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1.
Zhonghua Yi Xue Yi Chuan Xue Za Zhi ; 37(3): 263-268, 2020 Mar 10.
Artículo en Chino | MEDLINE | ID: mdl-32128742

RESUMEN

Spinal muscular atrophy (SMA) is one of the most common fatal autosomal recessive genetic disorders among infants. It is caused by mutations of motor neuron survival gene 1 (SMN1). The incidence of SMA among newborns is approximately 1/10 000 - 1/6000, and the carrier rate is 1/72 - 1/47 with an ethnic variance. Based on the time of onset and clinical phenotype, SMA can be divided into types I - IV. Approximately 95% of SMA patients have carried homozygous deletions of exon 7 of the (SMN1)] gene. For its significant phenotypic difference, abundant changes of (SMN1)] gene copy number, presence of pseudogene interference and high carrier rate, early diagnosis, genetic consultation, treatment and prevention of SMA can be difficult. This guideline summarizes the relevant research, guideline and consensus issued at home and abroad, clinical manifestations and pathogenesis of SMA patients, and experience in its diagnosis and genetic counseling, with an aim to promote a standardized diagnosis and treatment and reduce the births of children affected with the disease.


Asunto(s)
Atrofia Muscular Espinal/diagnóstico , Atrofia Muscular Espinal/terapia , Guías de Práctica Clínica como Asunto , Exones , Dosificación de Gen , Asesoramiento Genético , Heterocigoto , Humanos , Recién Nacido , Fenotipo , Eliminación de Secuencia , Proteína 1 para la Supervivencia de la Neurona Motora/genética
2.
BMC Med Genet ; 21(1): 26, 2020 02 06.
Artículo en Inglés | MEDLINE | ID: mdl-32028920

RESUMEN

BACKGROUND: While Miller-Dieker syndrome critical region deletions are well known delineated anomalies, submicroscopic duplications in this region have recently emerged as a new distinctive syndrome. So far, only few cases have been described overlapping 17p13.3 duplications. METHODS: In this study, we report on clinical and cytogenetic characterization of two new cases involving 17p13.3 and 3p26 chromosomal regions in two sisters with familial history of lissencephaly. Fluorescent In Situ Hybridization and array Comparative Genomic Hybridization were performed. RESULTS: A deletion including the critical region of the Miller-Dieker syndrome of at least 2,9 Mb and a duplication of at least 3,6 Mb on the short arm of chromosome 3 were highlighted in one case. The opposite rearrangements, 17p13.3 duplication and 3p deletion, were observed in the second case. This double chromosomal aberration is the result of an adjacent 1:1 meiotic segregation of a maternal reciprocal translocation t(3,17)(p26.2;p13.3). CONCLUSIONS: 17p13.3 and 3p26 deletions have a clear range of phenotypic features while duplications still have an uncertain clinical significance. However, we could suggest that regardless of the type of the rearrangement, the gene dosage and interactions of CNTN4, CNTN6 and CHL1 in the 3p26 and PAFAH1B1, YWHAE in 17p13.3 could result in different clinical spectrums.


Asunto(s)
Lisencefalias Clásicas y Heterotopias Subcorticales en Banda/genética , Lisencefalia/genética , Neuronas/patología , Translocación Genética/genética , 1-Alquil-2-acetilglicerofosfocolina Esterasa/genética , Proteínas 14-3-3/genética , Moléculas de Adhesión Celular/genética , Movimiento Celular/genética , Preescolar , Deleción Cromosómica , Cromosomas Humanos Par 17/genética , Cromosomas Humanos Par 3/genética , Lisencefalias Clásicas y Heterotopias Subcorticales en Banda/diagnóstico , Lisencefalias Clásicas y Heterotopias Subcorticales en Banda/fisiopatología , Hibridación Genómica Comparativa , Contactinas/genética , Femenino , Dosificación de Gen/genética , Estudios de Asociación Genética , Humanos , Hibridación Fluorescente in Situ , Lisencefalia/diagnóstico , Lisencefalia/fisiopatología , Meiosis/genética , Proteínas Asociadas a Microtúbulos/genética , Neuronas/metabolismo , Fenotipo , Trisomía/genética
3.
Medicine (Baltimore) ; 99(3): e18809, 2020 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-32011487

RESUMEN

In this article, the correlation between the copy number of survival motor neuron 2 (SMN2) gene, neuronal apoptosis inhibitory protein (NAIP), and the phenotype of spinal muscular atrophy patients were analyzed.Forty patients with spinal muscular atrophy (SMA) were included in the study at the Department of Medical Genetics of the First People's Hospital and the Department of Neurology of the Second People's Hospital in Yunnan Province from January 2012 to September 2018. Multiplex ligation-dependent probe amplification assay was performed to determine the copy numbers of SMN2 and NAIP genes. Statistical analysis was performed to determine the correlation between copy numbers of the SMN2 and NAIP genes and the clinical phenotypes of SMA.Our results show that among the 40 SMA patients, there were 13 type I cases, 16 type II cases and 11 type III cases. A total of 37 patients possessed a homozygous deletion of SMN1 exons 7 and 8, while the other 3 SMA patients possessed a single copy of SMN1 exon 8. There was no correlation between SMA subtypes and the deletion types of SMN1 exon 7 and 8 (P = .611). The percentage of 2, 3, and 4 copies of SMN2 exon 7 was 25.0%, 62.5%, and 12.5%, respectively. The percentage of 0, 1, and 2 copies of NAIP exon 5 was 10%, 57.5%, and 32.5%, respectively. The distributions of SMN2 and NAIP copy numbers among various SMA types were significantly different (all P < .05). Five combined SMN1-SMN2-NAIP genotypes were detected, of which 0-3-1 genotype had the highest proportion than the others, accounting for 42.5%. The copy number of SMN2 and NAIP gene had synergistic effect on SMA phenotype. The combined SMN1-SMN2-NAIP genotypes with fewer copies were associated with earlier onset age, higher mortality, and smaller average age at death in SMA patients.Therefore, we conclude that the copy number variance of SMN2 and NAIP is correlated with the SMA phenotype. Analysis of the copy number structure of the SMN1-SMN2-NAIP gene is helpful for SMA typing, disease prognosis prediction, and genetic counseling.


Asunto(s)
Dosificación de Gen , Atrofia Muscular Espinal/genética , Proteína Inhibidora de la Apoptosis Neuronal/genética , Adolescente , Niño , Preescolar , Femenino , Estudios de Asociación Genética , Predisposición Genética a la Enfermedad , Humanos , Lactante , Masculino , Proteína 1 para la Supervivencia de la Neurona Motora/genética , Proteína 2 para la Supervivencia de la Neurona Motora/genética , Adulto Joven
4.
Microbes Environ ; 35(1)2020.
Artículo en Inglés | MEDLINE | ID: mdl-31969532

RESUMEN

The pmoA gene, encoding particulate methane monooxygenase in methanotrophs, and nirS and nirK genes, encoding bacterial nitrite reductases, were examined in the root and rhizosphere sediment of three common emergent macrophytes (Phragmites australis, Typha angustifolia, and Scirpus triqueter) and unvegetated sediment from eutrophic Wuliangsuhai Lake in China. Sequencing analyses indicated that 334 out of 351 cloned pmoA sequences were phylogenetically the most closely related to type I methanotrophs (Gammaproteobacteria), and Methylomonas denitrificans-like organisms accounted for 44.4% of the total community. In addition, 244 out of 250 cloned nirS gene sequences belonged to type I methanotrophs, and 31.2% of nirS genes were the most closely related to paddy rice soil clone SP-2-12 in Methylomonas of the total community. Three genera of type I methanotrophs, Methylomonas, Methylobacter, and Methylovulum, were common in both pmoA and nirS clone libraries in each sample. A quantitative PCR (qPCR) analysis demonstrated that the copy numbers of the nirS and nirK genes were significantly higher in rhizosphere sediments than in unvegetated sediments in P. australis and T. angustifolia plants. In the same sample, the nirS gene copy number was significantly higher than that of nirK. Furthermore, type I methanotrophs were localized in the root tissues according to catalyzed reporter deposition-fluorescence in situ hybridization (CARD-FISH). Thus, nirS-carrying type I methanotrophs were enriched in macrophyte root and rhizosphere sediment and are expected to play important roles in carbon/nitrogen cycles in a eutrophic wetland.


Asunto(s)
Eutrofización , Gammaproteobacteria/genética , Genes Bacterianos/genética , Magnoliopsida/microbiología , Microbiología del Suelo , Humedales , Proteínas Bacterianas/genética , China , Gammaproteobacteria/clasificación , Gammaproteobacteria/metabolismo , Dosificación de Gen , Lagos/microbiología , Metano/metabolismo , Nitrito Reductasas/genética , Oxigenasas/genética , Raíces de Plantas/microbiología , Rizosfera
5.
J Clin Pathol ; 73(2): 107-111, 2020 Feb.
Artículo en Inglés | MEDLINE | ID: mdl-31471467

RESUMEN

AIMS: KDM1A/LSD1 and ZNF217 are involved in a protein complex that participates in transcriptional regulation. ZNF217 has been analysed in numerous cancers and its amplification has been associated with advanced stages of disease; however, a similar role for KDM1A/LSD1 has not been uncovered. In this study, we estimated the number of KDM1A/LSD1 and ZNF217 gene copies in tissue samples from patients diagnosed with colorectal cancer (CRC), as well as its association with clinicopathological features in patients with CRC. METHODS: Paraffin-embedded tumour samples from 50 patients with CRC with a histopathological diagnosis of CRC were included. The number of copies of KDM1A/LSD1 and ZNF217 genes was determined by fluorescence in situ hybridisation (FISH). We also analysed the association between copy numbers of selected genes and clinicopathological data based on multivariate analysis. RESULTS: Deletion of the KDM1A/LSD1 gene occurred in 19 samples (38%), whereas ZNF217 gene amplification was identified in 11 samples (22%). We found a significant association between lymph node metastasis or advanced tumour stage and KDM1A/LSD1 gene deletion (p value=0.0003 and p value=0.011, respectively). CONCLUSIONS: KDM1A/LSD1 gene deletion could be considered a novel prognostic biomarker of late-stage CRC.


Asunto(s)
Biomarcadores de Tumor/genética , Neoplasias Colorrectales/genética , Eliminación de Gen , Histona Demetilasas/genética , Adulto , Anciano , Anciano de 80 o más Años , Neoplasias Colorrectales/enzimología , Neoplasias Colorrectales/patología , Estudios Transversales , Femenino , Dosificación de Gen , Humanos , Hibridación Fluorescente in Situ , Metástasis Linfática , Masculino , Persona de Mediana Edad , Estadificación de Neoplasias , Valor Predictivo de las Pruebas , Estudios Retrospectivos , Factores de Riesgo , Transactivadores/genética
6.
PLoS Negl Trop Dis ; 13(12): e0007852, 2019 12.
Artículo en Inglés | MEDLINE | ID: mdl-31830027

RESUMEN

BACKGROUND: The yellow fever mosquito Aedes aegypti is the major vector of dengue, yellow fever, Zika, and Chikungunya viruses. Worldwide vector control is largely based on insecticide treatments but, unfortunately, vector control programs are facing operational challenges due to mosquitoes becoming resistant to commonly used insecticides. In Southeast Asia, resistance of Ae. aegypti to chemical insecticides has been documented in several countries but no data regarding insecticide resistance has been reported in Laos. To fill this gap, we assessed the insecticide resistance of 11 Ae. aegypti populations to larvicides and adulticides used in public health operations in the country. We also investigated the underlying molecular mechanisms associated with resistance, including target site mutations and detoxification enzymes putatively involved in metabolic resistance. METHODS AND RESULTS: Bioassays on adults and larvae collected in five provinces revealed various levels of resistance to organophosphates (malathion and temephos), organochlorine (DDT) and pyrethroids (permethrin and deltamethrin). Synergist bioassays showed a significant increased susceptibility of mosquitoes to insecticides after exposure to detoxification enzyme inhibitors. Biochemical assays confirmed these results by showing significant elevated activities of cytochrome P450 monooxygenases (P450), glutathione S-transferases (GST) and carboxylesterases (CCE) in adults. Two kdr mutations, V1016G and F1534C, were detected by qPCR at low and high frequency, respectively, in all populations tested. A significant negative association between the two kdr mutations was detected. No significant association between kdr mutations frequency (for both 1534C and 1016G) and survival rate to DDT or permethrin (P > 0.05) was detected. Gene Copy Number Variations (CNV) were detected for particular detoxification enzymes. At the population level, the presence of CNV affecting the carboxylesterase CCEAE3A and the two cytochrome P450 CYP6BB2 and CYP6P12 were significantly correlated to insecticide resistance. CONCLUSIONS: These results suggest that both kdr mutations and metabolic resistance mechanisms are present in Laos but their impact on phenotypic resistance may differ in proportion at the population or individual level. Molecular analyses suggest that CNV affecting CCEAE3A previously associated with temephos resistance is also associated with malathion resistance while CNV affecting CYP6BB2 and CYP6P12 are associated with pyrethroid and possibly DDT resistance. The presence of high levels of insecticide resistance in the main arbovirus vector in Laos is worrying and may have important implications for dengue vector control in the country.


Asunto(s)
Aedes/efectos de los fármacos , Resistencia a los Insecticidas , Insecticidas/farmacología , Control de Mosquitos/métodos , Mosquitos Vectores/efectos de los fármacos , Aedes/genética , Animales , Bioensayo , Sinergismo Farmacológico , Femenino , Dosificación de Gen , Genes de Insecto , Hidrocarburos Clorados/farmacología , Laos , Larva/efectos de los fármacos , Redes y Vías Metabólicas/genética , Mosquitos Vectores/genética , Mutación , Organofosfatos/farmacología , Piretrinas/farmacología
7.
Nat Commun ; 10(1): 5725, 2019 12 16.
Artículo en Inglés | MEDLINE | ID: mdl-31844058

RESUMEN

Many traits vary among isogenic individuals in homogeneous environments. In microbes, plants and animals, variation in the protein chaperone system affects many such traits. In the animal model C. elegans, the expression level of hsp-16.2 chaperone biomarkers correlates with or predicts the penetrance of mutations and lifespan after heat shock. But the physiological mechanisms causing cells to express different amounts of the biomarker were unknown. Here, we used an in vivo microscopy approach to dissect different contributions to cell-to-cell variation in hsp-16.2 expression in the intestines of young adult animals, which generate the most lifespan predicting signal. While we detected both cell autonomous intrinsic noise and signaling noise, we found both contributions were relatively unimportant. The major contributor to cell-to-cell variation in biomarker expression was general differences in protein dosage. The hsp-16.2 biomarker reveals states of high or low effective dosage for many genes.


Asunto(s)
Proteínas de Caenorhabditis elegans/genética , Dosificación de Gen , Proteínas de Choque Térmico/genética , Longevidad/genética , Penetrancia , Animales , Animales Modificados Genéticamente , Biomarcadores/metabolismo , Caenorhabditis elegans/fisiología , Proteínas de Caenorhabditis elegans/metabolismo , Genes Reporteros/genética , Proteínas de Choque Térmico/metabolismo , Microscopía Intravital/métodos , Proteínas Luminiscentes/genética , Proteínas Luminiscentes/metabolismo , Microscopía Fluorescente/métodos , Modelos Animales , Imagen Molecular , Transducción de Señal/genética
8.
PLoS One ; 14(12): e0226620, 2019.
Artículo en Inglés | MEDLINE | ID: mdl-31877167

RESUMEN

BACKGROUND: One of the most important susceptibility loci for cancer is the 8q24 human chromosomal region. The non-protein coding gene locus plasmacytoma variant translocation 1 (PVT1) is located at 8q24 and is dysregulated in prostate cancer. PVT1 gives rise to multiple transcripts which may have different functions. Here, we describe a real-time quantitative polymerase chain reaction (qPCR)-based assay for copy number-based quantitation of PVT1 exons 4A, 4B, and 9 to enable accurate, reproducible, and quantifiable detection. METHODS: PVT1 exons 4A, 4B, and 9 were cloned into a plasmid vector to create standards for subsequent creation of linear standard curves representing a broad range of concentrations. PCR was carried out using SYBR-Green signal detection to quantify PVT1 exons 4A, 4B, and 9. The efficacy of this assay was evaluated by using it to detect these transcripts in prostate epithelial and prostate cancer cell lines, normal and cancerous human prostate tissues, human serum, mouse plasma, and urine samples. RESULTS: The results indicate that the assay can be used to quantify both low and high copy numbers of PVT1-derived transcripts. This is the first report of a copy number-based quantification assay for non-invasive detection of PVT1 derived transcripts. CONCLUSIONS: This novel assay holds promise for routine non-invasive testing in diseases where PVT1 is dysregulated.


Asunto(s)
Dosificación de Gen , Neoplasias de la Próstata/genética , ARN Largo no Codificante/genética , Línea Celular , Línea Celular Tumoral , Exones , Regulación Neoplásica de la Expresión Génica , Humanos , Masculino , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Translocación Genética
9.
Adv Exp Med Biol ; 1185: 477-481, 2019.
Artículo en Inglés | MEDLINE | ID: mdl-31884657

RESUMEN

Evidence suggests that metabolic dysregulation plays an important role in disease etiology of retinal degenerations. Several studies suggest that preserving the retinal metabolic ecosystem may be protective against retinal degenerations. We investigated whether activation of 5' adenosine monophosphate protein kinase (AMPK) is protective to the retina in several preclinical mouse models of retinal degeneration and found that metformin-induced activation of AMPK was able to delay or prevent retinal degeneration in the rd10 model of retinitis pigmentosa, the NaIO3 model of RPE and retinal injury, and the light damage model of retinal degeneration. This protection was associated with increased mitochondrial DNA copy number, increased levels of ATP, and a reduction in oxidative stress and oxidative DNA damage. We propose that AMPK plays an important role in regulation of the retinal metabolic ecosystem and that activation of AMPK may promote metabolic processes to prevent retinal degeneration.


Asunto(s)
Proteínas Quinasas Activadas por AMP/metabolismo , Retina/enzimología , Degeneración Retiniana/prevención & control , Animales , Daño del ADN , ADN Mitocondrial/genética , Modelos Animales de Enfermedad , Dosificación de Gen , Metformina/farmacología , Ratones , Estrés Oxidativo , Retinitis Pigmentosa/enzimología , Retinitis Pigmentosa/prevención & control
10.
PLoS Comput Biol ; 15(11): e1007460, 2019 11.
Artículo en Inglés | MEDLINE | ID: mdl-31682594

RESUMEN

Radiation therapy is an important and effective treatment option for prostate cancer, but high-risk patients are prone to relapse due to radioresistance of cancer cells. Molecular mechanisms that contribute to radioresistance are not fully understood. Novel computational strategies are needed to identify radioresistance driver genes from hundreds of gene copy number alterations. We developed a network-based approach based on lasso regression in combination with network propagation for the analysis of prostate cancer cell lines with acquired radioresistance to identify clinically relevant marker genes associated with radioresistance in prostate cancer patients. We analyzed established radioresistant cell lines of the prostate cancer cell lines DU145 and LNCaP and compared their gene copy number and expression profiles to their radiosensitive parental cells. We found that radioresistant DU145 showed much more gene copy number alterations than LNCaP and their gene expression profiles were highly cell line specific. We learned a genome-wide prostate cancer-specific gene regulatory network and quantified impacts of differentially expressed genes with directly underlying copy number alterations on known radioresistance marker genes. This revealed several potential driver candidates involved in the regulation of cancer-relevant processes. Importantly, we found that ten driver candidates from DU145 (ADAMTS9, AKR1B10, CXXC5, FST, FOXL1, GRPR, ITGA2, SOX17, STARD4, VGF) and four from LNCaP (FHL5, LYPLAL1, PAK7, TDRD6) were able to distinguish irradiated prostate cancer patients into early and late relapse groups. Moreover, in-depth in vitro validations for VGF (Neurosecretory protein VGF) showed that siRNA-mediated gene silencing increased the radiosensitivity of DU145 and LNCaP cells. Our computational approach enabled to predict novel radioresistance driver gene candidates. Additional preclinical and clinical studies are required to further validate the role of VGF and other candidate genes as potential biomarkers for the prediction of radiotherapy responses and as potential targets for radiosensitization of prostate cancer.


Asunto(s)
Perfilación de la Expresión Génica/métodos , Recurrencia Local de Neoplasia/genética , Tolerancia a Radiación/genética , Apoptosis , Biomarcadores de Tumor/genética , Línea Celular Tumoral , Biología Computacional/métodos , Dosificación de Gen/genética , Regulación Neoplásica de la Expresión Génica/genética , Redes Reguladoras de Genes/genética , Silenciador del Gen , Humanos , Masculino , Factores de Crecimiento Nervioso/genética , Neoplasias de la Próstata/genética , ARN Mensajero/genética , ARN Interferente Pequeño , Recurrencia , Transducción de Señal/genética , Transcriptoma/genética
11.
Gynecol Oncol ; 155(3): 473-482, 2019 12.
Artículo en Inglés | MEDLINE | ID: mdl-31703812

RESUMEN

OBJECTIVE: We analyzed comprehensive genomic sequencing results from paired ovarian cancer samples to identify changes in mutational events over time. METHODS: DNA from paired FFPE tumor samples from 50 ovarian cancer patients in the Clearity Foundation Data Repository was analyzed for genomic mutations (GM), copy number alterations (CNA), microsatellite status (MS), tumor mutation burden (TMB), and loss of heterozygosity (LOH) by hybrid-capture, next-generation sequencing of up to 315 genes. Genomic profiles were compared between samples from the same patient. Poor quality results excluded 6 pairs from all analyses and 9 from CNA or LOH. RESULTS: Forty-four patients with predominantly advanced stage disease (34, 77%) and serous histology (31, 70%) received a median of 3 intervening treatment regimens (range 1-13). Analysis of 22 primary and recurrent sample pairs and 22 recurrent tumor pairs detected a median of 2 GM (range 0-5) and 1 CNA (range 0-6)/sample. TMB, MS, and LOH results were mostly concordant across paired samples. GM were consistent across most pairs [32/44 (73%) concordant], while CNA concordance was less [18/35 (51%)]. No changes were detected in therapeutically relevant GM, but 23% of patients had GM or CNA in the second sample that affect clinical trial eligibility. CONCLUSIONS: Paired ovarian cancer samples demonstrate stable genomic alterations across time. However, discordance was observed for some genes used as eligibility criteria for molecularly targeted clinical trials. Repeat tumor testing may be useful in cases where eligibility for such trials is deemed important after consideration of testing costs and potential clinical benefit.


Asunto(s)
Ensayos Clínicos como Asunto/métodos , Neoplasias Ováricas/genética , Selección de Paciente , Adulto , Anciano , Análisis Mutacional de ADN , ADN de Neoplasias/genética , Femenino , Dosificación de Gen , Secuenciación de Nucleótidos de Alto Rendimiento/métodos , Humanos , Pérdida de Heterocigocidad , Inestabilidad de Microsatélites , Persona de Mediana Edad , Estadificación de Neoplasias , Neoplasias Ováricas/patología , Neoplasias Ováricas/terapia
12.
Microb Cell Fact ; 18(1): 187, 2019 Nov 01.
Artículo en Inglés | MEDLINE | ID: mdl-31675969

RESUMEN

BACKGROUND: The PAOX1-based expression system is the most widely used for producing recombinant proteins in the methylotrophic yeast Pichia pastoris (Komagataella phaffii). Despite relevant recent advances in regulation of the methanol utilization (MUT) pathway have been made, the role of specific growth rate (µ) in AOX1 regulation remains unknown, and therefore, its impact on protein production kinetics is still unclear. RESULTS: The influence of heterologous gene dosage, and both, operational mode and strategy, on culture physiological state was studied by cultivating the two PAOX1-driven Candida rugosa lipase 1 (Crl1) producer clones. Specifically, a clone integrating a single expression cassette of CRL1 was compared with one containing three cassettes over broad dilution rate and µ ranges in both chemostat and fed-batch cultivations. Chemostat cultivations allowed to establish the impact of µ on the MUT-related MIT1 pool which leads to a bell-shaped relationship between µ and PAOX1-driven gene expression, influencing directly Crl1 production kinetics. Also, chemostat and fed-batch cultivations exposed the favorable effects of increasing the CRL1 gene dosage (up to 2.4 fold in qp) on Crl1 production with no significant detrimental effects on physiological capabilities. CONCLUSIONS: PAOX1-driven gene expression and Crl1 production kinetics in P. pastoris were successfully correlated with µ. In fact, µ governs MUT-related MIT1 amount that triggers PAOX1-driven gene expression-heterologous genes included-, thus directly influencing the production kinetics of recombinant protein.


Asunto(s)
Oxidorreductasas de Alcohol/genética , Proteínas Fúngicas/metabolismo , Metanol/metabolismo , Pichia/metabolismo , Proteínas Recombinantes/metabolismo , Dosificación de Gen , Expresión Génica , Regulación Fúngica de la Expresión Génica , Pichia/genética , Regiones Promotoras Genéticas
13.
Nat Commun ; 10(1): 5143, 2019 11 13.
Artículo en Inglés | MEDLINE | ID: mdl-31723142

RESUMEN

Molecular determinants governing the evolution of tumor subclones toward phylogenetic branches or fixation remain unknown. Using sequencing data, we model the propagation and selection of clones expressing distinct categories of BRAF mutations to estimate their evolutionary trajectories. We show that strongly activating BRAF mutations demonstrate hard sweep dynamics, whereas mutations with less pronounced activation of the BRAF signaling pathway confer soft sweeps or are subclonal. We use clonal reconstructions to estimate the strength of "driver" selection in individual tumors. Using tumors cells and human-derived murine xenografts, we show that tumor sweep dynamics can significantly affect responses to targeted inhibitors of BRAF/MEK or DNA damaging agents. Our study uncovers patterns of distinct BRAF clonal evolutionary dynamics and nominates therapeutic strategies based on the identity of the BRAF mutation and its clonal composition.


Asunto(s)
Evolución Clonal/genética , Neoplasias/genética , Proteínas Proto-Oncogénicas B-raf/genética , Adenocarcinoma del Pulmón/patología , Animales , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Células Clonales , Daño del ADN , Dosificación de Gen , Sitios Genéticos , Humanos , Ratones , Quinasas de Proteína Quinasa Activadas por Mitógenos/antagonistas & inhibidores , Quinasas de Proteína Quinasa Activadas por Mitógenos/metabolismo , Mutación/genética , Fenotipo , Inhibidores de Proteínas Quinasas/farmacología
14.
Vet Parasitol ; 276: 108977, 2019 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-31775104

RESUMEN

The aims of this study were to monitor the change in Theileria orientalis Ikeda type infection intensity, haematocrit, milk production and reproduction on three New Zealand spring calving dairy herds, over the 2014-2015 milking season. Three spring calving dairy farms, A, B and C, from high risk (endemically stable), low risk (endemically unstable), and zero risk (disease-free) tick areas respectively were followed through the 2014-2015 milking season. On Farms, A and B, 100 cows were randomly selected at the first visit, and the same cows blood sampled every month thereafter, whilst on Farm C, the whole herd was blood sampled bimonthly (140 cows). Blood samples were tested for haematocrit, by centrifugation, and Ikeda infection intensity, using qPCR. Animals that were Ikeda type PCR positive at the first sampling were described as prevalence cases and cows that were negative at the first sampling and became PCR positive during the sampling period were described as incidence cases. Production and reproduction data were accessed through LIC MINDA® and milk production data was standardised to energy corrected milk (ECM). In addition, the effect of buparvaquone (BPQ) treatment on milk production was estimated on Farm B. The prevalence of infection at the first sampling was 100 % on Farm A, 57 % on Farm B and 26 % on Farm C. The incidence risk of infection over the sampling period on Farms B and C was 25 % and 2 % and the incident rate was 0.026 and 0.002 cases per cow-month respectively. The average infection intensity for prevalence cases on all farms was low throughout the milking season, <7000 Ikeda organisms/µL however, cases of anaemia still occurred. There was no direct effect of infection intensity on milk production or from being a prevalence case compared to an uninfected cow on milk production, across all farms. However, on Farm B there was a loss of 266 kg (95 % CI 82 ̶ 450) ECM (∼20 kg milk solids) for incidence cases and a loss of 458 kg (95 % CI 211 ̶ 710) of ECM for buparvaquone treated cows, compared to uninfected cows. No significant effect of Ikeda infection on reproduction could be shown for Farms B and C, reproductive data for Farm A was not available. The effect of T. orientalis Ikeda type infection on production and reproduction appears to be minimal once animals have passed through the acute phase of infection and reached the chronic, asymptomatic carrier phase of infection.


Asunto(s)
Lactancia , Reproducción , Theileriosis/fisiopatología , Animales , Antiprotozoarios/uso terapéutico , Bovinos , ADN Protozoario/sangre , Industria Lechera , Femenino , Dosificación de Gen , Hematócrito/veterinaria , Incidencia , Estudios Longitudinales , Naftoquinonas/uso terapéutico , Prevalencia , Estudios Prospectivos , Reacción en Cadena en Tiempo Real de la Polimerasa/veterinaria , Theileria/efectos de los fármacos , Theileria/genética , Theileriosis/tratamiento farmacológico , Theileriosis/epidemiología , Theileriosis/parasitología
15.
J Microbiol ; 57(12): 1041-1047, 2019 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-31758393

RESUMEN

Suppression analysis is used for the identification of new genes and genetic interactions when there is a notable phenotype available for genetic selection or screening. A random genomic DNA library constructed on a multi-copy plasmid is a useful tool for suppression analysis when one expects that an overdose of a few genes will suppress the phenotype. These libraries have been successfully used to determine the function of a gene by revealing genes whose functions are related to the gene of interest. They have also been used to identify the targets of chemical or biological agents by increasing the number of unaffected target gene products in a cell. In this article, I will discuss important considerations for constructing multicopy genomic DNA libraries. The protocol provided in this paper should be a useful guide for constructing genomic DNA libraries in many bacterial species for which multi-copy plasmids are available.


Asunto(s)
Bacterias/genética , Biblioteca de Genes , Genómica , ADN Bacteriano , Dosificación de Gen , Genes Bacterianos , Fenotipo , Plásmidos , Selección Genética
16.
PLoS Genet ; 15(10): e1008357, 2019 10.
Artículo en Inglés | MEDLINE | ID: mdl-31609978

RESUMEN

Nonsyndromic orofacial cleft (NSOFC) is a severe birth defect that occurs early in embryonic development and includes the subtypes cleft palate only (CPO), cleft lip only (CLO) and cleft lip with cleft palate (CLP). Given a lack of specific genetic factor analysis for CPO and CLO, the present study aimed to dissect the landscape of genetic factors underlying the pathogenesis of these two subtypes using 6,986 cases and 10,165 controls. By combining a genome-wide association study (GWAS) for specific subtypes of CPO and CLO, as well as functional gene network and ontology pathway analysis, we identified 18 genes/loci that surpassed genome-wide significance (P < 5 × 10-8) responsible for NSOFC, including nine for CPO, seven for CLO, two for both conditions and four that contribute to the CLP subtype. Among these 18 genes/loci, 14 are novel and identified in this study and 12 contain developmental transcription factors (TFs), suggesting that TFs are the key factors for the pathogenesis of NSOFC subtypes. Interestingly, we observed an opposite effect of the genetic variants in the IRF6 gene for CPO and CLO. Moreover, the gene expression dosage effect of IRF6 with two different alleles at the same single-nucleotide polymorphism (SNP) plays important roles in driving CPO or CLO. In addition, PAX9 is a key TF for CPO. Our findings define subtypes of NSOFC using genetic factors and their functional ontologies and provide a clue to improve their diagnosis and treatment in the future.


Asunto(s)
Encéfalo/anomalías , Labio Leporino/genética , Fisura del Paladar/genética , Factores Reguladores del Interferón/genética , Factor de Transcripción PAX9/genética , Alelos , Encéfalo/fisiopatología , Labio Leporino/fisiopatología , Fisura del Paladar/fisiopatología , Dosificación de Gen/genética , Regulación de la Expresión Génica/genética , Redes Reguladoras de Genes/genética , Predisposición Genética a la Enfermedad , Estudio de Asociación del Genoma Completo , Genotipo , Humanos , Polimorfismo de Nucleótido Simple/genética
17.
Curr Med Sci ; 39(5): 759-765, 2019 Oct.
Artículo en Inglés | MEDLINE | ID: mdl-31612394

RESUMEN

Extrahepatic metastasis confers unfavorable patient prognosis in patients with hepatocellular carcinoma (HCC), however, reliable markers allowing prediction of extrahepatic metastasis at the time of initial diagnosis are still lacking. This study was to identify gene-level copy number aberrations (CNAs) related to extrahepatic metastasis-free survival of HCC patients, and further examine the associations between CNAs and gene expression. Array comparative genomic hybridization (aCGH) and expression array were used to analyze gene CNAs and expression levels, respectively. The associations between CNAs of a panel of 20 genes and extrahepatic metastasis-free survival were analyzed in 66 patients with follow-up period of 1.6-90.5 months. The gene expression levels between HCCs with and without gene CNA were compared in 109 patients with HCC. We observed that gains at MDM4 and BCL2L1, and losses at APC and FBXW7 were independent prognostic markers for extrahepatic metastasis-free survival of HCC patients. Integration analysis of aCGH and expression data showed that MDM4 and BCL2L1 were significantly upregulated in HCCs with gene gain, while APC and FBXW7 were significantly downregulated in HCCs with gene loss. We concluded that gene gains at MDM4 and BCL2L1, and losses at APC and FBXW7, with concordant expression changes, were associated with extrahepatic metastasis-free survival of HCC patients and have potential to act as novel prognostic markers.


Asunto(s)
Proteína de la Poliposis Adenomatosa del Colon/genética , Biomarcadores de Tumor/genética , Carcinoma Hepatocelular/genética , Proteínas de Ciclo Celular/genética , Proteína 7 que Contiene Repeticiones F-Box-WD/genética , Regulación Neoplásica de la Expresión Génica , Neoplasias Hepáticas/genética , Proteínas Proto-Oncogénicas/genética , Proteína bcl-X/genética , Proteína de la Poliposis Adenomatosa del Colon/metabolismo , Adulto , Biomarcadores de Tumor/metabolismo , Carcinoma Hepatocelular/diagnóstico , Carcinoma Hepatocelular/metabolismo , Carcinoma Hepatocelular/mortalidad , Proteínas de Ciclo Celular/metabolismo , Hibridación Genómica Comparativa , Proteína 7 que Contiene Repeticiones F-Box-WD/metabolismo , Femenino , Estudios de Seguimiento , Dosificación de Gen , Humanos , Neoplasias Hepáticas/diagnóstico , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/mortalidad , Metástasis Linfática , Masculino , Persona de Mediana Edad , Pronóstico , Proteínas Proto-Oncogénicas/metabolismo , Estudios Retrospectivos , Análisis de Supervivencia , Proteína bcl-X/metabolismo
18.
Nat Commun ; 10(1): 4495, 2019 10 03.
Artículo en Inglés | MEDLINE | ID: mdl-31582743

RESUMEN

Aneuploidy is a major source of gene dosage imbalance due to copy number alterations (CNA), and viable human trisomies are model disorders of altered gene expression. We study gene and allele-specific expression (ASE) of 9668 single-cell fibroblasts from trisomy 21 (T21) discordant twins and from mosaic T21, T18, T13 and T8. We examine 928 single cells with deep scRNAseq. Expected and observed overexpression of trisomic genes in trisomic vs. diploid bulk RNAseq is not detectable in trisomic vs. diploid single cells. Instead, for trisomic genes with low-to-average expression, their altered gene dosage is mainly due to the higher fraction of trisomic cells simultaneously expressing these genes, in agreement with a stochastic 2-state burst-like model of transcription. These results, confirmed in a further analysis of 8740 single fibroblasts with shallow scRNAseq, suggest that the specific transcriptional profile of each gene contributes to the phenotypic variability of trisomies. We propose an improved model to understand the effects of CNA and, generally, of gene regulation on gene dosage imbalance.


Asunto(s)
Variaciones en el Número de Copia de ADN , Dosificación de Gen , Modelos Genéticos , Transcriptoma/genética , Trisomía/genética , Alelos , Cromosomas Humanos Par 13/genética , Cromosomas Humanos Par 18/genética , Cromosomas Humanos Par 8/genética , Síndrome de Down/genética , Femenino , Fibroblastos , Perfilación de la Expresión Génica , Humanos , Masculino , Mosaicismo , Fenotipo , Análisis de la Célula Individual
19.
Aquat Toxicol ; 217: 105328, 2019 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-31629202

RESUMEN

Benthic ecosystems have come under intense pressure, due to eutrophication-driven oxygen decline and industrial metal contamination. One of the most toxic metals is Cadmium (Cd), which is lethal to many aquatic organisms already at low concentrations. Denitrification by facultative anaerobic microorganisms is an essential process to transform, but also to remove, excess nitrate in eutrophied systems. Cd has been shown to decrease denitrification and sequester free sulfide, which is available when oxygen is scarce and generally inhibits complete denitrification (i.e. N2O to N2). In polluted sediments, an interaction between oxygen and Cd may influence denitrification and this relationship has not been studied. For example, in the Baltic Sea some sediments are double exposed to both Cd and hypoxia. In this study, we examined how the double exposure of Cd and fluctuations in oxygen affects denitrification in Baltic Sea sediment. Results show that oxygen largely regulated N2O and N2 production after 21 days of exposure to Cd (ranging from 0 to 500 µg/L, 5 different treatments, measured by the isotope pairing technique (IPT)). In the high Cd treatment (500 µg/L) the variation in N2 production increased compared to the other treatments. Increases in N2 production are suggested to be an effect of 1) enhanced nitrification that increases NO3- availability thus stimulating denitrification, and 2) Cd successfully sequestrating sulfide (yielding CdS), which allows for full denitrification to N2. The in situ field sediment contained initially high Cd concentrations in the pore water (∼10 µg/L) and microbial communities might already have been adapted to metal stress, making the effect of low Cd levels negligible. Here we show that high levels of cadmium pollution might increase N2 production and influence nitrogen cycling in marine sediments.


Asunto(s)
Cadmio/toxicidad , Desnitrificación , Sedimentos Geológicos/química , Dosificación de Gen , Nitrato-Reductasa/genética , Nitrato-Reductasa/metabolismo , Nitratos/análisis , Nitrificación/efectos de los fármacos , Nitrógeno/análisis , Océanos y Mares , Oxidorreductasas/genética , Oxidorreductasas/metabolismo , Oxígeno/análisis , Porosidad , ARN Mensajero/genética , ARN Mensajero/metabolismo , Contaminantes Químicos del Agua/toxicidad
20.
Enzyme Microb Technol ; 131: 109382, 2019 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-31615681

RESUMEN

In an effort to find a suitable genetic background for efficient cellulolytic secretion, genetically diverse strains were transformed to produce core fungal cellulases namely, ß-glucosidase (BGLI), endoglucanase (EGII) and cellobiohydrolase (CBHI) in various combinations and expression configurations. The secreted enzyme activity levels, gene copy number, substrate specificities, as well as hydrolysis and fermentation yields of the transformants were analysed. The effectiveness of the partially cellulolytic yeast transformants to convert two different pre-treated corn residues, namely corn cob and corn husk was then explored. Higher secretion titers were achieved by cellulolytic strains with the YI13 genetic background and cellulolytic transformants produced up to 1.34 fold higher glucose concentrations (g/L) than a control composed of equal amounts of each enzyme type. The transformant co-producing BGLI and EGII in a secreted ratio of 1:15 (cellulase activity unit per gram dry cell weight) converted 56.5% of the cellulose present in corn cob to glucose in hydrolysis experiments and yielded 4.05 g/L ethanol in fermentations. We demonstrate that the choice of optimal genetic background and cellulase activity secretion ratio can improve cellulosic ethanol production by consolidated bioprocessing yeast strains.


Asunto(s)
Celulasa/metabolismo , Expresión Génica , Levaduras/enzimología , Levaduras/metabolismo , Zea mays/metabolismo , Biotransformación , Celulasa/genética , Diploidia , Fermentación , Dosificación de Gen , Hidrólisis , Levaduras/genética
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