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1.
Genet Med ; 2019 Jul 18.
Artigo em Inglês | MEDLINE | ID: mdl-31316167

RESUMO

PURPOSE: Congenital contractural arachnodactyly (CCA) is an autosomal dominant connective tissue disorder manifesting joint contractures, arachnodactyly, crumpled ears, and kyphoscoliosis as main features. Due to its rarity, rather aspecific clinical presentation, and overlap with other conditions including Marfan syndrome, the diagnosis is challenging, but important for prognosis and clinical management. CCA is caused by pathogenic variants in FBN2, encoding fibrillin-2, but locus heterogeneity has been suggested. We designed a clinical scoring system and diagnostic criteria to support the diagnostic process and guide molecular genetic testing. METHODS: In this retrospective study, we assessed 167 probands referred for FBN2 analysis and classified them into a FBN2-positive (n = 44) and FBN2-negative group (n = 123) following molecular analysis. We developed a 20-point weighted clinical scoring system based on the prevalence of ten main clinical characteristics of CCA in both groups. RESULTS: The total score was significantly different between the groups (P < 0.001) and was indicative for classifying patients into unlikely CCA (total score <7) and likely CCA (total score ≥7) groups. CONCLUSIONS: Our clinical score is helpful for clinical guidance for patients suspected to have CCA, and provides a quantitative tool for phenotyping in research settings.

2.
Hum Mutat ; 40(9): 1346-1363, 2019 09.
Artigo em Inglês | MEDLINE | ID: mdl-31209962

RESUMO

Intellectual disability (ID) and autism spectrum disorder (ASD) are clinically and genetically heterogeneous diseases. Recent whole exome sequencing studies indicated that genes associated with different neurological diseases are shared across disorders and converge on common functional pathways. Using the Ion Torrent platform, we developed a low-cost next-generation sequencing gene panel that has been transferred into clinical practice, replacing single disease-gene analyses for the early diagnosis of individuals with ID/ASD. The gene panel was designed using an innovative in silico approach based on disease networks and mining data from public resources to score disease-gene associations. We analyzed 150 unrelated individuals with ID and/or ASD and a confident diagnosis has been reached in 26 cases (17%). Likely pathogenic mutations have been identified in another 15 patients, reaching a total diagnostic yield of 27%. Our data also support the pathogenic role of genes recently proposed to be involved in ASD. Although many of the identified variants need further investigation to be considered disease-causing, our results indicate the efficiency of the targeted gene panel on the identification of novel and rare variants in patients with ID and ASD.

3.
Seizure ; 66: 81-85, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-30818181

RESUMO

PURPOSE: Mutations in SZT2 have been previously reported in several cases of early onset epilepsy and intellectual disability. In this study we investigate potential causal mutations in two male siblings affected by early onset epilepsy, intellectual disability and macrocephaly. METHODS: We use family-based whole-exome sequencing to identify candidate variants. RESULTS: We report the identification of two potential causal SZT2 mutations in compound heterozygous state. We observe considerable differences in the clinical phenotype severity of the two affected individuals. The cerebral MRI revealed no abnormalities in the older affected brother, while in the youngest one it revealed a right frontal polymicrogiria. Moreover, while good seizure control was achieved in the older affected individual the younger brother is affected by pharmacoresistant epilepsy, progressive spastic paraplegia, cortical myoclonus and a more severe intellectual disability. We also analyzed the relative location of the reported pathogenic mutations in the SZT2 protein. CONCLUSION: Variable phenotypic expressivity is observed for this condition, while the location and type of mutations in SZT2 also has a potential impact on epilepsy severity. These findings extend our knowledge of epileptogenic conditions related to SZT2 and mTOR signaling.


Assuntos
Epilepsia/genética , Saúde da Família , Deficiência Intelectual/genética , Megalencefalia/genética , Mutação/genética , Proteínas do Tecido Nervoso/genética , Adulto , Análise Mutacional de DNA , Epilepsia/complicações , Epilepsia/diagnóstico por imagem , Humanos , Deficiência Intelectual/complicações , Deficiência Intelectual/diagnóstico por imagem , Estudos Longitudinais , Imagem por Ressonância Magnética , Masculino , Megalencefalia/complicações , Megalencefalia/diagnóstico por imagem , Sequenciamento Completo do Exoma , Adulto Jovem
5.
Eur J Hum Genet ; 25(6): 694-701, 2017 06.
Artigo em Inglês | MEDLINE | ID: mdl-28422132

RESUMO

16q24 deletion involving the ANKRD11 gene, ranging from 137 kb to 2 Mb, have been associated with a microdeletion syndrome characterized by variable cognitive impairment, autism spectrum disorder, facial dysmorphisms with dental anomalies, brain abnormalities essentially affecting the corpus callosum and short stature. On the other hand, patients carrying either deletions encompassing solely ANKRD11 or its loss-of-function variants were reported in association with the KBG syndrome, characterized by a very similar phenotype, including mild-to-moderate intellectual disability, short stature and macrodontia of upper incisors, with inter and intrafamilial variability. To assess whether the haploinsufficiency of ANKRD11-flanking genes, such as ZFPM1, CDH15 and ZNF778, contributed to either the severity of the neurological impairment or was associated with other clinical features, we collected 12 new cases with a 16q24.2q24.3 deletion (de novo in 11 cases), ranging from 343 kb to 2.3 Mb. In 11 of them, the deletion involved the ANKRD11 gene, whereas in 1 case only flanking genes upstream to it were deleted. By comparing the clinical and genetic features of our patients with those previously reported, we show that the severity of the neurological phenotype and the frequency of congenital heart defects characterize the deletions that, besides ANKRD11, contain ZFPM1, CDH15 and ZNF778 as well. Moreover, the presence of thrombocytopenia and astigmatism should be taken into account to distinguish between 16q24 microdeletion syndrome and KBG syndrome. The single patient not deleted for ANKRD11, whose phenotype is characterized by milder psychomotor delay, cardiac congenital malformation, thrombocytopenia and astigmatism, confirms all this data.


Assuntos
Anormalidades Múltiplas/genética , Doenças do Desenvolvimento Ósseo/genética , Deleção Cromossômica , Cromossomos Humanos Par 16/genética , Haploinsuficiência , Deficiência Intelectual/genética , Proteínas Repressoras/genética , Anormalidades Dentárias/genética , Fatores de Transcrição/genética , Anormalidades Múltiplas/diagnóstico , Adolescente , Adulto , Doenças do Desenvolvimento Ósseo/diagnóstico , Caderinas/genética , Criança , Diagnóstico Diferencial , Facies , Feminino , Humanos , Deficiência Intelectual/diagnóstico , Masculino , Proteínas Nucleares/genética , Fenótipo , Anormalidades Dentárias/diagnóstico , Fatores de Transcrição/metabolismo
6.
Invest Ophthalmol Vis Sci ; 58(2): 821-832, 2017 02 01.
Artigo em Inglês | MEDLINE | ID: mdl-28159970

RESUMO

Purpose: The purpose of this study was to clinically characterize patients with CNGA3-linked achromatopsia (CNGA3-ACHM) in preparation of a gene therapy trial. Methods: Thirty-six patients (age 7-56 years) with complete (cACHM) or incomplete (iACHM) CNGA3-ACHM were examined, including detailed psychophysical tests, extended electrophysiology, and assessment of morphology by fundus autofluorescence and spectral-domain optical coherence tomography (SD-OCT). Results: Mean best-corrected visual acuity was 0.78 ± 0.14 logMAR. Color vision tests were consistent with a rod-dominated function in every cACHM patient. Microperimetry indicated an overall lowered retinal sensitivity within 20° of visual field. In electroretinography (ERG), photopic responses were nondetectable in cACHM patients, but residual cone responses were observed in the iACHM patients. Scotopic responses were altered referring to anomalies of photoreceptor and postreceptor signaling, whereas in voltage versus intensity functions, Vmax was significantly below normal values (P < 0.05). In contrast, slope (n) and semisaturation intensity (K) were found to be within normal limits. Spectral-domain OCT examination showed no specific changes in 14.7%, disruption of the ellipsoid zone (EZ) at the fovea in 38.2%, absent EZ in 17.7%, a hyporeflective zone in 20.5%, and outer retinal atrophy in 8.9% of all cases and foveal hypoplasia in 29 patients (85%). No correlation of retinal morphology with visual function or with a specific genotype was found. The severity of morphologic and functional changes lacked a robust association with age. Conclusions: Our extended investigations prove that even among such a genetically homogenous group of patients, no specific correlations regarding function and morphology severity and age can be observed. Therefore, the therapeutic window seems to be wider than previously indicated.


Assuntos
Defeitos da Visão Cromática/genética , Defeitos da Visão Cromática/terapia , Canais de Cátion Regulados por Nucleotídeos Cíclicos/genética , Terapia Genética , Fenótipo , Retina/patologia , Retina/fisiopatologia , Adolescente , Adulto , Criança , Percepção de Cores/fisiologia , Defeitos da Visão Cromática/patologia , Defeitos da Visão Cromática/fisiopatologia , Adaptação à Escuridão/fisiologia , Eletrorretinografia , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Estudos Prospectivos , Psicofísica , Limiar Sensorial , Tomografia de Coerência Óptica , Acuidade Visual/fisiologia , Campos Visuais/fisiologia , Adulto Jovem
7.
Clin Cancer Res ; 23(1): 273-282, 2017 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-27390349

RESUMO

PURPOSE: The majority of gastrointestinal stromal tumors (GIST) are driven by KIT, PDGFRA, or, less commonly, BRAF mutations, and SDH gene inactivation is involved in a limited fraction of gastric lesions. However, about 10% of GISTs are devoid of any of such alterations and are poorly responsive to standard treatments. This study aims to shed light on the molecular drivers of quadruple-negative GISTs. EXPERIMENTAL DESIGN: Twenty-two sporadic quadruple-negative GISTs with no prior association with Neurofibromatosis Type 1 syndrome were molecularly profiled for a panel of genes belonging to tyrosine kinase pathways or previously implicated in GISTs. For comparison purposes, 24 GISTs carrying KIT, PDGFRA, or SDH gene mutations were also analyzed. Molecular findings were correlated to clinicopathologic features. RESULTS: Most quadruple-negative GISTs featured intestinal localization, with a female predilection. About 60% (13/22) of quadruple-negative tumors carried NF1 pathogenic mutations, often associated with biallelic inactivation. The analysis of normal tissues, available in 11 cases, indicated the constitutional nature of the NF1 mutation in 7 of 11 cases, unveiling an unrecognized Neurofibromatosis Type 1 syndromic condition. Multifocality and a multinodular pattern of growth were common findings in NF1-mutated quadruple-negative GISTs. CONCLUSIONS: NF1 gene mutations are frequent in quadruple-negative GISTs and are often constitutional, indicating that a significant fraction of patients with apparently sporadic quadruple-negative GISTs are affected by unrecognized Neurofibromatosis Type 1 syndrome. Hence, a diagnosis of quadruple-negative GIST, especially if multifocal or with a multinodular growth pattern and a nongastric location, should alert the clinician to a possible Neurofibromatosis Type 1 syndromic condition. Clin Cancer Res; 23(1); 273-82. ©2016 AACR.


Assuntos
Biomarcadores Tumorais , Tumores do Estroma Gastrointestinal/genética , Neurofibromatose 1/genética , Adulto , Idoso , Idoso de 80 Anos ou mais , Alelos , Análise Mutacional de DNA , Feminino , Tumores do Estroma Gastrointestinal/diagnóstico , Tumores do Estroma Gastrointestinal/mortalidade , Tumores do Estroma Gastrointestinal/terapia , Genes da Neurofibromatose 1 , Genótipo , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Masculino , Pessoa de Meia-Idade , Mutação , Neurofibromatose 1/diagnóstico , Neurofibromatose 1/mortalidade , Neurofibromatose 1/terapia , Prognóstico , Proteínas Proto-Oncogênicas B-raf/genética , Proteínas Proto-Oncogênicas c-kit/genética , Receptor alfa de Fator de Crescimento Derivado de Plaquetas/genética
8.
Nat Genet ; 47(7): 757-65, 2015 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-26029869

RESUMO

Achromatopsia (ACHM) is an autosomal recessive disorder characterized by color blindness, photophobia, nystagmus and severely reduced visual acuity. Using homozygosity mapping and whole-exome and candidate gene sequencing, we identified ten families carrying six homozygous and two compound-heterozygous mutations in the ATF6 gene (encoding activating transcription factor 6A), a key regulator of the unfolded protein response (UPR) and cellular endoplasmic reticulum (ER) homeostasis. Patients had evidence of foveal hypoplasia and disruption of the cone photoreceptor layer. The ACHM-associated ATF6 mutations attenuate ATF6 transcriptional activity in response to ER stress. Atf6(-/-) mice have normal retinal morphology and function at a young age but develop rod and cone dysfunction with increasing age. This new ACHM-related gene suggests a crucial and unexpected role for ATF6A in human foveal development and cone function and adds to the list of genes that, despite ubiquitous expression, when mutated can result in an isolated retinal photoreceptor phenotype.


Assuntos
Fator 6 Ativador da Transcrição/genética , Defeitos da Visão Cromática/genética , Adolescente , Adulto , Idoso de 80 Anos ou mais , Animais , Criança , Feminino , Estudos de Associação Genética , Células HEK293 , Humanos , Masculino , Camundongos Endogâmicos C57BL , Camundongos Knockout , Pessoa de Meia-Idade , Mutação de Sentido Incorreto , Linhagem , Células Fotorreceptoras Retinianas Cones/patologia , Transcrição Genética , Resposta a Proteínas não Dobradas , Adulto Jovem
9.
Birth Defects Res A Clin Mol Teratol ; 103(12): 1003-10, 2015 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-26033879

RESUMO

BACKGROUND: Fanconi anemia (FA) is a rare genetic disease characterized by congenital malformations, aplastic anemia and increased risk of developing malignancies. FA is genetically heterogeneous as it is caused by at least 17 different genes. Among these, FANCA, FANCC, and FANCG account for approximately 85% of the patients whereas the remaining genes are mutated in only a small percentage of cases. For this reason, the molecular diagnostic process is complex and not always extended to all the FA genes, preventing the characterization of individuals belonging to rare groups. METHODS: The FA genes were analyzed using a next generation sequencing approach in two unrelated families. RESULTS: The analysis identified the same, c.484_485del, homozygous mutation of FANCF in both families. A careful examination of three electively aborted fetuses in one family and one affected girl in the other indicated an association of the FANCF loss-of-function mutation with a severe phenotype characterized by multiple malformations. CONCLUSION: The systematic use of next generation sequencing will allow the recognition of individuals from rare complementation groups, a better definition of their clinical phenotypes, and consequently, an appropriate genetic counseling.


Assuntos
Anemia de Fanconi/genética , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Mutação , Pré-Escolar , Feminino , Humanos , Masculino , Linhagem
10.
Elife ; 4: e06602, 2015 May 30.
Artigo em Inglês | MEDLINE | ID: mdl-26026149

RESUMO

Defective primary ciliogenesis or cilium stability forms the basis of human ciliopathies, including Joubert syndrome (JS), with defective cerebellar vermis development. We performed a high-content genome-wide small interfering RNA (siRNA) screen to identify genes regulating ciliogenesis as candidates for JS. We analyzed results with a supervised-learning approach, using SYSCILIA gold standard, Cildb3.0, a centriole siRNA screen and the GTex project, identifying 591 likely candidates. Intersection of this data with whole exome results from 145 individuals with unexplained JS identified six families with predominantly compound heterozygous mutations in KIAA0586. A c.428del base deletion in 0.1% of the general population was found in trans with a second mutation in an additional set of 9 of 163 unexplained JS patients. KIAA0586 is an orthologue of chick Talpid3, required for ciliogenesis and Sonic hedgehog signaling. Our results uncover a relatively high frequency cause for JS and contribute a list of candidates for future gene discoveries in ciliopathies.


Assuntos
Proteínas de Ciclo Celular/genética , Cerebelo/anormalidades , Predisposição Genética para Doença , Proteínas Mutantes/genética , Retina/anormalidades , Anormalidades Múltiplas/genética , Anormalidades do Olho/genética , Frequência do Gene , Testes Genéticos , Estudo de Associação Genômica Ampla , Heterozigoto , Humanos , Doenças Renais Císticas/genética , RNA Interferente Pequeno/genética
11.
Hum Genet ; 134(1): 123-6, 2015 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-25407461

RESUMO

Oral-facial-digital type VI syndrome (OFDVI) is a rare phenotype of Joubert syndrome (JS). Recently, C5orf42 was suggested as the major OFDVI gene, being mutated in 9 of 11 families (82 %). We sequenced C5orf42 in 313 JS probands and identified mutations in 28 (8.9 %), most with a phenotype of pure JS. Only 2 out of 17 OFDVI patients (11.7 %) were mutated. A comparison of mutated vs. non-mutated OFDVI patients showed that preaxial and mesoaxial polydactyly, hypothalamic hamartoma and other congenital defects may predict C5orf42 mutations, while tongue hamartomas are more common in negative patients.


Assuntos
Doenças Cerebelares/genética , Anormalidades do Olho/genética , Hamartoma/genética , Doenças Hipotalâmicas/genética , Doenças Renais Císticas/genética , Proteínas de Membrana/genética , Mutação/genética , Síndromes Orofaciodigitais/genética , Retina/anormalidades , Anormalidades Múltiplas , Doenças Cerebelares/patologia , Cerebelo/anormalidades , Estudos de Coortes , Anormalidades do Olho/patologia , Família , Feminino , Seguimentos , Hamartoma/patologia , Humanos , Doenças Hipotalâmicas/patologia , Doenças Renais Císticas/patologia , Masculino , Síndromes Orofaciodigitais/patologia , Fenótipo , Retina/patologia
12.
Am J Med Genet A ; 164A(8): 2084-90, 2014 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-24819041

RESUMO

NSD1 point mutations, submicroscopic deletions and intragenic deletions are the major cause of Sotos syndrome, characterized by pre-postnatal generalized overgrowth with advanced bone age, learning disability, seizures, distinctive facial phenotype. Reverse clinical phenotype due to 5q35 microduplication encompassing NSD1 gene has been reported so far in 27 cases presenting with delayed bone age, microcephaly, failure to thrive and seizures in some cases, further supporting a gene dosage effect of NSD1 on growth regulation and neurological functions. Here we depict the clinical presentation of three new cases with 5q35 microduplication outlining a novel syndrome characterized by microcephaly, short stature, developmental delay and in some cases delayed bone maturation, without any typical facial or osseous anomalies.


Assuntos
Deleção Cromossômica , Duplicação Cromossômica , Estudos de Associação Genética , Fenótipo , Síndrome de Sotos/diagnóstico , Síndrome de Sotos/genética , Adolescente , Pré-Escolar , Cromossomos Humanos Par 5 , Hibridização Genômica Comparativa , Facies , Feminino , Humanos , Hibridização in Situ Fluorescente , Masculino , Pessoa de Meia-Idade , Duplicações Segmentares Genômicas
13.
Gene ; 533(1): 398-402, 2014 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-24096233

RESUMO

BACKGROUND: Oculocutaneous Albinism (OCA) is a heterogeneous group of inherited diseases involving hair, skin and eyes. To date, six forms are recognized on the effects of different melanogenesis genes. OCA4 is caused by mutations in SLC45A2 showing a heterogeneous phenotype ranging from white hair, blue irides and nystagmus to brown/black hair, brown irides and no nystagmus. The high clinic variety often leads to misdiagnosis. Our aim is to contribute to OCA4 diagnosis defining SLC45A2 genetic variants in Italian patients with OCA without any TYR, OCA2 and TYRP1 gene defects. MATERIALS AND METHODS: After the clinical diagnosis of OCA, all patients received genetic counseling and genetic test. Automatic sequencing of TYR, OCA2, and TYRP1 genes was performed on DNA of 117 albino patients. Multiplex Ligation-dependent Probe Amplification (MLPA) was carried out on TYR and OCA2 genes to increase the mutation rate. SLC45A2 gene sequencing was then executed in the patients with a single mutation in one of the TYR, OCA2, TYRP1 genes and in the patients, which resulted negative at the screening of these genes. RESULTS: SLC45A2 gene analysis was performed in 41 patients and gene alterations were found in 5 patients. Four previously reported SLC45A2 mutations were found: p.G100S, p.W202C, p.A511E and c.986delC, and three novel variants were identified: p.M265L, p.H94D, and c.1156+1G>A. All the alterations have been detected in the group of patients without mutations in the other OCA genes. CONCLUSIONS: Three new variants were identified in OCA4 gene; the analysis allowed the classification of a patient previously misdiagnosed as OA1 because of skin and hair pigmentation presence. The molecular defects in SLC45A2 gene represent the 3.4% in this cohort of Italian patients, similar to other Caucasian populations; our data differ from those previously published by an Italian researcher group, obtained on a smaller cohort of patients.


Assuntos
Albinismo Oculocutâneo/genética , Antígenos de Neoplasias/genética , Proteínas de Membrana Transportadoras/genética , Mutação , Criança , Pré-Escolar , Estudos de Coortes , Feminino , Humanos , Lactente , Itália , Masculino
14.
Brain ; 136(Pt 12): 3634-44, 2013 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-24176978

RESUMO

Marinesco-Sjögren syndrome is a rare autosomal recessive multisystem disorder featuring cerebellar ataxia, early-onset cataracts, chronic myopathy, variable intellectual disability and delayed motor development. More recently, mutations in the SIL1 gene, which encodes an endoplasmic reticulum resident co-chaperone, were identified as the main cause of Marinesco-Sjögren syndrome. Here we describe the results of SIL1 mutation analysis in 62 patients presenting with early-onset ataxia, cataracts and myopathy or combinations of at least two of these. We obtained a mutation detection rate of 60% (15/25) among patients with the characteristic Marinesco-Sjögren syndrome triad (ataxia, cataracts, myopathy) whereas the detection rate in the group of patients with more variable phenotypic presentation was below 3% (1/37). We report 16 unrelated families with a total of 19 different SIL1 mutations. Among these mutations are 15 previously unreported changes, including single- and multi-exon deletions. Based on data from our screening cohort and data compiled from the literature we found that SIL1 mutations are invariably associated with the combination of a cerebellar syndrome and chronic myopathy. Cataracts were observed in all patients beyond the age of 7 years, but might be missing in infants. Six patients with SIL1 mutations had no intellectual disability, extending the known wide range of cognitive capabilities in Marinesco-Sjögren syndrome to include normal intelligence. Modestly constant features were somatic growth retardation, skeletal abnormalities and pyramidal tract signs. Examination of mutant SIL1 expression in cultured patient lymphoblasts suggested that SIL1 mutations result in severely reduced SIL1 protein levels irrespective of the type and position of mutations. Our data broaden the SIL1 mutation spectrum and confirm that SIL1 is the major Marinesco-Sjögren syndrome gene. SIL1 patients usually present with the characteristic triad but cataracts might be missing in young children. As cognitive impairment is not obligatory, patients without intellectual disability but a Marinesco-Sjögren syndrome-compatible phenotype should receive SIL1 mutation analysis. Despite allelic heterogeneity and many families with private mutations, the phenotype related to SIL1 mutations is relatively homogenous. Based on SIL1 expression studies we speculate that this may arise from a uniform effect of different mutations on protein expression.


Assuntos
Fatores de Troca do Nucleotídeo Guanina/genética , Mutação/genética , Degenerações Espinocerebelares/genética , Adolescente , Linfócitos B , Encéfalo/patologia , Encéfalo/ultraestrutura , Células Cultivadas , Análise Mutacional de DNA , Saúde da Família , Feminino , Humanos , Imagem por Ressonância Magnética , Masculino , Músculo Esquelético/patologia , Músculo Esquelético/ultraestrutura , Estudos Retrospectivos , Degenerações Espinocerebelares/patologia , Degenerações Espinocerebelares/fisiopatologia
15.
PLoS One ; 7(7): e41516, 2012.
Artigo em Inglês | MEDLINE | ID: mdl-22848519

RESUMO

Sandhoff disease (SD) is a lysosomal disorder caused by mutations in the HEXB gene. To date, 43 mutations of HEXB have been described, including 3 large deletions. Here, we have characterized 14 unrelated SD patients and developed a Multiplex Ligation-dependent Probe Amplification (MLPA) assay to investigate the presence of large HEXB deletions. Overall, we identified 16 alleles, 9 of which were novel, including 4 sequence variation leading to aminoacid changes [c.626C>T (p.T209I), c.634C>A (p.H212N), c.926G>T (p.C309F), c.1451G>A (p.G484E)] 3 intronic mutations (c.1082+5G>A, c.1242+1G>A, c.1169+5G>A), 1 nonsense mutation c.146C>A (p.S49X) and 1 small in-frame deletion c.1260_1265delAGTTGA (p.V421_E422del). Using the new MLPA assay, 2 previously described deletions were identified. In vitro expression studies showed that proteins bearing aminoacid changes p.T209I and p.G484E presented a very low or absent activity, while proteins bearing the p.H212N and p.C309F changes retained a significant residual activity. The detrimental effect of the 3 novel intronic mutations on the HEXB mRNA processing was demonstrated using a minigene assay. Unprecedentedly, minigene studies revealed the presence of a novel alternative spliced HEXB mRNA variant also present in normal cells. In conclusion, we provided new insights into the molecular basis of SD and validated an MLPA assay for detecting large HEXB deletions.


Assuntos
Processamento Alternativo/genética , Sequência de Bases , Doença de Sandhoff/genética , Deleção de Sequência , Cadeia beta da beta-Hexosaminidase/genética , Feminino , Células HEK293 , Humanos , Masculino , Reação em Cadeia da Polimerase Multiplex , RNA Mensageiro/biossíntese , RNA Mensageiro/genética , Doença de Sandhoff/metabolismo , Cadeia beta da beta-Hexosaminidase/metabolismo
16.
Am J Med Genet A ; 155A(11): 2860-4, 2011 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-21964829

RESUMO

Dominant mutations in the receptor calcium channel gene TRPV4 have been associated with a family of skeletal dysplasias (metatropic dysplasia, pseudo-Morquio type 2, spondylometaphyseal dysplasia, Kozlowski type, brachyolmia, and familial digital arthropathy) as well as with dominantly inherited neuropathies (hereditary motor and sensory neuropathy 2C, scapuloperoneal spinal muscular atrophy, and congenital distal spinal muscular atrophy). While there is phenotypic overlap between the various members of each group, the two groups were considered to be totally separate with the former being strictly a structural skeletal condition and the latter group being confined to the peripheral nervous system. We report here on fetal akinesia as the presenting feature of severe metatropic dysplasia, suggesting that certain TRPV4 mutations can cause both a skeletal and a neuropathic phenotype. Three cases were detected on prenatal ultrasound because of absent movements in the second trimester. Case 4 presented with multiple joint contractures and absent limb movements at birth and was diagnosed with "fetal akinesia syndrome". Post-interruption and post-natal X-rays showed typical features of metatropic dysplasia in all four. Sequencing of the TRPV4 gene confirmed the presence of de novo heterozygous mutations predicting G78W (Case 1), T740I (Cases 2 and 3), and K276E (Case 4). Although some degree of restriction of movements is not uncommon in fetuses with skeletal dysplasia, akinesia as leading sign is unusual and suggests that certain TRPV4 mutations produce both chondrodysplasia and a peripheral neuropathy resulting in a severe "overlap" phenotype.


Assuntos
Retardo do Crescimento Fetal/patologia , Atrofia Muscular Espinal/patologia , Osteocondrodisplasias/patologia , Fenótipo , Feminino , Retardo do Crescimento Fetal/diagnóstico , Retardo do Crescimento Fetal/genética , Feto/anormalidades , Feto/patologia , Heterozigoto , Humanos , Lactente , Recém-Nascido , Deformidades Congênitas dos Membros/diagnóstico , Deformidades Congênitas dos Membros/genética , Deformidades Congênitas dos Membros/patologia , Masculino , Atrofia Muscular Espinal/diagnóstico , Atrofia Muscular Espinal/genética , Mutação , Osteocondrodisplasias/diagnóstico , Osteocondrodisplasias/genética , Gravidez , Segundo Trimestre da Gravidez , Canais de Cátion TRPV/genética , Ultrassonografia Pré-Natal
17.
Orphanet J Rare Dis ; 6: 40, 2011 Jun 16.
Artigo em Inglês | MEDLINE | ID: mdl-21679407

RESUMO

BACKGROUND: The breadth of the clinical spectrum underlying Pelizaeus-Merzbacher disease and spastic paraplegia type 2 is due to the extensive allelic heterogeneity in the X-linked PLP1 gene encoding myelin proteolipid protein (PLP). PLP1 mutations range from gene duplications of variable size found in 60-70% of patients to intragenic lesions present in 15-20% of patients. METHODS: Forty-eight male patients from 38 unrelated families with a PLP1-related disorder were studied. All DNA samples were screened for PLP1 gene duplications using real-time PCR. PLP1 gene sequencing analysis was performed on patients negative for the duplication. The mutational status of all 14 potential carrier mothers of the familial PLP1 gene mutation was determined as well as 15/24 potential carrier mothers of the PLP1 duplication. RESULTS AND CONCLUSIONS: PLP1 gene duplications were identified in 24 of the unrelated patients whereas a variety of intragenic PLP1 mutations were found in the remaining 14 patients. Of the 14 different intragenic lesions, 11 were novel; these included one nonsense and 7 missense mutations, a 657-bp deletion, a microdeletion and a microduplication. The functional significance of the novel PLP1 missense mutations, all occurring at evolutionarily conserved residues, was analysed by the MutPred tool whereas their potential effect on splicing was ascertained using the Skippy algorithm and a neural network. Although MutPred predicted that all 7 novel missense mutations would be likely to be deleterious, in silico analysis indicated that four of them (p.Leu146Val, p.Leu159Pro, p.Thr230Ile, p.Ala247Asp) might cause exon skipping by altering exonic splicing elements. These predictions were then investigated in vitro for both p.Leu146Val and p.Thr230Ile by means of RNA or minigene studies and were subsequently confirmed in the case of p.Leu146Val. Peripheral neuropathy was noted in four patients harbouring intragenic mutations that altered RNA processing, but was absent from all PLP1-duplication patients. Unprecedentedly, family studies revealed the de novo occurrence of the PLP1 duplication at a frequency of 20%.


Assuntos
Duplicação Gênica , Proteína Proteolipídica de Mielina/genética , Doença de Pelizaeus-Merzbacher/genética , Paraplegia Espástica Hereditária/genética , Adolescente , Criança , Pré-Escolar , DNA/genética , Humanos , Lactente , Masculino , Adulto Jovem
18.
Hum Mutat ; 32(7): 760-72, 2011 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-21387466

RESUMO

Noonan syndrome (NS) is among the most common nonchromosomal disorders affecting development and growth. NS is caused by aberrant RAS-MAPK signaling and is genetically heterogeneous, which explains, in part, the marked clinical variability documented for this Mendelian trait. Recently, we and others identified SOS1 as a major gene underlying NS. Here, we explored further the spectrum of SOS1 mutations and their associated phenotypic features. Mutation scanning of the entire SOS1 coding sequence allowed the identification of 33 different variants deemed to be of pathological significance, including 16 novel missense changes and in-frame indels. Various mutation clusters destabilizing or altering orientation of regions of the protein predicted to contribute structurally to the maintenance of autoinhibition were identified. Two previously unappreciated clusters predicted to enhance SOS1's recruitment to the plasma membrane, thus promoting a spatial reorientation of domains contributing to inhibition, were also recognized. Genotype-phenotype analysis confirmed our previous observations, establishing a high frequency of ectodermal anomalies and a low prevalence of cognitive impairment and reduced growth. Finally, mutation analysis performed on cohorts of individuals with nonsyndromic pulmonic stenosis, atrial septal defects, and ventricular septal defects excluded a major contribution of germline SOS1 lesions to the isolated occurrence of these cardiac anomalies.


Assuntos
Síndrome de Noonan/diagnóstico , Síndrome de Noonan/genética , Proteína SOS1/genética , Adolescente , Adulto , Criança , Éxons , Feminino , Estudos de Associação Genética , Comunicação Interatrial/genética , Comunicação Interventricular/genética , Humanos , Mutação INDEL/genética , Íntrons , Masculino , Quinases de Proteína Quinase Ativadas por Mitógeno/genética , Mutação , Mutação de Sentido Incorreto/genética , Conformação Proteica , Estenose da Valva Pulmonar/genética , Proteína SOS1/química
19.
Hum Mutat ; 30(4): 695-702, 2009 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-19206169

RESUMO

Noonan, LEOPARD, and cardiofaciocutaneous syndromes (NS, LS, and CFCS) are developmental disorders with overlapping features including distinctive facial dysmorphia, reduced growth, cardiac defects, skeletal and ectodermal anomalies, and variable cognitive deficits. Dysregulated RAS-mitogen-activated protein kinase (MAPK) signal traffic has been established to represent the molecular pathogenic cause underlying these conditions. To investigate the phenotypic spectrum and molecular diversity of germline mutations affecting BRAF, which encodes a serine/threonine kinase functioning as a RAS effector frequently mutated in CFCS, subjects with a diagnosis of NS (N=270), LS (N=6), and CFCS (N=33), and no mutation in PTPN11, SOS1, KRAS, RAF1, MEK1, or MEK2, were screened for the entire coding sequence of the gene. Besides the expected high prevalence of mutations observed among CFCS patients (52%), a de novo heterozygous missense change was identified in one subject with LS (17%) and five individuals with NS (1.9%). Mutations mapped to multiple protein domains and largely did not overlap with cancer-associated defects. NS-causing mutations had not been documented in CFCS, suggesting that the phenotypes arising from germline BRAF defects might be allele specific. Selected mutant BRAF proteins promoted variable gain of function of the kinase, but appeared less activating compared to the recurrent cancer-associated p.Val600Glu mutant. Our findings provide evidence for a wide phenotypic diversity associated with mutations affecting BRAF, and occurrence of a clinical continuum associated with these molecular lesions.


Assuntos
Anormalidades Múltiplas/genética , Mutação em Linhagem Germinativa , Síndrome LEOPARD/genética , Síndrome de Noonan/genética , Proteínas Proto-Oncogênicas B-raf/genética , Anormalidades Múltiplas/patologia , Estudos de Coortes , Face/anormalidades , Feminino , Frequência do Gene , Variação Genética , Genótipo , Cardiopatias Congênitas/patologia , Humanos , Síndrome LEOPARD/patologia , Masculino , Mutação de Sentido Incorreto , Síndrome de Noonan/patologia , Fenótipo , Anormalidades da Pele
20.
Hum Mutat ; 22(2): 179-80, 2003 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-12872266

RESUMO

In the present study the entire NF1 coding region was analyzed for mutations in 132 unrelated Italian NF1 patients. Using PTT, SSCP, and DNA sequencing, we found 8 novel mutations. Clinical diagnosis of NF1 was established according to the NIH consensus criteria. We detected 59 truncated fragments, and 46 of them were characterized by SSCP and direct sequencing. Eight mutations represent novel changes that contribute to the germline mutational spectrum of the NF1 gene. In two patients, premature termination was due to substitutions at nucleotide c.3982C>T (Q1298X) and c.7411C>T (Q2471X), respectively. Two other mutations were caused by the deletions (1756delA, 4699delA), and two by the insertions (c.5266_5267insT, c.7464_7465insTCCA) of a small number of nucleotides. Lastly, we found 2 splice-site mutations (c.2252-2A>C, c.2251+1G>A).


Assuntos
Genes da Neurofibromatose 1 , Mutação/genética , Neurofibromatose 1/genética , Análise Mutacional de DNA , Mutação da Fase de Leitura/genética , Humanos , Itália , Mutagênese Insercional/genética , Deleção de Sequência/genética
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