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1.
Proc Natl Acad Sci U S A ; 118(4)2021 01 26.
Artigo em Inglês | MEDLINE | ID: mdl-33468668

RESUMO

Epistasis refers to the dependence of a mutation on other mutation(s) and the genetic context in general. In the context of human disorders, epistasis complicates the spectrum of disease symptoms and has been proposed as a major contributor to variations in disease outcome. The nonadditive relationship between mutations and the lack of complete understanding of the underlying physiological effects limit our ability to predict phenotypic outcome. Here, we report positive epistasis between intragenic mutations in the cystic fibrosis transmembrane conductance regulator (CFTR)-the gene responsible for cystic fibrosis (CF) pathology. We identified a synonymous single-nucleotide polymorphism (sSNP) that is invariant for the CFTR amino acid sequence but inverts translation speed at the affected codon. This sSNP in cis exhibits positive epistatic effects on some CF disease-causing missense mutations. Individually, both mutations alter CFTR structure and function, yet when combined, they lead to enhanced protein expression and activity. The most robust effect was observed when the sSNP was present in combination with missense mutations that, along with the primary amino acid change, also alter the speed of translation at the affected codon. Functional studies revealed that synergistic alteration in ribosomal velocity is the underlying mechanism; alteration of translation speed likely increases the time window for establishing crucial domain-domain interactions that are otherwise perturbed by each individual mutation.


Assuntos
Regulador de Condutância Transmembrana em Fibrose Cística/genética , Fibrose Cística/genética , Epistasia Genética , Biossíntese de Proteínas , Sequência de Aminoácidos/genética , Códon/genética , Fibrose Cística/patologia , Humanos , Mutação de Sentido Incorreto/genética , Polimorfismo de Nucleotídeo Único/genética , RNA Mensageiro/genética
2.
PLoS Genet ; 15(2): e1008007, 2019 02.
Artigo em Inglês | MEDLINE | ID: mdl-30807572

RESUMO

Cystic Fibrosis (CF) exhibits morbidity in several organs, including progressive lung disease in all patients and intestinal obstruction at birth (meconium ileus) in ~15%. Individuals with the same causal CFTR mutations show variable disease presentation which is partly attributed to modifier genes. With >6,500 participants from the International CF Gene Modifier Consortium, genome-wide association investigation identified a new modifier locus for meconium ileus encompassing ATP12A on chromosome 13 (min p = 3.83x10(-10)); replicated loci encompassing SLC6A14 on chromosome X and SLC26A9 on chromosome 1, (min p<2.2x10(-16), 2.81x10(-11), respectively); and replicated a suggestive locus on chromosome 7 near PRSS1 (min p = 2.55x10(-7)). PRSS1 is exclusively expressed in the exocrine pancreas and was previously associated with non-CF pancreatitis with functional characterization demonstrating impact on PRSS1 gene expression. We thus asked whether the other meconium ileus modifier loci impact gene expression and in which organ. We developed and applied a colocalization framework called the Simple Sum (SS) that integrates regulatory and genetic association information, and also contrasts colocalization evidence across tissues or genes. The associated modifier loci colocalized with expression quantitative trait loci (eQTLs) for ATP12A (p = 3.35x10(-8)), SLC6A14 (p = 1.12x10(-10)) and SLC26A9 (p = 4.48x10(-5)) in the pancreas, even though meconium ileus manifests in the intestine. The meconium ileus susceptibility locus on chromosome X appeared shifted in location from a previously identified locus for CF lung disease severity. Using the SS we integrated the lung disease association locus with eQTLs from nasal epithelia of 63 CF participants and demonstrated evidence of colocalization with airway-specific regulation of SLC6A14 (p = 2.3x10(-4)). Cystic Fibrosis is realizing the promise of personalized medicine, and identification of the contributing organ and understanding of tissue specificity for a gene modifier is essential for the next phase of personalizing therapeutic strategies.


Assuntos
Sistemas de Transporte de Aminoácidos Neutros/genética , Antiporters/genética , Fibrose Cística/genética , Perfilação da Expressão Gênica/métodos , Estudo de Associação Genômica Ampla/métodos , ATPase Trocadora de Hidrogênio-Potássio/genética , Transportadores de Sulfato/genética , Tripsina/genética , Sistemas de Transporte de Aminoácidos , Sistemas de Transporte de Aminoácidos Neutros/metabolismo , Antiporters/metabolismo , Fibrose Cística/metabolismo , Feminino , Regulação da Expressão Gênica , ATPase Trocadora de Hidrogênio-Potássio/metabolismo , Humanos , Pulmão/metabolismo , Masculino , Especificidade de Órgãos , Pâncreas Exócrino/metabolismo , Transportadores de Sulfato/metabolismo , Tripsina/metabolismo
3.
PLoS Comput Biol ; 16(10): e1008336, 2020 10.
Artigo em Inglês | MEDLINE | ID: mdl-33090994

RESUMO

Genome-wide association studies (GWAS) have primarily identified trait-associated loci in the non-coding genome. Colocalization analyses of SNP associations from GWAS with expression quantitative trait loci (eQTL) evidence enable the generation of hypotheses about responsible mechanism, genes and tissues of origin to guide functional characterization. Here, we present a web-based colocalization browsing and testing tool named LocusFocus (https://locusfocus.research.sickkids.ca). LocusFocus formally tests colocalization using our established Simple Sum method to identify the most relevant genes and tissues for a particular GWAS locus in the presence of high linkage disequilibrium and/or allelic heterogeneity. We demonstrate the utility of LocusFocus, following up on a genome-wide significant locus from a GWAS of meconium ileus (an intestinal obstruction in cystic fibrosis). Using LocusFocus for colocalization analysis with eQTL data suggests variation in ATP12A gene expression in the pancreas rather than intestine is responsible for the GWAS locus. LocusFocus has no operating system dependencies and may be installed in a local web server. LocusFocus is available under the MIT license, with full documentation and source code accessible on GitHub at https://github.com/naim-panjwani/LocusFocus.


Assuntos
Biologia Computacional/métodos , Estudo de Associação Genômica Ampla/métodos , Anotação de Sequência Molecular/métodos , Fibrose Cística/genética , Predisposição Genética para Doença/genética , Humanos , Internet , Desequilíbrio de Ligação/genética , Polimorfismo de Nucleotídeo Único/genética , Locos de Características Quantitativas/genética , Software
4.
Am J Respir Crit Care Med ; 199(9): 1116-1126, 2019 05 01.
Artigo em Inglês | MEDLINE | ID: mdl-30888834

RESUMO

Rationale: The advent of precision treatment for cystic fibrosis using small-molecule therapeutics has created a need to estimate potential clinical improvements attributable to increases in cystic fibrosis transmembrane conductance regulator (CFTR) function. Objectives: To derive CFTR function of a variety of CFTR genotypes and correlate with key clinical features (sweat chloride concentration, pancreatic exocrine status, and lung function) to develop benchmarks for assessing response to CFTR modulators. Methods: CFTR function assigned to 226 unique CFTR genotypes was correlated with the clinical data of 54,671 individuals enrolled in the Clinical and Functional Translation of CFTR (CFTR2) project. Cross-sectional FEV1% predicted measurements were plotted by age at which measurement was obtained. Shifts in sweat chloride concentration and lung function reported in CFTR modulator trials were compared with function-phenotype correlations to assess potential efficacy of therapies. Measurements and Main Results: CFTR genotype function exhibited a logarithmic relationship with each clinical feature. Modest increases in CFTR function related to differing genotypes were associated with clinically relevant improvements in cross-sectional FEV1% predicted over a range of ages (6-82 yr). Therapeutic responses to modulators corresponded closely to predictions from the CFTR2-derived relationship between CFTR genotype function and phenotype. Conclusions: Increasing CFTR function in individuals with severe disease will have a proportionally greater effect on outcomes than similar increases in CFTR function in individuals with mild disease and should reverse a substantial fraction of the disease process. This study provides reference standards for clinical outcomes that may be achieved by increasing CFTR function.


Assuntos
Regulador de Condutância Transmembrana em Fibrose Cística/genética , Fibrose Cística/genética , Adolescente , Adulto , Criança , Fibrose Cística/fisiopatologia , Regulador de Condutância Transmembrana em Fibrose Cística/fisiologia , Feminino , Volume Expiratório Forçado , Estudos de Associação Genética , Humanos , Masculino , Pessoa de Meia-Idade , Medicina de Precisão/métodos , Adulto Jovem
5.
Hum Mol Genet ; 25(20): 4590-4600, 2016 10 15.
Artigo em Inglês | MEDLINE | ID: mdl-28171547

RESUMO

Cystic fibrosis is realizing the promise of personalized medicine. Recent advances in drug development that target the causal CFTR directly result in lung function improvement, but variability in response is demanding better prediction of outcomes to improve management decisions. The genetic modifier SLC26A9 contributes to disease severity in the CF pancreas and intestine at birth and here we assess its relationship with disease severity and therapeutic response in the airways. SLC26A9 association with lung disease was assessed in individuals from the Canadian and French CF Gene Modifier consortia with CFTR-gating mutations and in those homozygous for the common Phe508del mutation. Variability in response to a CFTR-directed therapy attributed to SLC26A9 genotype was assessed in Canadian patients with gating mutations. A primary airway model system determined if SLC26A9 shows modification of Phe508del CFTR function upon treatment with a CFTR corrector. In those with gating mutations that retain cell surface-localized CFTR we show that SLC26A9 modifies lung function while this is not the case in individuals homozygous for Phe508del where cell surface expression is lacking. Treatment response to ivacaftor, which aims to improve CFTR-channel opening probability in patients with gating mutations, shows substantial variability in response, 28% of which can be explained by rs7512462 in SLC26A9 (P = 0.0006). When homozygous Phe508del primary bronchial cells are treated to restore surface CFTR, SLC26A9 likewise modifies treatment response (P = 0.02). Our findings indicate that SLC26A9 airway modification requires CFTR at the cell surface, and that a common variant in SLC26A9 may predict response to CFTR-directed therapeutics.


Assuntos
Aminofenóis/metabolismo , Antiporters/genética , Fibrose Cística/metabolismo , Genes Modificadores , Pulmão/metabolismo , Variantes Farmacogenômicos , Quinolonas/metabolismo , Aminofenóis/farmacocinética , Aminofenóis/farmacologia , Aminofenóis/uso terapêutico , Antiporters/metabolismo , Canadá , Células Cultivadas , Agonistas dos Canais de Cloreto/metabolismo , Agonistas dos Canais de Cloreto/farmacocinética , Agonistas dos Canais de Cloreto/farmacologia , Agonistas dos Canais de Cloreto/uso terapêutico , Fibrose Cística/tratamento farmacológico , Fibrose Cística/genética , Fibrose Cística/patologia , Regulador de Condutância Transmembrana em Fibrose Cística/agonistas , Feminino , França , Estudos de Associação Genética , Humanos , Pulmão/efeitos dos fármacos , Pulmão/patologia , Masculino , Modelos Genéticos , Gravidade do Paciente , Polimorfismo de Nucleotídeo Único , Medicina de Precisão , Quinolonas/farmacocinética , Quinolonas/farmacologia , Quinolonas/uso terapêutico , Transportadores de Sulfato
6.
Am J Hum Genet ; 97(1): 125-38, 2015 Jul 02.
Artigo em Inglês | MEDLINE | ID: mdl-26140448

RESUMO

Gene-based, pathway, and other multivariate association methods are motivated by the possibility of GxG and GxE interactions; however, accounting for such interactions is limited by the challenges associated with adequate modeling information. Here we propose an easy-to-implement joint location-scale (JLS) association testing framework for single-variant and multivariate analysis that accounts for interactions without explicitly modeling them. We apply the JLS method to a gene-set analysis of cystic fibrosis (CF) lung disease, which is influenced by multiple environmental and genetic factors. We identify and replicate an association between the constituents of the apical plasma membrane and CF lung disease (p = 0.0099 and p = 0.0180, respectively) and highlight a role for the SLC9A3-SLC9A3R1/2-EZR complex in contributing to CF lung disease. Many association studies could benefit from re-analysis with the JLS method that leverages complex genetic architecture for SNP, gene, and pathway identification. Analytical verification, simulation, and additional proof-of-principle applications support our approach.


Assuntos
Membrana Celular/metabolismo , Fibrose Cística/genética , Complicações do Diabetes/genética , Estudos de Associação Genética/métodos , Complexos Multiproteicos/genética , Polimorfismo de Nucleotídeo Único/genética , Simulação por Computador , Fibrose Cística/metabolismo , Complicações do Diabetes/metabolismo , Humanos , Fosfoproteínas/genética , Trocador 3 de Sódio-Hidrogênio , Trocadores de Sódio-Hidrogênio/genética
7.
Am J Hum Genet ; 96(2): 318-28, 2015 Feb 05.
Artigo em Inglês | MEDLINE | ID: mdl-25640674

RESUMO

Variation in cystic fibrosis (CF) phenotypes, including lung disease severity, age of onset of persistent Pseudomonas aeruginosa (P. aeruginosa) lung infection, and presence of meconium ileus (MI), has been partially explained by genome-wide association studies (GWASs). It is not expected that GWASs alone are sufficiently powered to uncover all heritable traits associated with CF phenotypic diversity. Therefore, we utilized gene expression association from lymphoblastoid cells lines from 754 p.Phe508del CF-affected homozygous individuals to identify genes and pathways. LPAR6, a G protein coupled receptor, associated with lung disease severity (false discovery rate q value = 0.0006). Additional pathway analyses, utilizing a stringent permutation-based approach, identified unique signals for all three phenotypes. Pathways associated with lung disease severity were annotated in three broad categories: (1) endomembrane function, containing p.Phe508del processing genes, providing evidence of the importance of p.Phe508del processing to explain lung phenotype variation; (2) HLA class I genes, extending previous GWAS findings in the HLA region; and (3) endoplasmic reticulum stress response genes. Expression pathways associated with lung disease were concordant for some endosome and HLA pathways, with pathways identified using GWAS associations from 1,978 CF-affected individuals. Pathways associated with age of onset of persistent P. aeruginosa infection were enriched for HLA class II genes, and those associated with MI were related to oxidative phosphorylation. Formal testing demonstrated that genes showing differential expression associated with lung disease severity were enriched for heritable genetic variation and expression quantitative traits. Gene expression provided a powerful tool to identify unrecognized heritable variation, complementing ongoing GWASs in this rare disease.


Assuntos
Fibrose Cística/genética , Fibrose Cística/patologia , Genes MHC Classe I/genética , Variação Genética , Fenótipo , Receptores de Ácidos Lisofosfatídicos/genética , Estresse do Retículo Endoplasmático/genética , Perfilação da Expressão Gênica , Humanos , Modelos Lineares , Deleção de Sequência/genética
8.
PLoS Genet ; 11(6): e1005288, 2015 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-26057580

RESUMO

Genetic models of ribosome dysfunction show selective organ failure, highlighting a gap in our understanding of cell-type specific responses to translation insufficiency. Translation defects underlie a growing list of inherited and acquired cancer-predisposition syndromes referred to as ribosomopathies. We sought to identify molecular mechanisms underlying organ failure in a recessive ribosomopathy, with particular emphasis on the pancreas, an organ with a high and reiterative requirement for protein synthesis. Biallelic loss of function mutations in SBDS are associated with the ribosomopathy Shwachman-Diamond syndrome, which is typified by pancreatic dysfunction, bone marrow failure, skeletal abnormalities and neurological phenotypes. Targeted disruption of Sbds in the murine pancreas resulted in p53 stabilization early in the postnatal period, specifically in acinar cells. Decreased Myc expression was observed and atrophy of the adult SDS pancreas could be explained by the senescence of acinar cells, characterized by induction of Tgfß, p15(Ink4b) and components of the senescence-associated secretory program. This is the first report of senescence, a tumour suppression mechanism, in association with SDS or in response to a ribosomopathy. Genetic ablation of p53 largely resolved digestive enzyme synthesis and acinar compartment hypoplasia, but resulted in decreased cell size, a hallmark of decreased translation capacity. Moreover, p53 ablation resulted in expression of acinar dedifferentiation markers and extensive apoptosis. Our findings indicate a protective role for p53 and senescence in response to Sbds ablation in the pancreas. In contrast to the pancreas, the Tgfß molecular signature was not detected in fetal bone marrow, liver or brain of mouse models with constitutive Sbds ablation. Nevertheless, as observed with the adult pancreas phenotype, disease phenotypes of embryonic tissues, including marked neuronal cell death due to apoptosis, were determined to be p53-dependent. Our findings therefore point to cell/tissue-specific responses to p53-activation that include distinction between apoptosis and senescence pathways, in the context of translation disruption.


Assuntos
Doenças da Medula Óssea/genética , Senescência Celular , Insuficiência Pancreática Exócrina/genética , Lipomatose/genética , Pâncreas/metabolismo , Proteínas/metabolismo , Ribossomos/metabolismo , Células Acinares/metabolismo , Células Acinares/patologia , Células Acinares/fisiologia , Animais , Apoptose , Doenças da Medula Óssea/metabolismo , Doenças da Medula Óssea/patologia , Células Cultivadas , Insuficiência Pancreática Exócrina/metabolismo , Insuficiência Pancreática Exócrina/patologia , Lipomatose/metabolismo , Lipomatose/patologia , Camundongos , Camundongos Endogâmicos C57BL , Pâncreas/crescimento & desenvolvimento , Pâncreas/patologia , Biossíntese de Proteínas , Proteínas/genética , Ribossomos/genética , Síndrome de Shwachman-Diamond , Fator de Crescimento Transformador beta/metabolismo , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/metabolismo
9.
Am J Respir Crit Care Med ; 194(11): 1375-1382, 2016 12 01.
Artigo em Inglês | MEDLINE | ID: mdl-27258095

RESUMO

RATIONALE: Expanding the use of cystic fibrosis transmembrane conductance regulator (CFTR) potentiators and correctors for the treatment of cystic fibrosis (CF) requires precise and accurate biomarkers. Sweat chloride concentration provides an in vivo assessment of CFTR function, but it is unknown the degree to which CFTR mutations account for sweat chloride variation. OBJECTIVES: To estimate potential sources of variation for sweat chloride measurements, including demographic factors, testing variability, recording biases, and CFTR genotype itself. METHODS: A total of 2,639 sweat chloride measurements were obtained in 1,761 twins/siblings from the CF Twin-Sibling Study, French CF Modifier Gene Study, and Canadian Consortium for Genetic Studies. Variance component estimation was performed by nested mixed modeling. MEASUREMENTS AND MAIN RESULTS: Across the tested CF population as a whole, CFTR gene mutations were found to be the primary determinant of sweat chloride variability (56.1% of variation) with contributions from variation over time (e.g., factors related to testing on different days; 13.8%), environmental factors (e.g., climate, family diet; 13.5%), other residual factors (e.g., test variability; 9.9%), and unique individual factors (e.g., modifier genes, unique exposures; 6.8%) (likelihood ratio test, P < 0.001). Twin analysis suggested that modifier genes did not play a significant role because the heritability estimate was negligible (H2 = 0; 95% confidence interval, 0.0-0.35). For an individual with CF, variation in sweat chloride was primarily caused by variation over time (58.1%) with the remainder attributable to residual/random factors (41.9%). CONCLUSIONS: Variation in the CFTR gene is the predominant cause of sweat chloride variation; most of the non-CFTR variation is caused by testing variability and unique environmental factors. If test precision and accuracy can be improved, sweat chloride measurement could be a valuable biomarker for assessing response to therapies directed at mutant CFTR.


Assuntos
Cloretos/metabolismo , Fibrose Cística/metabolismo , Suor/metabolismo , Adulto , Biomarcadores/metabolismo , Regulador de Condutância Transmembrana em Fibrose Cística/metabolismo , Feminino , Humanos , Masculino , Adulto Jovem
10.
Genet Med ; 18(4): 333-40, 2016 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-26087176

RESUMO

RATIONALE: Meconium ileus (MI) is a perinatal complication in cystic fibrosis (CF), which is only minimally influenced by environmental factors. We derived and examined MI prevalence (MIP) scores to assess CFTR phenotype-phenotype correlation for severe mutations. METHOD: MIP scores were established using a Canadian CF population (n = 2,492) as estimates of the proportion of patients with MI among all patients carrying the same CFTR mutation, focusing on patients with p.F508del as the second allele. Comparisons were made to the registries from the US CF Foundation (n = 43,432), Italy (Veneto/Trentino/Alto Adige regions) (n = 1,788), and Germany (n = 3,596). RESULTS: The prevalence of MI varied among the different registries (13-21%). MI was predominantly prevalent in patients with pancreatic insufficiency carrying "severe" CFTR mutations. In this severe spectrum MIP scores further distinguished between mutation types, for example, G542X (0.31) with a high, F508del (0.22) with a moderate, and G551D (0.08) with a low MIP score. Higher MIP scores were associated with more severe clinical phenotypes, such as a lower forced expiratory volume in 1 second (P = 0.01) and body mass index z score (P = 0.04). CONCLUSIONS: MIP scores can be used to rank CFTR mutations according to their clinical severity and provide a means to expand delineation of CF phenotypes.Genet Med 18 4, 333-340.


Assuntos
Regulador de Condutância Transmembrana em Fibrose Cística/genética , Fibrose Cística/complicações , Fibrose Cística/genética , Íleus/epidemiologia , Íleus/etiologia , Mecônio , Mutação , Adolescente , Adulto , Alelos , Canadá/epidemiologia , Criança , Fibrose Cística/diagnóstico , Fibrose Cística/epidemiologia , Feminino , Estudos de Associação Genética , Genótipo , Humanos , Íleus/diagnóstico , Masculino , Fenótipo , Prevalência , Sistema de Registros , Testes de Função Respiratória , Índice de Gravidade de Doença , Adulto Jovem
11.
Nature ; 464(7290): 852-7, 2010 Apr 08.
Artigo em Inglês | MEDLINE | ID: mdl-20305640

RESUMO

Mesenchymal cells contribute to the 'stroma' of most normal and malignant tissues, with specific mesenchymal cells participating in the regulatory niches of stem cells. By examining how mesenchymal osteolineage cells modulate haematopoiesis, here we show that deletion of Dicer1 specifically in mouse osteoprogenitors, but not in mature osteoblasts, disrupts the integrity of haematopoiesis. Myelodysplasia resulted and acute myelogenous leukaemia emerged that had acquired several genetic abnormalities while having intact Dicer1. Examining gene expression altered in osteoprogenitors as a result of Dicer1 deletion showed reduced expression of Sbds, the gene mutated in Schwachman-Bodian-Diamond syndrome-a human bone marrow failure and leukaemia pre-disposition condition. Deletion of Sbds in mouse osteoprogenitors induced bone marrow dysfunction with myelodysplasia. Therefore, perturbation of specific mesenchymal subsets of stromal cells can disorder differentiation, proliferation and apoptosis of heterologous cells, and disrupt tissue homeostasis. Furthermore, primary stromal dysfunction can result in secondary neoplastic disease, supporting the concept of niche-induced oncogenesis.


Assuntos
Osso e Ossos/patologia , Leucemia Mieloide Aguda/patologia , Síndromes Mielodisplásicas/patologia , Células-Tronco/patologia , Animais , Medula Óssea/metabolismo , Medula Óssea/patologia , Osso e Ossos/metabolismo , Diferenciação Celular , Linhagem da Célula , Feminino , Deleção de Genes , Hematopoese/genética , Leucemia Mieloide Aguda/genética , Leucemia Mieloide Aguda/metabolismo , Masculino , Mesoderma/citologia , Camundongos , Síndromes Mielodisplásicas/genética , Síndromes Mielodisplásicas/metabolismo , Osteoblastos/metabolismo , Osteoblastos/patologia , Fenótipo , Proteínas/genética , Proteínas/metabolismo , Ribonuclease III/deficiência , Ribonuclease III/genética , Ribonuclease III/metabolismo , Sarcoma Mieloide/genética , Sarcoma Mieloide/metabolismo , Sarcoma Mieloide/patologia , Nicho de Células-Tronco/metabolismo , Nicho de Células-Tronco/patologia , Células-Tronco/metabolismo , Células Estromais/metabolismo , Células Estromais/patologia
12.
J Pediatr ; 166(5): 1152-1157.e6, 2015 May.
Artigo em Inglês | MEDLINE | ID: mdl-25771386

RESUMO

OBJECTIVES: To test the hypothesis that multiple constituents of the apical plasma membrane residing alongside the causal cystic fibrosis (CF) transmembrane conductance regulator protein, including known CF modifiers SLC26A9, SLC6A14, and SLC9A3, would be associated with prenatal exocrine pancreatic damage as measured by newborn screened (NBS) immunoreactive trypsinogen (IRT) levels. STUDY DESIGN: NBS IRT measures and genome-wide genotype data were available on 111 subjects from Colorado, 37 subjects from Wisconsin, and 80 subjects from France. Multiple linear regression was used to determine whether any of 8 single nucleotide polymorphisms (SNPs) in SLC26A9, SLC6A14, and SLC9A3 were associated with IRT and whether other constituents of the apical plasma membrane contributed to IRT. RESULTS: In the Colorado sample, 3 SLC26A9 SNPs were associated with NBS IRT (min P=1.16×10(-3); rs7512462), but no SLC6A14 or SLC9A3 SNPs were associated (P>.05). The rs7512462 association replicated in the Wisconsin sample (P=.03) but not in the French sample (P=.76). Furthermore, rs7512462 was the top-ranked apical membrane constituent in the combined Colorado and Wisconsin sample. CONCLUSIONS: NBS IRT is a biomarker of prenatal exocrine pancreatic disease in patients with CF, and a SNP in SLC26A9 accounts for significant IRT variability. This work suggests SLC26A9 as a potential therapeutic target to ameliorate exocrine pancreatic disease.


Assuntos
Antiporters/genética , Fibrose Cística/genética , Pâncreas Exócrino/anormalidades , Biomarcadores/sangue , Membrana Celular/metabolismo , Colorado , Regulador de Condutância Transmembrana em Fibrose Cística/genética , Feminino , França , Predisposição Genética para Doença , Genótipo , Humanos , Recém-Nascido , Modelos Lineares , Masculino , Mutação , Triagem Neonatal , Polimorfismo de Nucleotídeo Único , Controle de Qualidade , Transportadores de Sulfato , Tripsinogênio/sangue , Wisconsin
13.
Haematologica ; 100(10): 1285-93, 2015 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-26185170

RESUMO

Shwachman-Diamond syndrome is a congenital bone marrow failure disorder characterized by debilitating neutropenia. The disease is associated with loss-of-function mutations in the SBDS gene, implicated in ribosome biogenesis, but the cellular and molecular events driving cell specific phenotypes in ribosomopathies remain poorly defined. Here, we established what is to our knowledge the first mammalian model of neutropenia in Shwachman-Diamond syndrome through targeted downregulation of Sbds in hematopoietic stem and progenitor cells expressing the myeloid transcription factor CCAAT/enhancer binding protein α (Cebpa). Sbds deficiency in the myeloid lineage specifically affected myelocytes and their downstream progeny while, unexpectedly, it was well tolerated by rapidly cycling hematopoietic progenitor cells. Molecular insights provided by massive parallel sequencing supported cellular observations of impaired cell cycle exit and formation of secondary granules associated with the defect of myeloid lineage progression in myelocytes. Mechanistically, Sbds deficiency activated the p53 tumor suppressor pathway and induced apoptosis in these cells. Collectively, the data reveal a previously unanticipated, selective dependency of myelocytes and downstream progeny, but not rapidly cycling progenitors, on this ubiquitous ribosome biogenesis protein, thus providing a cellular basis for the understanding of myeloid lineage biased defects in Shwachman-Diamond syndrome.


Assuntos
Diferenciação Celular/genética , Linhagem da Célula/genética , Células-Tronco Hematopoéticas/metabolismo , Células Mieloides/citologia , Células Mieloides/metabolismo , Neutropenia/genética , Proteínas/genética , Animais , Apoptose/genética , Doenças da Medula Óssea/genética , Proteínas Estimuladoras de Ligação a CCAAT/genética , Proteínas Estimuladoras de Ligação a CCAAT/metabolismo , Ciclo Celular/genética , Modelos Animais de Doenças , Insuficiência Pancreática Exócrina/genética , Deleção de Genes , Hematopoese/genética , Transplante de Células-Tronco Hematopoéticas , Células-Tronco Hematopoéticas/citologia , Lipomatose/genética , Camundongos , Camundongos Knockout , Síndrome de Shwachman-Diamond , Transdução de Sinais , Proteína Supressora de Tumor p53/metabolismo
14.
Nat Genet ; 38(1): 93-100, 2006 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-16311595

RESUMO

Methylmalonic aciduria and homocystinuria, cblC type (OMIM 277400), is the most common inborn error of vitamin B(12) (cobalamin) metabolism, with about 250 known cases. Affected individuals have developmental, hematological, neurological, metabolic, ophthalmologic and dermatologic clinical findings. Although considered a disease of infancy or childhood, some individuals develop symptoms in adulthood. The cblC locus was mapped to chromosome region 1p by linkage analysis. We refined the chromosomal interval using homozygosity mapping and haplotype analyses and identified the MMACHC gene. In 204 individuals, 42 different mutations were identified, many consistent with a loss of function of the protein product. One mutation, 271dupA, accounted for 40% of all disease alleles. Transduction of wild-type MMACHC into immortalized cblC fibroblast cell lines corrected the cellular phenotype. Molecular modeling predicts that the C-terminal region of the gene product folds similarly to TonB, a bacterial protein involved in energy transduction for cobalamin uptake.


Assuntos
Proteínas de Transporte/genética , Homocistinúria/genética , Erros Inatos do Metabolismo/genética , Ácido Metilmalônico/urina , Mutação , Sequência de Aminoácidos , Proteínas de Bactérias/química , Proteínas de Transporte/química , Proteínas de Transporte/metabolismo , Linhagem Celular , Mapeamento Cromossômico , Sequência Conservada , Fibroblastos/metabolismo , Haplótipos/genética , Humanos , Proteínas de Membrana/química , Dados de Sequência Molecular , Oxirredutases , Dobramento de Proteína , Homologia Estrutural de Proteína , Vitamina B 12/metabolismo
16.
Gastroenterology ; 143(2): 481-92, 2012 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-22510201

RESUMO

BACKGROUND & AIMS: Shwachman-Diamond syndrome (SDS) is the second leading cause of hereditary exocrine pancreatic dysfunction. More than 90% of patients with SDS have biallelic loss-of-function mutations in the Shwachman-Bodian Diamond syndrome (SBDS) gene, which encodes a factor involved in ribosome function. We investigated whether mutations in Sbds lead to similar pancreatic defects in mice. METHODS: Pancreas-specific knock-out mice were generated using a floxed Sbds allele and bred with mice carrying a null or disease-associated missense Sbds allele. Cre recombinase, regulated by the pancreatic transcription factor 1a promoter, was used to disrupt Sbds specifically in the pancreas. Models were assessed for pancreatic dysfunction and growth impairment. RESULTS: Disruption of Sbds in the mouse pancreas was sufficient to recapitulate SDS phenotypes. Pancreata of mice with Sbds mutations had decreased mass, fat infiltration, but general preservation of ductal and endocrine compartments. Pancreatic extracts from mutant mice had defects in formation of the 80S ribosomal complex. The exocrine compartment of mutant mice was hypoplastic and individual acini produced few zymogen granules. The null Sbds allele resulted in an earlier onset of phenotypes as well as endocrine impairment. Mutant mice had reduced serum levels of digestive enzymes and overall growth impairment. CONCLUSIONS: We developed a mouse model of SDS with pancreatic phenotypes similar to those of the human disease. This model could be used to investigate organ-specific consequences of Sbds-associated ribosomopathy. Sbds genotypes correlated with phenotypes. Defects developed specifically in the pancreata of mice, reducing growth of mice and production of digestive enzymes. SBDS therefore appears to be required for normal pancreatic development and function.


Assuntos
Doenças da Medula Óssea/genética , Modelos Animais de Doenças , Precursores Enzimáticos/metabolismo , Insuficiência Pancreática Exócrina/genética , Lipomatose/genética , Camundongos , Pâncreas Exócrino/metabolismo , Proteínas/genética , Vesículas Secretórias/metabolismo , Animais , Biomarcadores/metabolismo , Doenças da Medula Óssea/metabolismo , Doenças da Medula Óssea/patologia , Doenças da Medula Óssea/fisiopatologia , Insuficiência Pancreática Exócrina/metabolismo , Insuficiência Pancreática Exócrina/patologia , Insuficiência Pancreática Exócrina/fisiopatologia , Estudos de Associação Genética , Lipomatose/metabolismo , Lipomatose/patologia , Lipomatose/fisiopatologia , Camundongos Knockout , Pâncreas Exócrino/patologia , Pâncreas Exócrino/fisiopatologia , Proteínas/metabolismo , Reação em Cadeia da Polimerase em Tempo Real , Síndrome de Shwachman-Diamond
17.
Nat Genet ; 33(1): 97-101, 2003 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-12496757

RESUMO

Shwachman-Diamond syndrome (SDS; OMIM 260400) is an autosomal recessive disorder with clinical features that include pancreatic exocrine insufficiency, hematological dysfunction and skeletal abnormalities. Here, we report identification of disease-associated mutations in an uncharacterized gene, SBDS, in the interval of 1.9 cM at 7q11 previously shown to be associated with the disease. We report that SBDS has a 1.6-kb transcript and encodes a predicted protein of 250 amino acids. A pseudogene copy (SBDSP) with 97% nucleotide sequence identity resides in a locally duplicated genomic segment of 305 kb. We found recurring mutations resulting from gene conversion in 89% of unrelated individuals with SDS (141 of 158), with 60% (95 of 158) carrying two converted alleles. Converted segments consistently included at least one of two pseudogene-like sequence changes that result in protein truncation. SDBS is a member of a highly conserved protein family of unknown function with putative orthologs in diverse species including archaea and eukaryotes. Archaeal orthologs are located within highly conserved operons that include homologs of RNA-processing genes, suggesting that SDS may be caused by a deficiency in an aspect of RNA metabolism that is essential for development of the exocrine pancreas, hematopoiesis and chrondrogenesis.


Assuntos
Insuficiência Pancreática Exócrina/genética , Doenças Hematológicas/genética , Anormalidades Musculoesqueléticas/genética , Proteínas/genética , Alelos , Sequência de Bases , Cromossomos Humanos Par 7/genética , Sequência Conservada , Análise Mutacional de DNA , Feminino , Conversão Gênica , Perfilação da Expressão Gênica , Humanos , Escore Lod , Masculino , Dados de Sequência Molecular , Mutação , Pseudogenes/genética , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Síndrome
18.
Blood ; 115(6): 1264-6, 2010 Feb 11.
Artigo em Inglês | MEDLINE | ID: mdl-20007542

RESUMO

Quebec platelet disorder (QPD) is an autosomal dominant bleeding disorder linked to a region on chromosome 10 that includes PLAU, the urokinase plasminogen activator gene. QPD increases urokinase plasminogen activator mRNA levels, particularly during megakaryocyte differentiation, without altering expression of flanking genes. Because PLAU sequence changes were excluded as the cause of this bleeding disorder, we investigated whether the QPD mutation involved PLAU copy number variation. All 38 subjects with QPD had a direct tandem duplication of a 78-kb genomic segment that includes PLAU. This mutation was specific to QPD as it was not present in any unaffected family members (n = 114), unrelated French Canadians (n = 221), or other persons tested (n = 90). This new information on the genetic mutation will facilitate diagnostic testing for QPD and studies of its pathogenesis and prevalence. QPD is the first bleeding disorder to be associated with a gene duplication event and a PLAU mutation.


Assuntos
Transtornos Plaquetários/diagnóstico , Transtornos Plaquetários/genética , Dosagem de Genes , Duplicação Gênica , Ativador de Plasminogênio Tipo Uroquinase/genética , Adulto , Sequência de Bases , Transtornos Plaquetários/sangue , Cromossomos Humanos Par 10/genética , Feminino , Humanos , Hibridização in Situ Fluorescente , Cariotipagem , Masculino , Dados de Sequência Molecular , Prognóstico , Homologia de Sequência do Ácido Nucleico
19.
Pediatr Blood Cancer ; 59(5): 945-6, 2012 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-22213587

RESUMO

Shwachman Diamond syndrome (SDS) is a rare inherited bone marrow failure syndrome (IBMFS) characterized by neutropenia, exocrine pancreatic dysfunction, and cancer predisposition. Patients are at risk for myelodysplastic syndrome (MDS) and acute myelogenous leukemia (AML) but, unlike other IBMFS, there have been no reported cases of solid tumors. We report a novel case of a solid tumor in a patient with SDS and biallelic mutations in the Shwachman Bodian Diamond Syndrome gene (SBDS). Whether the development of breast cancer in this patient is due to SDS or an isolated case due to unknown factors requires further study.


Assuntos
Alelos , Doenças da Medula Óssea/genética , Neoplasias da Mama/genética , Insuficiência Pancreática Exócrina/genética , Lipomatose/genética , Mutação , Proteínas/genética , Adulto , Doenças da Medula Óssea/complicações , Insuficiência Pancreática Exócrina/complicações , Feminino , Humanos , Lipomatose/complicações , Síndrome de Shwachman-Diamond
20.
J Cyst Fibros ; 21(4): 616-622, 2022 07.
Artigo em Inglês | MEDLINE | ID: mdl-35190293

RESUMO

BACKGROUND: Variation in respiratory response to cystic fibrosis (CF) small molecule therapies is due in part to the contribution of CF lung disease modifier genes. Cultured human bronchial epithelia (HBE) is the gold standard respiratory model for assessing CF therapeutic efficacy but it is hard to access. Cultured human nasal epithelia (HNE) is proposed as a more accessible surrogate model but it is unknown whether the expression profile of the modifier genes are comparable between HNE and HBE which we assess here. METHODS: RNA-sequencing was conducted on paired cultured and fresh HNE and HBE (n = 71 samples) collected from 21 individuals with CF. Genome-wide gene expression was first compared between cultured and fresh cells and then between cultured HNE and HBE based on an equivalence testing procedure we implemented. The co-expression relationships of CFTR and CF lung disease modifier genes were compared between cultured HNE and HBE to determine equivalent interactions. RESULTS: The culturing process had little impact on the expression level of CF lung disease modifier genes. Over 90% of expressed genes showed significant equivalent expression level across cultured HNE and HBE (expression fold-change<2, FDR<0.1), including CFTR and CF lung disease modifier genes. The difference in co-expression relationships among these genes was not significant (p-value=0.99), suggesting their functional interactions are likely to be consistent in the two models. CONCLUSIONS: Cultured HNE recapitulates the expression profile of CF lung disease modifier genes in cultured HBE, suggesting the biological processes involving these genes are likely to be consistent across the two models.


Assuntos
Fibrose Cística , Células Cultivadas , Fibrose Cística/terapia , Regulador de Condutância Transmembrana em Fibrose Cística/metabolismo , Genes Modificadores , Humanos , Mucosa Nasal/metabolismo , Mucosa Respiratória/metabolismo
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