RESUMEN
The collapsin response mediator protein (CRMP) family proteins are intracellular mediators of neurotrophic factors regulating neurite structure/spine formation and are essential for dendrite patterning and directional axonal pathfinding during brain developmental processes. Among this family, CRMP5/DPYSL5 plays a significant role in neuronal migration, axonal guidance, dendrite outgrowth, and synapse formation by interacting with microtubules. Here, we report the identification of missense mutations in DPYSL5 in nine individuals with brain malformations, including corpus callosum agenesis and/or posterior fossa abnormalities, associated with variable degrees of intellectual disability. A recurrent de novo p.Glu41Lys variant was found in eight unrelated patients, and a p.Gly47Arg variant was identified in one individual from the first family reported with Ritscher-Schinzel syndrome. Functional analyses of the two missense mutations revealed impaired dendritic outgrowth processes in young developing hippocampal primary neuronal cultures. We further demonstrated that these mutations, both located in the same loop on the surface of DPYSL5 monomers and oligomers, reduced the interaction of DPYSL5 with neuronal cytoskeleton-associated proteins MAP2 and ßIII-tubulin. Our findings collectively indicate that the p.Glu41Lys and p.Gly47Arg variants impair DPYSL5 function on dendritic outgrowth regulation by preventing the formation of the ternary complex with MAP2 and ßIII-tubulin, ultimately leading to abnormal brain development. This study adds DPYSL5 to the list of genes implicated in brain malformation and in neurodevelopmental disorders.
Asunto(s)
Agenesia del Cuerpo Calloso/genética , Cerebelo/anomalías , Mutación Missense/genética , Trastornos del Neurodesarrollo/genética , Adulto , Agenesia del Cuerpo Calloso/diagnóstico por imagen , Cerebelo/diagnóstico por imagen , Niño , Preescolar , Femenino , Humanos , Hidrolasas/química , Hidrolasas/genética , Discapacidad Intelectual/diagnóstico por imagen , Discapacidad Intelectual/genética , Masculino , Proteínas Asociadas a Microtúbulos/química , Proteínas Asociadas a Microtúbulos/genética , Proteínas Asociadas a Microtúbulos/metabolismo , Modelos Moleculares , Trastornos del Neurodesarrollo/diagnóstico por imagen , Tubulina (Proteína)/metabolismo , Adulto JovenRESUMEN
The ALF transcription factor paralogs, AFF1, AFF2, AFF3, and AFF4, are components of the transcriptional super elongation complex that regulates expression of genes involved in neurogenesis and development. We describe an autosomal dominant disorder associated with de novo missense variants in the degron of AFF3, a nine amino acid sequence important for its binding to ubiquitin ligase, or with de novo deletions of this region. The sixteen affected individuals we identified, along with two previously reported individuals, present with a recognizable pattern of anomalies, which we named KINSSHIP syndrome (KI for horseshoe kidney, NS for Nievergelt/Savarirayan type of mesomelic dysplasia, S for seizures, H for hypertrichosis, I for intellectual disability, and P for pulmonary involvement), partially overlapping the AFF4-associated CHOPS syndrome. Whereas homozygous Aff3 knockout mice display skeletal anomalies, kidney defects, brain malformations, and neurological anomalies, knockin animals modeling one of the microdeletions and the most common of the missense variants identified in affected individuals presented with lower mesomelic limb deformities like KINSSHIP-affected individuals and early lethality, respectively. Overexpression of AFF3 in zebrafish resulted in body axis anomalies, providing some support for the pathological effect of increased amount of AFF3. The only partial phenotypic overlap of AFF3- and AFF4-associated syndromes and the previously published transcriptome analyses of ALF transcription factors suggest that these factors are not redundant and each contributes uniquely to proper development.
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Encefalopatías/genética , Epilepsia/genética , Riñón Fusionado/genética , Discapacidad Intelectual/genética , Mutación Missense , Proteínas Nucleares/genética , Osteocondrodisplasias/genética , Adolescente , Secuencia de Aminoácidos , Animales , Encefalopatías/etiología , Niño , Preescolar , Epilepsia/complicaciones , Evolución Molecular , Femenino , Frecuencia de los Genes , Humanos , Lactante , Masculino , Ratones , Modelos Moleculares , Proteínas Nucleares/química , Proteínas Nucleares/deficiencia , Fenotipo , Estabilidad Proteica , Síndrome , Factores de Elongación Transcripcional/química , Factores de Elongación Transcripcional/genética , Adulto Joven , Pez Cebra/genéticaRESUMEN
NKAP is a ubiquitously expressed nucleoplasmic protein that is currently known as a transcriptional regulatory molecule via its interaction with HDAC3 and spliceosomal proteins. Here, we report a disorder of transcriptional regulation due to missense mutations in the X chromosome gene, NKAP. These mutations are clustered in the C-terminal region of NKAP where NKAP interacts with HDAC3 and post-catalytic spliceosomal complex proteins. Consistent with a role for the C-terminal region of NKAP in embryogenesis, nkap mutant zebrafish with a C-terminally truncated NKAP demonstrate severe developmental defects. The clinical features of affected individuals are highly conserved and include developmental delay, hypotonia, joint contractures, behavioral abnormalities, Marfanoid habitus, and scoliosis. In affected cases, transcriptome analysis revealed the presence of a unique transcriptome signature, which is characterized by the downregulation of long genes with higher exon numbers. These observations indicate the critical role of NKAP in transcriptional regulation and demonstrate that perturbations of the C-terminal region lead to developmental defects in both humans and zebrafish.
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Disfunción Cognitiva/genética , Mutación Missense/genética , Proteínas Represoras/genética , Transcripción Genética/genética , Secuencia de Aminoácidos , Animales , Regulación hacia Abajo/genética , Exones/genética , Regulación de la Expresión Génica/genética , Genes Ligados a X/genética , Histona Desacetilasas/genética , Humanos , Alineación de Secuencia , Transcriptoma/genética , Pez Cebra/genéticaRESUMEN
The RNA polymerase II complex (pol II) is responsible for transcription of all â¼21,000 human protein-encoding genes. Here, we describe sixteen individuals harboring de novo heterozygous variants in POLR2A, encoding RPB1, the largest subunit of pol II. An iterative approach combining structural evaluation and mass spectrometry analyses, the use of S. cerevisiae as a model system, and the assessment of cell viability in HeLa cells allowed us to classify eleven variants as probably disease-causing and four variants as possibly disease-causing. The significance of one variant remains unresolved. By quantification of phenotypic severity, we could distinguish mild and severe phenotypic consequences of the disease-causing variants. Missense variants expected to exert only mild structural effects led to a malfunctioning pol II enzyme, thereby inducing a dominant-negative effect on gene transcription. Intriguingly, individuals carrying these variants presented with a severe phenotype dominated by profound infantile-onset hypotonia and developmental delay. Conversely, individuals carrying variants expected to result in complete loss of function, thus reduced levels of functional pol II from the normal allele, exhibited the mildest phenotypes. We conclude that subtle variants that are central in functionally important domains of POLR2A cause a neurodevelopmental syndrome characterized by profound infantile-onset hypotonia and developmental delay through a dominant-negative effect on pol-II-mediated transcription of DNA.
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ARN Polimerasas Dirigidas por ADN/genética , Hipotonía Muscular/patología , Mutación , Trastornos del Neurodesarrollo/patología , Saccharomyces cerevisiae/crecimiento & desarrollo , Adolescente , Edad de Inicio , Niño , Preescolar , Femenino , Células HeLa , Heterocigoto , Humanos , Masculino , Hipotonía Muscular/enzimología , Hipotonía Muscular/genética , Trastornos del Neurodesarrollo/enzimología , Trastornos del Neurodesarrollo/genética , Fenotipo , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismoRESUMEN
PURPOSE: Proline Rich 12 (PRR12) is a gene of unknown function with suspected DNA-binding activity, expressed in developing mice and human brains. Predicted loss-of-function variants in this gene are extremely rare, indicating high intolerance of haploinsufficiency. METHODS: Three individuals with intellectual disability and iris anomalies and truncating de novo PRR12 variants were described previously. We add 21 individuals with similar PRR12 variants identified via matchmaking platforms, bringing the total number to 24. RESULTS: We observed 12 frameshift, 6 nonsense, 1 splice-site, and 2 missense variants and one patient with a gross deletion involving PRR12. Three individuals had additional genetic findings, possibly confounding the phenotype. All patients had developmental impairment. Variable structural eye defects were observed in 12/24 individuals (50%) including anophthalmia, microphthalmia, colobomas, optic nerve and iris abnormalities. Additional common features included hypotonia (61%), heart defects (52%), growth failure (54%), and kidney anomalies (35%). PrediXcan analysis showed that phecodes most strongly associated with reduced predicted PRR12 expression were enriched for eye- (7/30) and kidney- (4/30) phenotypes, such as wet macular degeneration and chronic kidney disease. CONCLUSION: These findings support PRR12 haploinsufficiency as a cause for a novel disorder with a wide clinical spectrum marked chiefly by neurodevelopmental and eye abnormalities.
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Haploinsuficiencia , Discapacidad Intelectual , Animales , Haploinsuficiencia/genética , Humanos , Discapacidad Intelectual/genética , Ratones , Hipotonía Muscular , Mutación Missense , FenotipoRESUMEN
Mutations that alter signaling of RAS/MAPK-family proteins give rise to a group of Mendelian diseases known as RASopathies. However, among RASopathies, the matrix of genotype-phenotype relationships is still incomplete, in part because there are many RAS-related proteins and in part because the phenotypic consequences may be variable and/or pleiotropic. Here, we describe a cohort of ten cases, drawn from six clinical sites and over 16,000 sequenced probands, with de novo protein-altering variation in RALA, a RAS-like small GTPase. All probands present with speech and motor delays, and most have intellectual disability, low weight, short stature, and facial dysmorphism. The observed rate of de novo RALA variants in affected probands is significantly higher (p = 4.93 x 10(-11)) than expected from the estimated random mutation rate. Further, all de novo variants described here affect residues within the GTP/GDP-binding region of RALA; in fact, six alleles arose at only two codons, Val25 and Lys128. The affected residues are highly conserved across both RAL- and RAS-family genes, are devoid of variation in large human population datasets, and several are homologous to positions at which disease-associated variants have been observed in other GTPase genes. We directly assayed GTP hydrolysis and RALA effector-protein binding of the observed variants, and found that all but one tested variant significantly reduced both activities compared to wild-type. The one exception, S157A, reduced GTP hydrolysis but significantly increased RALA-effector binding, an observation similar to that seen for oncogenic RAS variants. These results show the power of data sharing for the interpretation and analysis of rare variation, expand the spectrum of molecular causes of developmental disability to include RALA, and provide additional insight into the pathogenesis of human disease caused by mutations in small GTPases.
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Discapacidades del Desarrollo/genética , Discapacidad Intelectual/genética , Proteínas Mitocondriales/genética , Mutación , Dominios y Motivos de Interacción de Proteínas/genética , Proteínas de Unión al GTP ral/genética , Proteínas ras/genética , Facies , Genotipo , Guanosina Difosfato/metabolismo , Guanosina Trifosfato/metabolismo , Humanos , Proteínas Mitocondriales/química , Modelos Moleculares , Mutación Missense , Fenotipo , Conformación Proteica , Proteínas de Unión al GTP ral/química , Proteínas ras/químicaRESUMEN
PURPOSE: Nontruncating variants in SMARCA2, encoding a catalytic subunit of SWI/SNF chromatin remodeling complex, cause Nicolaides-Baraitser syndrome (NCBRS), a condition with intellectual disability and multiple congenital anomalies. Other disorders due to SMARCA2 are unknown. METHODS: By next-generation sequencing, we identified candidate variants in SMARCA2 in 20 individuals from 18 families with a syndromic neurodevelopmental disorder not consistent with NCBRS. To stratify variant interpretation, we functionally analyzed SMARCA2 variants in yeasts and performed transcriptomic and genome methylation analyses on blood leukocytes. RESULTS: Of 20 individuals, 14 showed a recognizable phenotype with recurrent features including epicanthal folds, blepharophimosis, and downturned nasal tip along with variable degree of intellectual disability (or blepharophimosis intellectual disability syndrome [BIS]). In contrast to most NCBRS variants, all SMARCA2 variants associated with BIS are localized outside the helicase domains. Yeast phenotype assays differentiated NCBRS from non-NCBRS SMARCA2 variants. Transcriptomic and DNA methylation signatures differentiated NCBRS from BIS and those with nonspecific phenotype. In the remaining six individuals with nonspecific dysmorphic features, clinical and molecular data did not permit variant reclassification. CONCLUSION: We identified a novel recognizable syndrome named BIS associated with clustered de novo SMARCA2 variants outside the helicase domains, phenotypically and molecularly distinct from NCBRS.
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Blefarofimosis , Hipotricosis , Discapacidad Intelectual , Facies , Deformidades Congénitas del Pie , Humanos , Discapacidad Intelectual/genética , Fenotipo , Factores de Transcripción/genéticaRESUMEN
PURPOSE: Variants in IQSEC2, escaping X inactivation, cause X-linked intellectual disability with frequent epilepsy in males and females. We aimed to investigate sex-specific differences. METHODS: We collected the data of 37 unpublished patients (18 males and 19 females) with IQSEC2 pathogenic variants and 5 individuals with variants of unknown significance and reviewed published variants. We compared variant types and phenotypes in males and females and performed an analysis of IQSEC2 isoforms. RESULTS: IQSEC2 pathogenic variants mainly led to premature truncation and were scattered throughout the longest brain-specific isoform, encoding the synaptic IQSEC2/BRAG1 protein. Variants occurred de novo in females but were either de novo (2/3) or inherited (1/3) in males, with missense variants being predominantly inherited. Developmental delay and intellectual disability were overall more severe in males than in females. Likewise, seizures were more frequently observed and intractable, and started earlier in males than in females. No correlation was observed between the age at seizure onset and severity of intellectual disability or resistance to antiepileptic treatments. CONCLUSION: This study provides a comprehensive overview of IQSEC2-related encephalopathy in males and females, and suggests that an accurate dosage of IQSEC2 at the synapse is crucial during normal brain development.
Asunto(s)
Encefalopatías/genética , Factores de Intercambio de Guanina Nucleótido/genética , Discapacidad Intelectual/genética , Convulsiones/genética , Encéfalo/crecimiento & desarrollo , Encéfalo/metabolismo , Encefalopatías/epidemiología , Encefalopatías/fisiopatología , Femenino , Humanos , Lactante , Recién Nacido , Discapacidad Intelectual/epidemiología , Discapacidad Intelectual/fisiopatología , Masculino , Mutación , Linaje , Fenotipo , Isoformas de Proteínas/genética , Convulsiones/epidemiología , Convulsiones/fisiopatología , Caracteres SexualesRESUMEN
This Article was originally published under Nature Research's License to Publish, but has now been made available under a CC BY 4.0 license. The PDF and HTML versions of the Article have been modified accordingly.
RESUMEN
The importance of gonadal mosaicism in families with apparently de novo mutations is being increasingly recognized. We report on two affected brothers initially suggestive of X-linked or autosomal recessive inheritance. Malan syndrome due to shared NFIX variants was diagnosed in the brothers using exome sequencing. The boys shared the same paternal but not maternal haplotype around NFIX, and deep amplicon sequencing showed ~7% of the variant in paternal sperm but not in paternal blood and saliva. We performed review of previous cases of gonadal mosaicism, which suggests that the phenomenon is not uncommon. Gonadal mosaicism is often not accompanied by somatic mosaicism in tissues routinely used for testing, and if both types of mosaicism are present, the frequency of the variant in sperm is often higher than in somatic cells. In families with shared apparently de novo variants without evidence of parental somatic mosaicism, the transmitting parent may be determined through haplotyping of exome variants. Gonadal mosaicism has important consequences for recurrence risks and should be considered in genetic counseling in families with de novo variants.
Asunto(s)
Anomalías Múltiples/genética , Gónadas/patología , Mosaicismo , Mutación/genética , Factores de Transcripción NFI/genética , Hermanos , Secuencia de Aminoácidos , Secuencia de Bases , Niño , Preescolar , Femenino , Humanos , Masculino , Factores de Transcripción NFI/química , Linaje , Embarazo , Síndrome , Adulto JovenRESUMEN
BACKGROUND: Genetic testing rapidly penetrates into all medical specialties and medical students must acquire skills in this area. However, many of them consider it difficult. Furthermore, many find these topics less appealing and not connected to their future specialization in different fields of clinical medicine. Student-centred strategies such as problem-based learning, gamification and the use of real data can increase the appeal of a difficult topic such as genetic testing, a field at the crossroads of genetics, molecular biology and bioinformatics. METHODS: We designed an electronic teaching application which students registered in the undergraduate Medical Biology course can access online. A study was carried out to assess the influence of implementation of the new method. We performed pretest/posttest evaluation and analyzed the results using the sign test with median values. We also collected students' personal comments. RESULTS: The newly developed interactive application simulates the process of molecular genetic diagnostics of a hereditary disorder in a family. Thirteen tasks guide students through clinical and laboratory steps needed to reach the final diagnosis. Genetics and genomics are fields strongly dependent on electronic databases and computer-based data analysis tools. The tasks employ publicly available internet bioinformatic resources used routinely in medical genetics departments worldwide. Authenticity is assured by the use of modified and de-identified clinical and laboratory data from real families analyzed in our previous research projects. Each task contains links to databases and data processing tools needed to solve the task, and an answer box. If the entered answer is correct, the system allows the user to proceed to the next task. The solving of consecutive tasks arranged into a single narrative resembles a computer game, making the concept appealing. There was a statistically significant improvement of knowledge and skills after the practical class, and most comments on the application were positive. A demo version is available at https://medbio.lf2.cuni.cz/demo_m/ . Full version is available on request from the authors. CONCLUSIONS: Our concept proved to be appealing to the students and effective in teaching medical molecular genetics. It can be modified for training in the use of electronic information resources in other medical specialties.
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Instrucción por Computador , Educación de Pregrado en Medicina/métodos , Pruebas Genéticas , Genética Médica/educación , Biología Computacional/educación , Enfermedades Genéticas Congénitas/diagnóstico , Humanos , Medicina Molecular/educación , Aprendizaje Basado en Problemas , Enseñanza , Interfaz Usuario-Computador , Juegos de VideoRESUMEN
Analyses at nucleotide resolution reveal unexpected complexity of seemingly simple and balanced chromosomal rearrangements. Chromothripsis is a rare complex aberration involving local shattering of one or more chromosomes and reassembly of the resulting DNA segments. This can influence gene expression and cause abnormal phenotypes. We studied the structure and mechanism of a seemingly balanced de novo complex rearrangement of four chromosomes in a boy with developmental and growth delay. Microarray analysis revealed two paternal de novo deletions of 0.7 and 2.5 Mb at two of the breakpoints in 1q24.3 and 6q24.1-q24.2, respectively, which could explain most symptoms of the patient. Subsequent whole-genome mate-pair sequencing confirmed the chromothriptic nature of the rearrangement. The four participating chromosomes were broken into 29 segments longer than 1 kb. Sanger sequencing of all breakpoint junctions revealed additional complexity compatible with the involvement of different repair pathways. We observed translocation of a 33 bp long DNA fragment, which may have implications for the definition of the lower size limit of structural variants. Our observations and literature review indicate that even very small fragments from shattered chromosomes can be detected and handled by the repair machinery during germline chromothriptic chromosome reassembly.
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Cromotripsis , Reparación del ADN , ADN/genética , Células Germinativas/metabolismo , Adolescente , Adulto , Secuencia de Bases , Preescolar , Cromosomas Humanos/genética , Humanos , Lactante , Recién Nacido , Cariotipo , MasculinoRESUMEN
The prenatal finding of a small supernumerary marker chromosome (sSMC) is a challenge for genetic counseling. Our analytic algorithm is based on sSMC frequencies and multicolor FISH to accelerate the procedure. The chromosomal origin, size, and degree of mosaicism of the sSMC then determine the prognosis. We illustrate the effectiveness on 4 prenatally identified de novo mosaic sSMCs derived from chromosomes 13/21, X, 3, and 17. Three sSMC carriers had a good prognosis and apparently healthy children were born, showing no abnormality till the last examination at the age of 4 years. One case had a poor prognosis, and the parents decided to terminate the pregnancy. Our work contributes to the laboratory and clinical management of prenatally detected sSMCs. FISH is a reliable method for fast sSMC evaluation and prognosis assessment; it prevents unnecessary delays and uncertainty, allows informed decision making, and reduces unnecessary pregnancy terminations.
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Aberraciones Cromosómicas , Heterocigoto , Diagnóstico Prenatal , Adulto , Algoritmos , Preescolar , Femenino , Estudios de Asociación Genética , Asesoramiento Genético , Humanos , Hibridación Fluorescente in Situ , Lactante , Cariotipificación , Masculino , Edad Materna , Embarazo , PronósticoRESUMEN
Microdeletions of 17q24.2-q24.3 have been described in several patients with developmental and speech delay, growth retardation, and other features. The relatively large size and limited overlap of the deletions complicate the genotype-phenotype correlation. We identified a girl with intellectual disability, growth retardation, dysmorphic features, and a de novo 2.8 Mb long deletion of 17q24.2-q24.3. Her phenotype was strikingly similar to one previously described boy with Dubowitz syndrome (MIM 223370) and a de novo 3.9 Mb long deletion encompassing the deletion of our patient. In addition, both patients had the shortest telomeres among normal age-matched controls. Our review of all 17q24.2-q24.3 deletion patients revealed additional remarkable phenotypic features shared by the patients, some of which have consequences for their management. Proposed novel genotype-phenotype correlations based on new literature information on the region include the role of PSMD12 and BPTF, the genes recently associated with syndromic neurodevelopmental disorders, and a possible role of the complex topologically associated domain structure of the region, which may explain some of the phenotypic discrepancies observed between patients with similar but not identical deletions. Nevertheless, although different diagnoses including the Dubowitz, Nijmegen breakage (MIM 251260), Silver-Russell (MIM 180860), or Myhre (MIM 139210) syndromes were originally considered in the 17q24.2-q24.3 deletion patients, they clearly belong to one diagnostic entity defined by their deletions and characterized especially by developmental delay, specific facial dysmorphism, abnormalities of extremities and other phenotypes, and possibly also short telomere length.
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Cromosomas Humanos Par 17 , Discapacidades del Desarrollo/genética , Telómero , Niño , Deleción Cromosómica , Cromosomas Humanos Par 17/genética , Discapacidades del Desarrollo/etiología , Eccema/etiología , Cara/anomalías , Facies , Femenino , Fibromatosis Gingival/genética , Trastornos del Crecimiento/etiología , Humanos , Hipertricosis/genética , Discapacidad Intelectual/etiología , Microcefalia/etiología , FenotipoRESUMEN
BACKGROUND: Mutations in forkhead box protein P1 (FOXP1) cause intellectual disability (ID) and specific language impairment (SLI), with or without autistic features (MIM: 613670). Despite multiple case reports no specific phenotype emerged so far. METHODS: We correlate clinical and molecular data of 25 novel and 23 previously reported patients with FOXP1 defects. We evaluated FOXP1 activity by an in vitro luciferase model and assessed protein stability in vitro by western blotting. RESULTS: Patients show ID, SLI, neuromotor delay (NMD) and recurrent facial features including a high broad forehead, bent downslanting palpebral fissures, ptosis and/or blepharophimosis and a bulbous nasal tip. Behavioural problems and autistic features are common. Brain, cardiac and urogenital malformations can be associated. More severe ID and NMD, sensorineural hearing loss and feeding difficulties are more common in patients with interstitial 3p deletions (14 patients) versus patients with monogenic FOXP1 defects (34 patients). Mutations result in impaired transcriptional repression and/or reduced protein stability. CONCLUSIONS: FOXP1-related ID syndrome is a recognisable entity with a wide clinical spectrum and frequent systemic involvement. Our data will be helpful to evaluate genotype-phenotype correlations when interpreting next-generation sequencing data obtained in patients with ID and/or SLI and will guide clinical management.
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Factores de Transcripción Forkhead/genética , Discapacidad Intelectual/genética , Proteínas Represoras/genética , Trastorno del Espectro Autista/genética , Cara/anomalías , Femenino , Factores de Transcripción Forkhead/química , Factores de Transcripción Forkhead/metabolismo , Humanos , Trastornos del Lenguaje/genética , Masculino , Trastornos de la Destreza Motora/genética , Mutación , Mutación Missense , Trastornos del Neurodesarrollo/genética , Fenotipo , Estabilidad Proteica , Proteínas Represoras/química , Proteínas Represoras/metabolismo , Síndrome , Transcripción GenéticaRESUMEN
Both gain- and loss-of-function mutations have recently implicated HCFC1 in neurodevelopmental disorders. Here, we extend our previous HCFC1 over-expression studies by employing short hairpin RNA to reduce the expression of Hcfc1 in embryonic neural cells. We show that in contrast to over-expression, loss of Hcfc1 favoured proliferation of neural progenitor cells at the expense of differentiation and promoted axonal growth of post-mitotic neurons. To further support the involvement of HCFC1 in neurological disorders, we report two novel HCFC1 missense variants found in individuals with intellectual disability (ID). One of these variants, together with three previously reported HCFC1 missense variants of unknown pathogenicity, were functionally assessed using multiple cell-based assays. We show that three out of the four variants tested result in a partial loss of HCFC1 function. While over-expression of the wild-type HCFC1 caused reduction in HEK293T cell proliferation and axonal growth of neurons, these effects were alleviated upon over-expression of three of the four HCFC1 variants tested. One of these partial loss-of-function variants disrupted a nuclear localization sequence and the resulting protein displayed reduced ability to localize to the cell nucleus. The other two variants displayed negative effects on the expression of the HCFC1 target gene MMACHC, which is responsible for the metabolism of cobalamin, suggesting that these individuals may also be susceptible to cobalamin deficiency. Together, our work identifies plausible cellular consequences of missense HCFC1 variants and identifies likely and relevant disease mechanisms that converge on embryonic stages of brain development.
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Encéfalo/citología , Factor C1 de la Célula Huésped/genética , Mutación , Células-Madre Neurales/metabolismo , Transporte Activo de Núcleo Celular , Secuencia de Aminoácidos , Sustitución de Aminoácidos , Animales , Encéfalo/embriología , Proteínas Portadoras/genética , Diferenciación Celular/genética , Proliferación Celular , Células Cultivadas , Femenino , Expresión Génica , Células HEK293 , Factor C1 de la Célula Huésped/química , Factor C1 de la Célula Huésped/metabolismo , Humanos , Discapacidad Intelectual/genética , Masculino , Ratones , Células-Madre Neurales/citología , Oxidorreductasas , Linaje , Interferencia de ARN , ARN Interferente Pequeño/genética , Transducción GenéticaRESUMEN
BACKGROUND: Whole exome sequencing is a powerful tool for the analysis of genetically heterogeneous conditions. The prioritization of variants identified often focuses on nonsense, frameshift and canonical splice site mutations, and highly deleterious missense variants, although other defects can also play a role. The definition of the phenotype range and course of rare genetic conditions requires long-term clinical follow-up of patients. CASE PRESENTATION: We report an adult female patient with severe intellectual disability, severe speech delay, epilepsy, autistic features, aggressiveness, sleep problems, broad-based clumsy gait and constipation. Whole exome sequencing identified a de novo mutation in the SYNGAP1 gene. The variant was located in the broader splice donor region of intron 10 and replaced G by A at position +5 of the splice site. The variant was predicted in silico and shown experimentally to abolish the regular splice site and to activate a cryptic donor site within exon 10, causing frameshift and premature termination. The overall clinical picture of the patient corresponded well with the characteristic SYNGAP1-associated phenotype observed in previously reported patients. However, our patient was 31 years old which contrasted with most other published SYNGAP1 cases who were much younger. Our patient had a significant growth delay and microcephaly. Both features normalised later, although the head circumference stayed only slightly above the lower limit of the norm. The patient had a delayed puberty. Her cognitive and language performance remained at the level of a one-year-old child even in adulthood and showed a slow decline. Myopathic facial features and facial dysmorphism became more pronounced with age. Although the gait of the patient was unsteady in childhood, more severe gait problems developed in her teens. While the seizures remained well-controlled, her aggressive behaviour worsened with age and required extensive medication. CONCLUSIONS: The finding in our patient underscores the notion that the interpretation of variants identified using whole exome sequencing should focus not only on variants in the canonical splice dinucleotides GT and AG, but also on broader splice regions. The long-term clinical follow-up of our patient contributes to the knowledge of the developmental trajectory in individuals with SYNGAP1 gene defects.
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Discapacidad Intelectual/genética , Proteínas Activadoras de ras GTPasa/genética , Adulto , Secuencia de Bases , Exoma , Femenino , Estudios de Seguimiento , Variación Genética , Genómica , Humanos , Discapacidad Intelectual/diagnóstico , Cariotipificación , Análisis por Micromatrices , Microcefalia/diagnóstico , Microcefalia/genética , Mutación , Fenotipo , Análisis de Secuencia de ADNRESUMEN
We identified a de novo deletion of 14q11.2 in a Czech patient with developmental delay, mild autistic features, macrosomy, macrocephaly, orthognathic deformities, and dysmorphic facial features. The clinical follow-up of the patient lasting 14 years documented changes in the facial dysmorphism from infancy to adolescence. The deletion affects approximately 200 kb of DNA with five protein-coding genes and two snoRNA genes. Two of the protein-coding genes, SUPT16H and CHD8, have been proposed as candidate genes for a new microdeletion syndrome. Our patient further supports the existence of this syndrome and extends its phenotypic spectrum, especially points to the possibility that orthognathic deformities may be associated with microdeletions of 14q11.2. CHD8 mutations have been found in patients with neurodevelopmental disorders and macrocephaly. The HNRNPC gene, repeatedly deleted in patients with developmental delay, is another candidate as its 5Ì end is adjacent to the deletion, and the expression of this gene may be affected by position effect.
Asunto(s)
Anomalías Múltiples/diagnóstico , Deleción Cromosómica , Cromosomas Humanos Par 14/genética , Proteínas de Unión al ADN/genética , Factores de Transcripción/genética , Anomalías Múltiples/genética , Adolescente , Discapacidades del Desarrollo/diagnóstico , Discapacidades del Desarrollo/genética , Estudios de Seguimiento , Humanos , Discapacidad Intelectual/diagnóstico , Discapacidad Intelectual/genética , Masculino , Megalencefalia/diagnóstico , Megalencefalia/genéticaRESUMEN
Chromosome 17q21.31 microdeletion syndrome is a genomic disorder caused by a recurrent 600 kb long deletion. The deletion affects the region of a common inversion present in about 20% of Europeans. The inversion is associated with the H2 haplotype carrying additional low-copy repeats susceptible to non-allelic homologous recombination, and this haplotype is prone to deletion. No instances of 17q21.31 deletions inherited from an affected parent have been reported, and the deletions always affected a parental chromosome with the H2 haplotype. The syndrome is characterized clinically by intellectual disability, hypotonia, friendly behavior and specific facial dysmorphism with long face, large tubular or pear-shaped nose and bulbous nasal tip. We present monozygotic twin sisters showing the typical clinical picture of the syndrome. The phenotype of the sisters was very similar, with a slightly more severe presentation in Twin B. The 17q21.31 microdeletion was confirmed in both patients but in neither of their parents. Potential copy number differences between the genomes of the twins were subsequently searched using high-resolution single nucleotide polymorphism (SNP) and comparative genome hybridisation (CGH) arrays. However, these analyses identified no additional aberrations or genomic differences that could potentially be responsible for the subtle phenotypic differences. These could possibly be related to the more severe perinatal history of Twin B, or to the variable expressivity of the disorder. In accord with the expectations, one of the parents (the mother) was shown to carry the H2 haplotype, and the maternal allele of chromosome 17q21.31 was missing in the twins.
Asunto(s)
Haplotipos , Discapacidad Intelectual/genética , Polimorfismo de Nucleótido Simple , Gemelos Monocigóticos/genética , Adulto , Deleción Cromosómica , Cromosomas Humanos Par 17/genética , Femenino , Humanos , Discapacidad Intelectual/patología , Masculino , Síndrome de Smith-MagenisRESUMEN
Pre-mRNA splicing is a highly coordinated process. While its dysregulation has been linked to neurological deficits, our understanding of the underlying molecular and cellular mechanisms remains limited. We implicated pathogenic variants in U2AF2 and PRPF19, encoding spliceosome subunits in neurodevelopmental disorders (NDDs), by identifying 46 unrelated individuals with 23 de novo U2AF2 missense variants (including 7 recurrent variants in 30 individuals) and 6 individuals with de novo PRPF19 variants. Eight U2AF2 variants dysregulated splicing of a model substrate. Neuritogenesis was reduced in human neurons differentiated from human pluripotent stem cells carrying two U2AF2 hyper-recurrent variants. Neural loss of function (LoF) of the Drosophila orthologs U2af50 and Prp19 led to lethality, abnormal mushroom body (MB) patterning, and social deficits, which were differentially rescued by wild-type and mutant U2AF2 or PRPF19. Transcriptome profiling revealed splicing substrates or effectors (including Rbfox1, a third splicing factor), which rescued MB defects in U2af50-deficient flies. Upon reanalysis of negative clinical exomes followed by data sharing, we further identified 6 patients with NDD who carried RBFOX1 missense variants which, by in vitro testing, showed LoF. Our study implicates 3 splicing factors as NDD-causative genes and establishes a genetic network with hierarchy underlying human brain development and function.