares/genética , Adulto JovenRESUMEN
PURPOSE: The clinical spectrum of motile ciliopathies includes laterality defects, hydrocephalus, and infertility as well as primary ciliary dyskinesia when impaired mucociliary clearance results in otosinopulmonary disease. Importantly, approximately 30% of patients with primary ciliary dyskinesia lack a genetic diagnosis. METHODS: Clinical, genomic, biochemical, and functional studies were performed alongside in vivo modeling of DAW1 variants. RESULTS: In this study, we identified biallelic DAW1 variants associated with laterality defects and respiratory symptoms compatible with motile cilia dysfunction. In early mouse embryos, we showed that Daw1 expression is limited to distal, motile ciliated cells of the node, consistent with a role in left-right patterning. daw1 mutant zebrafish exhibited reduced cilia motility and left-right patterning defects, including cardiac looping abnormalities. Importantly, these defects were rescued by wild-type, but not mutant daw1, gene expression. In addition, pathogenic DAW1 missense variants displayed reduced protein stability, whereas DAW1 loss-of-function was associated with distal type 2 outer dynein arm assembly defects involving axonemal respiratory cilia proteins, explaining the reduced cilia-induced fluid flow in particle tracking velocimetry experiments. CONCLUSION: Our data define biallelic DAW1 variants as a cause of human motile ciliopathy and determine that the disease mechanism involves motile cilia dysfunction, explaining the ciliary beating defects observed in affected individuals.
Asunto(s)
Trastornos de la Motilidad Ciliar , Ciliopatías , Proteínas del Citoesqueleto , Animales , Humanos , Ratones , Axonema/genética , Cilios/metabolismo , Trastornos de la Motilidad Ciliar/genética , Trastornos de la Motilidad Ciliar/metabolismo , Trastornos de la Motilidad Ciliar/patología , Ciliopatías/genética , Ciliopatías/metabolismo , Ciliopatías/patología , Proteínas del Citoesqueleto/genética , Mutación , Proteínas/genética , Pez Cebra/genéticaRESUMEN
PURPOSE: We previously defined biallelic HYAL2 variants causing a novel disorder in 2 families, involving orofacial clefting, facial dysmorphism, congenital heart disease, and ocular abnormalities, with Hyal2 knockout mice displaying similar phenotypes. In this study, we better define the phenotype and pathologic disease mechanism. METHODS: Clinical and genomic investigations were undertaken alongside molecular studies, including immunoblotting and immunofluorescence analyses of variant/wild-type human HYAL2 expressed in mouse fibroblasts, and in silico modeling of putative pathogenic variants. RESULTS: Ten newly identified individuals with this condition were investigated, and they were associated with 9 novel pathogenic variants. Clinical studies defined genotype-phenotype correlations and confirmed a recognizable craniofacial phenotype in addition to myopia, cleft lip/palate, and congenital cardiac anomalies as the most consistent manifestations of the condition. In silico modeling of missense variants identified likely deleterious effects on protein folding. Consistent with this, functional studies indicated that these variants cause protein instability and a concomitant cell surface absence of HYAL2 protein. CONCLUSION: These studies confirm an association between HYAL2 alterations and syndromic cleft lip/palate, provide experimental evidence for the pathogenicity of missense alleles, enable further insights into the pathomolecular basis of the disease, and delineate the core and variable clinical outcomes of the condition.
Asunto(s)
Labio Leporino , Fisura del Paladar , Alelos , Animales , Moléculas de Adhesión Celular/genética , Labio Leporino/genética , Fisura del Paladar/genética , Proteínas Ligadas a GPI/genética , Estudios de Asociación Genética , Humanos , Hialuronoglucosaminidasa/genética , Ratones , FenotipoRESUMEN
Phosphatidylinositol 4-kinase IIIα (PI4KIIIα/PI4KA/OMIM:600286) is a lipid kinase generating phosphatidylinositol 4-phosphate (PI4P), a membrane phospholipid with critical roles in the physiology of multiple cell types. PI4KIIIα's role in PI4P generation requires its assembly into a heterotetrameric complex with EFR3, TTC7 and FAM126. Sequence alterations in two of these molecular partners, TTC7 (encoded by TTC7A or TCC7B) and FAM126, have been associated with a heterogeneous group of either neurological (FAM126A) or intestinal and immunological (TTC7A) conditions. Here we show that biallelic PI4KA sequence alterations in humans are associated with neurological disease, in particular hypomyelinating leukodystrophy. In addition, affected individuals may present with inflammatory bowel disease, multiple intestinal atresia and combined immunodeficiency. Our cellular, biochemical and structural modelling studies indicate that PI4KA-associated phenotypical outcomes probably stem from impairment of PI4KIIIα-TTC7-FAM126's organ-specific functions, due to defective catalytic activity or altered intra-complex functional interactions. Together, these data define PI4KA gene alteration as a cause of a variable phenotypical spectrum and provide fundamental new insight into the combinatorial biology of the PI4KIIIα-FAM126-TTC7-EFR3 molecular complex.
Asunto(s)
Enfermedades Desmielinizantes del Sistema Nervioso Central Hereditarias/genética , Atresia Intestinal/genética , Antígenos de Histocompatibilidad Menor/genética , Fosfotransferasas (Aceptor de Grupo Alcohol)/genética , Enfermedades de Inmunodeficiencia Primaria/genética , Femenino , Humanos , Masculino , Linaje , Polimorfismo de Nucleótido SimpleRESUMEN
BACKGROUND: Pathogenicity predictors are integral to genomic variant interpretation but, despite their widespread usage, an independent validation of performance using a clinically relevant dataset has not been undertaken. METHODS: We derive two validation datasets: an 'open' dataset containing variants extracted from publicly available databases, similar to those commonly applied in previous benchmarking exercises, and a 'clinically representative' dataset containing variants identified through research/diagnostic exome and panel sequencing. Using these datasets, we evaluate the performance of three recent meta-predictors, REVEL, GAVIN and ClinPred, and compare their performance against two commonly used in silico tools, SIFT and PolyPhen-2. RESULTS: Although the newer meta-predictors outperform the older tools, the performance of all pathogenicity predictors is substantially lower in the clinically representative dataset. Using our clinically relevant dataset, REVEL performed best with an area under the receiver operating characteristic curve of 0.82. Using a concordance-based approach based on a consensus of multiple tools reduces the performance due to both discordance between tools and false concordance where tools make common misclassification. Analysis of tool feature usage may give an insight into the tool performance and misclassification. CONCLUSION: Our results support the adoption of meta-predictors over traditional in silico tools, but do not support a consensus-based approach as in current practice.
Asunto(s)
Biología Computacional/métodos , Variación Genética/genética , Genómica/métodos , Exoma/genética , Humanos , Curva ROCRESUMEN
Motor neuron diseases (MNDs) encompass an extensive and heterogeneous group of upper and/or lower motor neuron degenerative disorders, in which the particular clinical outcomes stem from the specific neuronal component involved in each condition. While mutations in a large number of molecules associated with lipid metabolism are known to be implicated in MNDs, there remains a lack of clarity regarding the key functional pathways involved, and their inter-relationships. This review highlights evidence that defines defects within two specific lipid (cholesterol/oxysterol and phosphatidylethanolamine) biosynthetic cascades as being centrally involved in MND, particularly hereditary spastic paraplegia. We also identify how other MND-associated molecules may impact these cascades, in particular through impaired organellar interfacing, to propose 'subcellular lipidome imbalance' as a likely common pathomolecular theme in MND. Further exploration of this mechanism has the potential to identify new therapeutic targets and management strategies for modulation of disease progression in hereditary spastic paraplegias and other MNDs.
Asunto(s)
Metabolismo de los Lípidos/fisiología , Neuronas Motoras/metabolismo , Enfermedades Neurodegenerativas/metabolismo , Animales , Humanos , Enfermedades Neurodegenerativas/fisiopatologíaRESUMEN
Heterozygous mutations in KMT2B are associated with an early-onset, progressive and often complex dystonia (DYT28). Key characteristics of typical disease include focal motor features at disease presentation, evolving through a caudocranial pattern into generalized dystonia, with prominent oromandibular, laryngeal and cervical involvement. Although KMT2B-related disease is emerging as one of the most common causes of early-onset genetic dystonia, much remains to be understood about the full spectrum of the disease. We describe a cohort of 53 patients with KMT2B mutations, with detailed delineation of their clinical phenotype and molecular genetic features. We report new disease presentations, including atypical patterns of dystonia evolution and a subgroup of patients with a non-dystonic neurodevelopmental phenotype. In addition to the previously reported systemic features, our study has identified co-morbidities, including the risk of status dystonicus, intrauterine growth retardation, and endocrinopathies. Analysis of this study cohort (n = 53) in tandem with published cases (n = 80) revealed that patients with chromosomal deletions and protein truncating variants had a significantly higher burden of systemic disease (with earlier onset of dystonia) than those with missense variants. Eighteen individuals had detailed longitudinal data available after insertion of deep brain stimulation for medically refractory dystonia. Median age at deep brain stimulation was 11.5 years (range: 4.5-37.0 years). Follow-up after deep brain stimulation ranged from 0.25 to 22 years. Significant improvement of motor function and disability (as assessed by the Burke Fahn Marsden's Dystonia Rating Scales, BFMDRS-M and BFMDRS-D) was evident at 6 months, 1 year and last follow-up (motor, P = 0.001, P = 0.004, and P = 0.012; disability, P = 0.009, P = 0.002 and P = 0.012). At 1 year post-deep brain stimulation, >50% of subjects showed BFMDRS-M and BFMDRS-D improvements of >30%. In the long-term deep brain stimulation cohort (deep brain stimulation inserted for >5 years, n = 8), improvement of >30% was maintained in 5/8 and 3/8 subjects for the BFMDRS-M and BFMDRS-D, respectively. The greatest BFMDRS-M improvements were observed for trunk (53.2%) and cervical (50.5%) dystonia, with less clinical impact on laryngeal dystonia. Improvements in gait dystonia decreased from 20.9% at 1 year to 16.2% at last assessment; no patient maintained a fully independent gait. Reduction of BFMDRS-D was maintained for swallowing (52.9%). Five patients developed mild parkinsonism following deep brain stimulation. KMT2B-related disease comprises an expanding continuum from infancy to adulthood, with early evidence of genotype-phenotype correlations. Except for laryngeal dysphonia, deep brain stimulation provides a significant improvement in quality of life and function with sustained clinical benefit depending on symptoms distribution.
Asunto(s)
Trastornos Distónicos/genética , N-Metiltransferasa de Histona-Lisina/genética , Adolescente , Adulto , Niño , Preescolar , Deleción Cromosómica , Estudios de Cohortes , Simulación por Computador , Estimulación Encefálica Profunda , Progresión de la Enfermedad , Trastornos Distónicos/terapia , Enfermedades del Sistema Endocrino/complicaciones , Enfermedades del Sistema Endocrino/genética , Femenino , Retardo del Crecimiento Fetal/genética , Trastornos Neurológicos de la Marcha/etiología , Trastornos Neurológicos de la Marcha/terapia , Humanos , Enfermedades de la Laringe/etiología , Enfermedades de la Laringe/terapia , Masculino , Mutación , Mutación Missense , Fenotipo , Calidad de Vida , Resultado del Tratamiento , Adulto JovenRESUMEN
We identified a homozygous missense alteration (c.75C>A, p.D25E) in CLCC1, encoding a presumptive intracellular chloride channel highly expressed in the retina, associated with autosomal recessive retinitis pigmentosa (arRP) in eight consanguineous families of Pakistani descent. The p.D25E alteration decreased CLCC1 channel function accompanied by accumulation of mutant protein in granules within the ER lumen, while siRNA knockdown of CLCC1 mRNA induced apoptosis in cultured ARPE-19 cells. TALEN KO in zebrafish was lethal 11 days post fertilization. The depressed electroretinogram (ERG) cone response and cone spectral sensitivity of 5 dpf KO zebrafish and reduced eye size, retinal thickness, and expression of rod and cone opsins could be rescued by injection of wild type CLCC1 mRNA. Clcc1+/- KO mice showed decreased ERGs and photoreceptor number. Together these results strongly suggest that intracellular chloride transport by CLCC1 is a critical process in maintaining retinal integrity, and CLCC1 is crucial for survival and function of retinal cells.