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1.
Cell Rep ; 32(12): 108185, 2020 09 22.
Artigo em Inglês | MEDLINE | ID: covidwho-743905

RESUMO

One of the features distinguishing SARS-CoV-2 from its more pathogenic counterpart SARS-CoV is the presence of premature stop codons in its ORF3b gene. Here, we show that SARS-CoV-2 ORF3b is a potent interferon antagonist, suppressing the induction of type I interferon more efficiently than its SARS-CoV ortholog. Phylogenetic analyses and functional assays reveal that SARS-CoV-2-related viruses from bats and pangolins also encode truncated ORF3b gene products with strong anti-interferon activity. Furthermore, analyses of approximately 17,000 SARS-CoV-2 sequences identify a natural variant in which a longer ORF3b reading frame was reconstituted. This variant was isolated from two patients with severe disease and further increased the ability of ORF3b to suppress interferon induction. Thus, our findings not only help to explain the poor interferon response in COVID-19 patients but also describe the emergence of natural SARS-CoV-2 quasispecies with an extended ORF3b gene that may potentially affect COVID-19 pathogenesis.


Assuntos
Betacoronavirus/genética , Infecções por Coronavirus/patologia , Interferon Tipo I/antagonistas & inibidores , Pneumonia Viral/patologia , Proteínas Virais Reguladoras e Acessórias/genética , Adulto , Sequência de Aminoácidos/genética , Animais , Betacoronavirus/imunologia , Quirópteros/virologia , Códon sem Sentido/genética , Eutérios/virologia , Humanos , Masculino , Pandemias , Proteínas Virais Reguladoras e Acessórias/metabolismo
2.
Cell Rep ; 32(12): 108185, 2020 09 22.
Artigo em Inglês | MEDLINE | ID: mdl-32941788

RESUMO

One of the features distinguishing SARS-CoV-2 from its more pathogenic counterpart SARS-CoV is the presence of premature stop codons in its ORF3b gene. Here, we show that SARS-CoV-2 ORF3b is a potent interferon antagonist, suppressing the induction of type I interferon more efficiently than its SARS-CoV ortholog. Phylogenetic analyses and functional assays reveal that SARS-CoV-2-related viruses from bats and pangolins also encode truncated ORF3b gene products with strong anti-interferon activity. Furthermore, analyses of approximately 17,000 SARS-CoV-2 sequences identify a natural variant in which a longer ORF3b reading frame was reconstituted. This variant was isolated from two patients with severe disease and further increased the ability of ORF3b to suppress interferon induction. Thus, our findings not only help to explain the poor interferon response in COVID-19 patients but also describe the emergence of natural SARS-CoV-2 quasispecies with an extended ORF3b gene that may potentially affect COVID-19 pathogenesis.


Assuntos
Betacoronavirus/genética , Infecções por Coronavirus/patologia , Interferon Tipo I/antagonistas & inibidores , Pneumonia Viral/patologia , Proteínas Virais Reguladoras e Acessórias/genética , Adulto , Sequência de Aminoácidos/genética , Animais , Betacoronavirus/imunologia , Quirópteros/virologia , Códon sem Sentido/genética , Eutérios/virologia , Humanos , Masculino , Pandemias , Proteínas Virais Reguladoras e Acessórias/metabolismo
3.
Medicine (Baltimore) ; 99(34): e21843, 2020 Aug 21.
Artigo em Inglês | MEDLINE | ID: mdl-32846832

RESUMO

RATIONALE: Hypertrophic cardiomyopathy (HCM) is an inherited myocardial disease and a common cause of sudden cardiac death, heart failure, atrial fibrillation and stroke. In families affected by HCM, genotyping is useful for identifying susceptible relatives. In the present study, we investigated the disease-causing mutations in a three-generation Chinese family with HCM using whole exome sequencing (WES). PATIENT CONCERNS: The proband, a 50-year-old man, was diagnosed with HCM at the age of 41 years. He presented with an asymmetric hypertrophic interventricular septum and a maximum interventricular septum thickness of 18.04 mm. His third elder sister, niece and daughter were also clinically affected by HCM. DIAGNOSIS: Autosomal dominant HCM. INTERVENTIONS: Seven family members, including 4 affected members, accepted WES. The genetic variants were subsequently called using Genome Analysis Toolkit and annotated using the InterVar program. Following frequency filtration by the Genome Aggregation Database, the variants were evaluated using an in-house bioinformatics analysis pipeline. OUTCOMES: HCM was transmitted as an autosomal dominant trait in the family. An extremely rare stop gained mutation, rs796925245 (g.1:201359630G>A, c.835C>T, p.Gln279Ter) in the troponin T2 (TNNT2) gene was identified as the disease-causing mutation. The stop gained mutation was predicted to result in a truncated troponin T protein in cardiac sarcomere. An adolescent family member who had normal echocardiographic measurements was found to carry the same disease-causing mutation. LESSONS: A novel nonsense TNNT2 mutation was identified as the HCM-causing mutation in this Chinese pedigree. Since HCM shows a low penetrance by clinical criteria in adolescents, the adolescent mutation carrier, who is still clinically unaffected, should be offered routine follow-ups and sport activity recommendations to prevent adverse events including sudden cardiac death in the future.


Assuntos
Cardiomiopatia Hipertrófica Familiar/genética , Troponina T/genética , Adolescente , Adulto , Idoso , Grupo com Ancestrais do Continente Asiático/genética , Cardiomiopatia Hipertrófica Familiar/complicações , Cardiomiopatia Hipertrófica Familiar/diagnóstico , Códon sem Sentido , Morte Súbita Cardíaca/etiologia , Ecocardiografia/métodos , Feminino , Humanos , Hipertrofia/diagnóstico por imagem , Masculino , Pessoa de Meia-Idade , Linhagem , Fenótipo , Septo Interventricular/patologia , Sequenciamento Completo do Exoma/métodos
5.
PLoS One ; 15(8): e0237803, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32813700

RESUMO

A nonsense mutation adds a premature stop signal that hinders any further translation of a protein-coding gene, usually resulting in a null allele. To investigate the possible exceptions, we used the DMD gene as an ideal model. First, because dystrophin absence causes Duchenne muscular dystrophy (DMD), while its reduction causes Becker muscular dystrophy (BMD). Second, the DMD gene is X-linked and there is no second allele that can interfere in males. Third, databases are accumulating reports on many mutations and phenotypic data. Finally, because DMD mutations may have important therapeutic implications. For our study, we analyzed large databases (LOVD, HGMD and ClinVar) and literature and revised critically all data, together with data from our internal patients. We totally collected 2593 patients. Positioning these mutations along the dystrophin transcript, we observed a nonrandom distribution of BMD-associated mutations within selected exons and concluded that the position can be predictive of the phenotype. Nonsense mutations always cause DMD when occurring at any point in fifty-one exons. In the remaining exons, we found milder BMD cases due to early 5' nonsense mutations, if reinitiation can occur, or due to late 3' nonsense when the shortened product retains functionality. In the central part of the gene, all mutations in some in-frame exons, such as in exons 25, 31, 37 and 38 cause BMD, while mutations in exons 30, 32, 34 and 36 cause DMD. This may have important implication in predicting the natural history and the efficacy of therapeutic use of drug-stimulated translational readthrough of premature termination codons, also considering the action of internal natural rescuers. More in general, our survey confirm that a nonsense mutation should be not necessarily classified as a null allele and this should be considered in genetic counselling.


Assuntos
Códon sem Sentido/genética , Distrofina/genética , Distrofia Muscular de Duchenne/genética , Distrofia Muscular de Duchenne/patologia , Mutação/genética , Inquéritos e Questionários , Sequência de Aminoácidos , Sequência de Bases , Distrofina/química , Éxons/genética , Humanos , Fenótipo
6.
PLoS One ; 15(8): e0236808, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32750061

RESUMO

BACKGROUND: Ataxia with oculomotor apraxia type 1 (AOA1) is a rare autosomal recessive cerebellar ataxia, caused by mutations in the APTX gene. The disease is characterized by early-onset cerebellar ataxia, oculomotor apraxia and severe axonal polyneuropathy. The aim of this study was to detect the disease-causing variants in two unrelated consanguineous Jordanian families with cerebellar ataxia using whole exome sequencing (WES), and to correlate the identified mutation(s) with the clinical and cellular phenotypes. METHODS: WES was performed in three affected individuals and segregation analysis of p.W279* APTX candidate variant was performed. Expression levels of APTX were measured in patients' skin fibroblasts and peripheral blood mononuclear cells, followed by western blot analysis in skin fibroblasts. Genotoxicity assay was performed to detect the sensitivity of APTX mutated cells to H2O2, MMC, MMS and etoposide. RESULTS: A recurrent homozygous nonsense variant in APTX gene (c.837G>A, p.W279*) was revealed in all affected individuals. qRT-PCR showed normal APTX levels in peripheral blood and lower levels in fibroblast cells. However, western blot showed the absence of APTX protein in patients' skin fibroblasts. Significant hypersensitivity to H2O2, MMC and etoposide and lack of sensitivity to MMS were noted. CONCLUSIONS: This is the first study to report the identification of a nonsense variant in the APTX gene (c.837G>A; p.W279*) in AOA1 patients within the Jordanian population. This study confirmed the need of WES to assist in the diagnosis of cerebellar ataxia and it emphasizes the importance of studying the pathophysiology of the APTX gene.


Assuntos
Ataxia Cerebelar/genética , Códon sem Sentido , Dano ao DNA , Proteínas de Ligação a DNA/genética , Proteínas Nucleares/genética , Criança , Pré-Escolar , Consanguinidade , DNA/efeitos dos fármacos , Feminino , Humanos , Masculino , Mutagênicos/farmacologia , Sequenciamento Completo do Exoma
7.
Gene ; 761: 145023, 2020 Nov 30.
Artigo em Inglês | MEDLINE | ID: mdl-32758581

RESUMO

The clinical pictures of the disease of two Russian patients with cystic fibrosis with a rare nonsense variant c.831G>A (p.Trp277*) are described. The first case is a patient with the genotype comprising variant c.54-5940_273+10250del21kb (CFTRdele2,3), and the genotype of the second case included variant c.1521_1523delCTT (F508del). Patient 1, whose genotype had two class I genetic variants, revealed severe violations of CFTR synthesis based on the intestinal current measurements (ICM) and results obtained in the intestinal organoids. In both cases of patients with genetic variant c.831G>A, a severe course of cystic fibrosis was observed.


Assuntos
Canais de Cloreto/genética , Fibrose Cística/genética , Criança , Códon sem Sentido/genética , Fibrose Cística/metabolismo , Regulador de Condutância Transmembrana em Fibrose Cística/genética , Feminino , Genótipo , Humanos , Masculino , Mutação , Federação Russa
8.
PLoS Genet ; 16(8): e1008691, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32764743

RESUMO

Primary ciliary dyskinesia (PCD) is characterized by chronic airway disease, reduced fertility, and randomization of the left/right body axis. It is caused by defects of motile cilia and sperm flagella. We screened a cohort of affected individuals that lack an obvious axonemal defect for pathogenic variants using whole exome capture, next generation sequencing, and bioinformatic analysis assuming an autosomal recessive trait. We identified one subject with an apparently homozygous nonsense variant [(c.1762C>T), p.(Arg588*)] in the uncharacterized CFAP57 gene. Interestingly, the variant results in the skipping of exon 11 (58 amino acids), which may be due to disruption of an exonic splicing enhancer. In normal human nasal epithelial cells, CFAP57 localizes throughout the ciliary axoneme. Nasal cells from the PCD patient express a shorter, mutant version of CFAP57 and the protein is not incorporated into the axoneme. The missing 58 amino acids include portions of WD repeats that may be important for loading onto the intraflagellar transport (IFT) complexes for transport or docking onto the axoneme. A reduced beat frequency and an alteration in ciliary waveform was observed. Knockdown of CFAP57 in human tracheobronchial epithelial cells (hTECs) recapitulates these findings. Phylogenetic analysis showed that CFAP57 is highly conserved in organisms that assemble motile cilia. CFAP57 is allelic with the BOP2/IDA8/FAP57 gene identified previously in Chlamydomonas reinhardtii. Two independent, insertional fap57 Chlamydomonas mutant strains show reduced swimming velocity and altered waveforms. Tandem mass tag (TMT) mass spectroscopy shows that FAP57 is missing, and the "g" inner dyneins (DHC7 and DHC3) and the "d" inner dynein (DHC2) are reduced, but the FAP57 paralog FBB7 is increased. Together, our data identify a homozygous variant in CFAP57 that causes PCD that is likely due to a defect in the inner dynein arm assembly process.


Assuntos
Axonema/metabolismo , Transtornos da Motilidade Ciliar/genética , Códon sem Sentido , Dineínas/metabolismo , Proteínas/genética , Células 3T3 , Adulto , Animais , Axonema/fisiologia , Células Cultivadas , Chlamydomonas reinhardtii , Cílios/metabolismo , Cílios/fisiologia , Transtornos da Motilidade Ciliar/patologia , Sequência Conservada , Humanos , Masculino , Camundongos , Proteínas de Plantas/química , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Proteínas/química , Proteínas/metabolismo , Mucosa Respiratória/metabolismo
9.
PLoS One ; 15(7): e0236597, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32735634

RESUMO

INTRODUCTION: McArdle disease presents clinical and genetic heterogeneity. There is no obvious association between genotype and phenotype. PYGM (muscle glycogen phosphorylase gene) mRNA expression and its association with clinical, morphological, and genetic aspects of the disease as a set have not been studied previously. METHODS: We investigated genetic variation in PYGM considering the number of PTCs (premature termination codon) per sample and compared mRNA expression in skeletal muscle samples from 15 patients with McArdle disease and 16 controls to PTCs number and different aspects of the disease. RESULTS: The main variant found was c.148C>T (PTC-premature termination codon). Patients with two PTCs showed 42% mRNA expression compared to the control group. Most cases showed an inversely proportional relation among PTCs and mRNA expression. Association between mRNA expression and other aspects of the disease showed no statistically significant difference (p> 0.05). DISCUSSION: mRNA expression is not useful as a predictor factor for the prognosis and severity of the disease. Different mechanisms as post-transcriptional events, epigenetics factors or protein function may be involved.


Assuntos
Demografia , Regulação Enzimológica da Expressão Gênica , Glicogênio Fosforilase Muscular/genética , Doença de Depósito de Glicogênio Tipo V/genética , Adulto , Códon sem Sentido/genética , Estudos Transversais , Feminino , Doença de Depósito de Glicogênio Tipo V/epidemiologia , Doença de Depósito de Glicogênio Tipo V/patologia , Humanos , Masculino , Pessoa de Meia-Idade , RNA Mensageiro/genética , Adulto Jovem
10.
Proc Natl Acad Sci U S A ; 117(28): 16456-16464, 2020 07 14.
Artigo em Inglês | MEDLINE | ID: mdl-32616572

RESUMO

Duchenne muscular dystrophy (DMD) is caused by mutations in the DMD gene leading to the presence of premature termination codons (PTC). Previous transcriptional studies have shown reduced DMD transcript levels in DMD patient and animal model muscles when PTC are present. Nonsense-mediated decay (NMD) has been suggested to be responsible for the observed reduction, but there is no experimental evidence supporting this claim. In this study, we aimed to investigate the mechanism responsible for the drop in DMD expression levels in the presence of PTC. We observed that the inhibition of NMD does not normalize DMD gene expression in DMD. Additionally, in situ hybridization showed that DMD messenger RNA primarily localizes in the nuclear compartment, confirming that a cytoplasmic mechanism like NMD indeed cannot be responsible for the observed reduction. Sequencing of nascent RNA to explore DMD transcription dynamics revealed a lower rate of DMD transcription in patient-derived myotubes compared to healthy controls, suggesting a transcriptional mechanism involved in reduced DMD transcript levels. Chromatin immunoprecipitation in muscle showed increased levels of the repressive histone mark H3K9me3 in mdx mice compared to wild-type mice, indicating a chromatin conformation less prone to transcription in mdx mice. In line with this finding, treatment with the histone deacetylase inhibitor givinostat caused a significant increase in DMD transcript expression in mdx mice. Overall, our findings show that transcription dynamics across the DMD locus are affected by the presence of PTC, hinting at a possible epigenetic mechanism responsible for this process.


Assuntos
Códon sem Sentido/genética , Distrofina/genética , Distrofia Muscular de Duchenne/genética , RNA Mensageiro/genética , Animais , Códon sem Sentido/metabolismo , Modelos Animais de Doenças , Distrofina/metabolismo , Humanos , Camundongos , Camundongos Endogâmicos mdx , Distrofia Muscular de Duchenne/metabolismo , Degradação do RNAm Mediada por Códon sem Sentido , RNA Mensageiro/metabolismo
11.
Proc Natl Acad Sci U S A ; 117(27): 15799-15808, 2020 07 07.
Artigo em Inglês | MEDLINE | ID: mdl-32571908

RESUMO

The transcriptome of eukaryotic cells is constantly monitored for errors to avoid the production of undesired protein variants. The evolutionarily conserved nonsense-mediated mRNA decay (NMD) pathway degrades aberrant mRNAs, but also functions in the regulation of transcript abundance in response to changed physiological states. Here, we describe a zebrafish mutant of upf1, encoding the central component of the NMD machinery. Fish homozygous for the upf1 t20450 allele (Y163X) survive until day 10 after fertilization, presenting with impaired T cell development as one of the most conspicuous features of the mutant phenotype. Analysis of differentially expressed genes identified dysregulation of the pre-mRNA splicing pathway, accompanied by perturbed autoregulation of canonical splicing activators (SRSF) and repressors (HNRNP). In upf1-deficient mutants, NMD-susceptible transcripts of ribosomal proteins that are known for their role as noncanonical splicing regulators were greatly increased, most notably, rpl10a When the levels of NMD-susceptible rpl10a transcripts were artificially increased in zebrafish larvae, T cell development was significantly impaired, suggesting that perturbed autoregulation of rpl10a splicing contributes to failing T cell development in upf1 deficiency. Our results identify an extraribosomal tissue-specific function to rpl10a in the immune system, and thus exemplify the advantages of the zebrafish model to study the effects of upf1-deficiency in the context of a vertebrate organism.


Assuntos
Glutationa/análogos & derivados , Degradação do RNAm Mediada por Códon sem Sentido/genética , Processamento de RNA/genética , Proteínas de Ligação a RNA/genética , Linfócitos T/imunologia , Proteínas de Peixe-Zebra/genética , Animais , Códon sem Sentido/genética , Fertilização/genética , Regulação da Expressão Gênica no Desenvolvimento/genética , Glutationa/genética , Homozigoto , Humanos , Degradação do RNAm Mediada por Códon sem Sentido/imunologia , RNA Mensageiro/genética , Fatores de Transcrição/genética , Transcriptoma/genética , Peixe-Zebra/genética
12.
Nat Commun ; 11(1): 3106, 2020 06 19.
Artigo em Inglês | MEDLINE | ID: mdl-32561765

RESUMO

Nonsense-mediated mRNA decay (NMD) typifies an mRNA surveillance pathway. Because NMD necessitates a translation event to recognize a premature termination codon on mRNAs, truncated misfolded polypeptides (NMD-polypeptides) could potentially be generated from NMD substrates as byproducts. Here, we show that when the ubiquitin-proteasome system is overwhelmed, various misfolded polypeptides including NMD-polypeptides accumulate in the aggresome: a perinuclear nonmembranous compartment eventually cleared by autophagy. Hyperphosphorylation of the key NMD factor UPF1 is required for selective targeting of the misfolded polypeptide aggregates toward the aggresome via the CTIF-eEF1A1-DCTN1 complex: the aggresome-targeting cellular machinery. Visualization at a single-particle level reveals that UPF1 increases the frequency and fidelity of movement of CTIF aggregates toward the aggresome. Furthermore, the apoptosis induced by proteotoxic stresses is suppressed by UPF1 hyperphosphorylation. Altogether, our data provide evidence that UPF1 functions in the regulation of a protein surveillance as well as an mRNA quality control.


Assuntos
Degradação do RNAm Mediada por Códon sem Sentido , Complexo de Endopeptidases do Proteassoma/metabolismo , RNA Helicases/metabolismo , RNA Mensageiro/metabolismo , Transativadores/metabolismo , Resposta a Proteínas não Dobradas/genética , Autofagia , Códon sem Sentido , Complexo Dinactina/metabolismo , Fatores de Iniciação em Eucariotos/metabolismo , Células HEK293 , Células HeLa , Humanos , Imagem Molecular , Fator 1 de Elongação de Peptídeos/metabolismo , Fosforilação , Agregados Proteicos/genética , Imagem Individual de Molécula , Ubiquitina/metabolismo
13.
Tunis Med ; 98(5): 420-422, 2020 May.
Artigo em Inglês | MEDLINE | ID: mdl-32548846

RESUMO

Cystic Fibrosis (CF) is a lethal autosomal recessive condition due to a defect at the level of the transmembrane conductance regulator gene which plays a role in cell homeostasis. Numerous mutations have been identified as the cause of this gene defect, with delF508 being one of the most common mutations in Tunisia. This is a case report describing, up to our knowledge, the second case of a patient with CF carrying a rare mutation: W19X. W19X is a nonsense mutation that has been previously identified in only one other Tunisian patient with CF. Since both incidence of this mutation have been described in Tunisia, it seems as if W19X is specific to Tunisian CF patient with significant morbidities. The information provided by this study contributes to defining the molecular spectrum of CF in Tunisia, in the aim of improving genetic testing and prenatal diagnosis.


Assuntos
Códon sem Sentido , Regulador de Condutância Transmembrana em Fibrose Cística/genética , Fibrose Cística/genética , Anormalidades Múltiplas/diagnóstico , Anormalidades Múltiplas/genética , Anormalidades Múltiplas/patologia , Fibrose Cística/diagnóstico , Fibrose Cística/patologia , Frequência do Gene , Testes Genéticos , Humanos , Triptofano/genética , Tunísia
14.
PLoS One ; 15(5): e0232637, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32365113

RESUMO

ADAMTS13 regulates the hemostatic activity of von Willebrand factor (VWF). Determined by static assays, proteolytic activity <10IU/dL in patient plasma, in absence of ADAMTS13 autoantibodies, indicates Upshaw-Schulman syndrome (USS); the congenital form of Thrombotic Thrombocytopenic Purpura (TTP). We have recently functionally characterized sixteen USS-associated ADAMTS13 missense variants under static conditions. Here, we used two assays under shear flow conditions to analyze the activity of those seven mutants with sufficiently high residual secretion plus two newly identified variants. One assay determines cleavage of VWF strings bound to the surface of endothelial cells. The other, light transmission aggregometry-based assay, mimics degradation of VWF-platelet complexes, which are likely to be present in the circulation during TTP bouts. We found that 100 ng/ml of all variants were able to cleave about 80-90% of VWF strings even though 5 out of 9 exhibited activity ≤1% in the state-of-the-art static assay at the same concentration. These data indicate underestimation of ADAMTS13 activity by the used static assay. In simulated circulation, two variants, with missense mutations in the vicinity of the catalytic domain, exhibited only minor residual activity while all other variants were able to effectively break down VWF-platelet complexes. In both assays, significant proteolytic activity could be observed down to 100 ng/ml ADAMTS13. It is thus intriguing to postulate that most variants would have ample activity if secretion of 10% of normal plasma levels could be achieved.


Assuntos
Proteína ADAMTS13/genética , Variação Genética , Mutação de Sentido Incorreto , Púrpura Trombocitopênica Trombótica/congênito , Púrpura Trombocitopênica Trombótica/genética , Plaquetas/metabolismo , Domínio Catalítico , Códon sem Sentido , Células Endoteliais/metabolismo , Células HEK293 , Hemostasia , Humanos , Agregação Plaquetária , Proteínas Recombinantes/genética , Resistência ao Cisalhamento , Fatores de Tempo , Fator de von Willebrand
15.
Zhong Nan Da Xue Xue Bao Yi Xue Ban ; 45(2): 198-203, 2020 Feb 28.
Artigo em Inglês, Chinês | MEDLINE | ID: mdl-32386048

RESUMO

Rubinstein-Taybi syndrome (RSTS), also known as broad thumb-great toe syndrome or broad digits syndrome, is a rare autosomal dominant genetic disease. The main features of the patients are craniofacial dysmorphisms, skeletal malformations, and delay of growth and psychomotor development. In this case, the child has a typical RSTS specific face and growth retardation, with atypical indirect inguinalhemia. A heterozygous mutation, C. 4492 C>T (p. Arg1498Ter), was found in the exon of CREBBP gene by gene sequencing. It was a nonsense mutation, which leads to the premature termination of peptide synthesis. The mutation was not observed in the child's parents, which may be a de Novo mutation. The disease is lack of effective therapy so far.


Assuntos
Proteína de Ligação a CREB/genética , Síndrome de Rubinstein-Taybi , Códon sem Sentido , Éxons , Humanos , Lactente , Mutação
16.
BMC Med Genet ; 21(1): 97, 2020 05 07.
Artigo em Inglês | MEDLINE | ID: mdl-32380970

RESUMO

BACKGROUND: Amelogenesis imperfecta (AI) is a highly heterogeneous group of hereditary developmental abnormalities which mainly affects the dental enamel during tooth development in terms of its thickness, structure, and composition. It appears both in syndromic as well as non-syndromic forms. In the affected individuals, the enamel is usually thin, soft, rough, brittle, pitted, chipped, and abraded, having reduced functional ability and aesthetics. It leads to severe complications in the patient, like early tooth loss, severe discomfort, pain, dental caries, chewing difficulties, and discoloration of teeth from yellow to yellowish-brown or creamy type. The study aimed to identify the disease-causing variant in a consanguineous family. METHODS: We recruited a consanguineous Pashtun family of Pakistani origin. Exome sequencing analysis was followed by Sanger sequencing to identify the pathogenic variant in this family. RESULTS: Clinical analysis revealed hypomaturation AI having generalized yellow-brown or creamy type of discoloration in affected members. We identified a novel nonsense sequence variant c.1192C > T (p.Gln398*) in exon-12 of SLC24A4 by using exome sequencing. Later, its co-segregation within the family was confirmed by Sanger sequencing. The human gene mutation database (HGMD, 2019) has a record of five pathogenic variants in SLC24A4, causing AI phenotype. CONCLUSION: This nonsense sequence variant c.1192C > T (p.Gln398*) is the sixth disease-causing variant in SLC24A4, which extends its mutation spectrum and confirms the role of this gene in the morphogenesis of human tooth enamel. The identified variant highlights the critical role of SLC24A4 in causing a rare AI type in humans.


Assuntos
Amelogênese Imperfeita/genética , Antiporters/genética , Cárie Dentária/genética , Predisposição Genética para Doença , Adulto , Amelogênese Imperfeita/epidemiologia , Amelogênese Imperfeita/patologia , Códon sem Sentido/genética , Cárie Dentária/epidemiologia , Cárie Dentária/patologia , Esmalte Dentário/metabolismo , Éxons/genética , Feminino , Humanos , Masculino , Morfogênese/genética , Paquistão/epidemiologia , Linhagem , Perda de Dente/genética , Perda de Dente/fisiopatologia , Sequenciamento Completo do Exoma , Adulto Jovem
17.
Neurology ; 94(23): e2441-e2447, 2020 06 09.
Artigo em Inglês | MEDLINE | ID: mdl-32467133

RESUMO

OBJECTIVE: Facioscapulohumeral muscular dystrophy (FSHD) is a heterogenetic disorder predominantly characterized by progressive facial and scapular muscle weakness. Patients with FSHD either have a contraction of the D4Z4 repeat on chromosome 4q35 or mutations in D4Z4 chromatin modifiers SMCHD1 and DNMT3B, both causing D4Z4 chromatin relaxation and inappropriate expression of the D4Z4-encoded DUX4 gene in skeletal muscle. In this study, we tested the hypothesis whether LRIF1, a known SMCHD1 protein interactor, is a disease gene for idiopathic FSHD2. METHODS: Clinical examination of a patient with idiopathic FSHD2 was combined with pathologic muscle biopsy examination and with genetic, epigenetic, and molecular studies. RESULTS: A homozygous LRIF1 mutation was identified in a patient with a clinical phenotype consistent with FSHD. This mutation resulted in the absence of the long isoform of LRIF1 protein, D4Z4 chromatin relaxation, and DUX4 and DUX4 target gene expression in myonuclei, all molecular and epigenetic hallmarks of FSHD. In concordance, LRIF1 was shown to bind to the D4Z4 repeat, and knockdown of the LRIF1 long isoform in muscle cells results in DUX4 and DUX4 target gene expression. CONCLUSION: LRIF1 is a bona fide disease gene for FSHD2. This study further reinforces the unifying genetic mechanism, which postulates that FSHD is caused by D4Z4 chromatin relaxation, resulting in inappropriate DUX4 expression in skeletal muscle.


Assuntos
Proteínas de Ciclo Celular/genética , Distrofia Muscular Facioescapuloumeral/genética , Biópsia , Proteínas de Ciclo Celular/química , Proteínas de Ciclo Celular/metabolismo , Células Cultivadas , Cromatina/ultraestrutura , Proteínas Cromossômicas não Histona/metabolismo , Cromossomos Humanos Par 4/genética , Códon sem Sentido , Consanguinidade , Fibroblastos , Mutação da Fase de Leitura , Duplicação Gênica , Regulação da Expressão Gênica , Proteínas de Homeodomínio/biossíntese , Proteínas de Homeodomínio/genética , Homozigoto , Humanos , Masculino , Pessoa de Meia-Idade , Fibras Musculares Esqueléticas/metabolismo , Músculo Esquelético/patologia , Linhagem , Isoformas de Proteínas/genética , Sequências Repetitivas de Ácido Nucleico
18.
RNA ; 26(9): 1247-1256, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32434780

RESUMO

We have previously shown that when the uridine of a stop codon (UAA, UAG, or UGA) is pseudouridylated, the ribosome reads through the modified stop codon. However, it is not clear as to whether or not the pseudouridine (Ψ)-mediated readthrough is dependent on the sequence context of mRNA. Here, we use several different approaches and the yeast system to address this question. We show that when a stop codon (premature termination codon, PTC) is introduced into the coding region of a reporter mRNA at several different positions (with different sequence contexts) and pseudouridylated, we detect similar levels of readthrough. Using mutational and selection/screen analyses, we also show that the upstream sequence (relative to PTC) as well as the nucleotides surrounding the PTC (upstream and downstream) play a minimal role (if at all) in Ψ-mediated ribosome readthrough. Interestingly, we detect no suppression of NMD (nonsense-mediated mRNA decay) by targeted PTC pseudouridylation in the yeast system. Our results indicate that Ψ-mediated nonsense suppression occurs at the translational level, and that the suppression is sequence context-independent, unlike some previously characterized rare stop codon readthrough events.


Assuntos
Códon sem Sentido/genética , Códon de Terminação/genética , Pseudouridina/genética , Saccharomyces cerevisiae/genética , Mutação/genética , Degradação do RNAm Mediada por Códon sem Sentido/genética , Nucleotídeos/genética , Biossíntese de Proteínas/genética , RNA Mensageiro/genética , Ribossomos/genética
19.
BMC Med Genet ; 21(1): 72, 2020 04 06.
Artigo em Inglês | MEDLINE | ID: mdl-32252659

RESUMO

BACKGROUND: Propionic acidemia (PA) is an autosomal recessive metabolic disorder caused by the deficiency of the mitochondrial protein propionyl-CoA carboxylase (PCC) and is associated with pathogenic variants in either of the two genes PCCA or PCCB. The present study aimed to identify the genetic cause of three Chinese patients with PA. CASE PRESENTATION: Three Chinese PA patients were diagnosed by using gas chromatography-mass spectrometry(GC-MS), tandem mass spectrometry (MS/MS) and molecular diagnostic methods. All patients had onset in the neonatal period. One patient died of infection and metabolic decompensation, and the other two had mild to moderate developmental delay/mental retardation. Mutation analysis of the PCCA gene identified that patient 1 carried the compound heterozygous c.1288C > T(p.R430X) and c.2002G > A(p.G668R), and patient 2 was homozygous for the c.1426C > T(p.R476X) mutation. Mutation analysis of the PCCB gene identified that patient 3 harbored the compound heterozygous mutations c.359_360del AT(p.Y120Cfs*40) and c.1398 + 1G > A. Among these mutations, three (c.1288C > T, c.359_360del AT and c.1398 + 1G > A) are novel. CONCLUSIONS: We reported three Chinese PA patients who had PCCA or PCCB mutants. Among them, in the PCCA gene, c.1288C > T(p.R430X) was a nonsense mutation, resulting in a truncated protein. c.359_360del AT was a frameshift mutation, leading to a p.Y120Cfs*40 change in the amino acid sequence in the PCCB protein. c.1398 + 1G > A was a splicing mutation, causing skipping of the exons 13-14. In conclusion, the novel mutations uncovered in this study will expands the mutation spectrum of PA.


Assuntos
Carbono-Carbono Ligases/genética , Metilmalonil-CoA Descarboxilase/genética , Mutação , Acidemia Propiônica/genética , Pré-Escolar , China , Códon sem Sentido , Análise Mutacional de DNA/métodos , Feminino , Mutação da Fase de Leitura , Cromatografia Gasosa-Espectrometria de Massas , Humanos , Lactente , Recém-Nascido , Masculino , Polimorfismo Genético , Deleção de Sequência
20.
PLoS One ; 15(4): e0232216, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32348326

RESUMO

BACKGROUND: The knowledge of hereditary predisposition has changed our understanding of Pulmonary Arterial Hypertension. Genetic testing has been widely extended and the application of Pulmonary Arterial Hypertension specific gene panels has allowed its inclusion in the diagnostic workup and increase the diagnostic ratio compared to the traditional sequencing techniques. This is particularly important in the differential diagnosis between Pulmonary Arterial Hypertension and Pulmonary Venoocclusive Disease. METHODS: Since November 2011, genetic testing is offered to all patients with idiopathic, hereditable and associated forms of Pulmonary Arterial Hypertension or Pulmonary Venoocclusive Disease included in the Spanish Registry of Pulmonary Arterial Hypertension. Herein, we present the clinical phenotype and prognosis of all Pulmonary Arterial Hypertension patients with disease-associated variants in TBX4. RESULTS: Out of 579 adults and 45 children, we found in eight patients from seven families, disease-causing associated variants in TBX4. All adult patients had a moderate-severe reduction in diffusion capacity. However, we observed a wide spectrum of clinical presentations, including Pulmonary Venoocclusive Disease suspicion, interstitial lung disease, pulmonary vascular abnormalities and congenital heart disease. CONCLUSIONS: Genetic testing is now essential for a correct diagnosis work-up in Pulmonary Arterial Hypertension. TBX4-associated Pulmonary Arterial Hypertension has marked clinical heterogeneity. In this regard, a genetic study is extremely useful to obtain an accurate diagnosis and provide appropriate management.


Assuntos
Hipertensão Pulmonar Primária Familiar/genética , Variação Genética , Proteínas com Domínio T/genética , Adolescente , Adulto , Criança , Pré-Escolar , Códon sem Sentido , Diagnóstico Diferencial , Hipertensão Pulmonar Primária Familiar/diagnóstico , Hipertensão Pulmonar Primária Familiar/diagnóstico por imagem , Feminino , Deleção de Genes , Predisposição Genética para Doença , Humanos , Masculino , Pessoa de Meia-Idade , Mutação de Sentido Incorreto , Linhagem , Fenótipo , Polimorfismo de Nucleotídeo Único , Prognóstico , Pneumopatia Veno-Oclusiva/diagnóstico , Pneumopatia Veno-Oclusiva/genética
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