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1.
Nat Commun ; 13(1): 2300, 2022 04 28.
Artículo en Inglés | MEDLINE | ID: mdl-35484108

RESUMEN

While the genomes of normal tissues undergo dynamic changes over time, little is understood about the temporal-spatial dynamics of genomes in premalignant tissues that progress to cancer compared to those that remain cancer-free. Here we use whole genome sequencing to contrast genomic alterations in 427 longitudinal samples from 40 patients with stable Barrett's esophagus compared to 40 Barrett's patients who progressed to esophageal adenocarcinoma (ESAD). We show the same somatic mutational processes are active in Barrett's tissue regardless of outcome, with high levels of mutation, ESAD gene and focal chromosomal alterations, and similar mutational signatures. The critical distinction between stable Barrett's versus those who progress to cancer is acquisition and expansion of TP53-/- cell populations having complex structural variants and high-level amplifications, which are detectable up to six years prior to a cancer diagnosis. These findings reveal the timing of common somatic genome dynamics in stable Barrett's esophagus and define key genomic features specific to progression to esophageal adenocarcinoma, both of which are critical for cancer prevention and early detection strategies.


Asunto(s)
Adenocarcinoma , Esófago de Barrett , Neoplasias Esofágicas , Adenocarcinoma/patología , Esófago de Barrett/genética , Esófago de Barrett/patología , Progresión de la Enfermedad , Neoplasias Esofágicas/patología , Humanos
2.
Cell ; 183(1): 197-210.e32, 2020 10 01.
Artículo en Inglés | MEDLINE | ID: mdl-33007263

RESUMEN

Cancer genomes often harbor hundreds of somatic DNA rearrangement junctions, many of which cannot be easily classified into simple (e.g., deletion) or complex (e.g., chromothripsis) structural variant classes. Applying a novel genome graph computational paradigm to analyze the topology of junction copy number (JCN) across 2,778 tumor whole-genome sequences, we uncovered three novel complex rearrangement phenomena: pyrgo, rigma, and tyfonas. Pyrgo are "towers" of low-JCN duplications associated with early-replicating regions, superenhancers, and breast or ovarian cancers. Rigma comprise "chasms" of low-JCN deletions enriched in late-replicating fragile sites and gastrointestinal carcinomas. Tyfonas are "typhoons" of high-JCN junctions and fold-back inversions associated with expressed protein-coding fusions, breakend hypermutation, and acral, but not cutaneous, melanomas. Clustering of tumors according to genome graph-derived features identified subgroups associated with DNA repair defects and poor prognosis.


Asunto(s)
Variación Estructural del Genoma/genética , Genómica/métodos , Neoplasias/genética , Inversión Cromosómica/genética , Cromotripsis , Variaciones en el Número de Copia de ADN/genética , Reordenamiento Génico/genética , Genoma Humano/genética , Humanos , Mutación/genética , Secuenciación Completa del Genoma/métodos
3.
BMC Bioinformatics ; 20(1): 431, 2019 Aug 19.
Artículo en Inglés | MEDLINE | ID: mdl-31426747

RESUMEN

BACKGROUND: Protein pulldown using Methyl-CpG binding domain (MBD) proteins followed by high-throughput sequencing is a common method to determine DNA methylation. Algorithms have been developed to estimate absolute methylation level from read coverage generated by affinity enrichment-based techniques, but the most accurate one for MBD-seq data requires additional data from an SssI-treated Control experiment. RESULTS: Using our previous characterizations of Methyl-CpG/MBD2 binding in the context of an MBD pulldown experiment, we build a model of expected MBD pulldown reads as drawn from SssI-treated DNA. We use the program BayMeth to evaluate the effectiveness of this model by substituting calculated SssI Control data for the observed SssI Control data. By comparing methylation predictions against those from an RRBS data set, we find that BayMeth run with our modeled SssI Control data performs better than BayMeth run with observed SssI Control data, on both 100 bp and 10 bp windows. Adapting the model to an external data set solely by changing the average fragment length, our calculated data still informs the BayMeth program to a similar level as observed data in predicting methylation state on a pulldown data set with matching WGBS estimates. CONCLUSION: In both internal and external MBD pulldown data sets tested in this study, BayMeth used with our modeled pulldown coverage performs better than BayMeth run without the inclusion of any estimate of SssI Control pulldown, and is comparable to - and in some cases better than - using observed SssI Control data with the BayMeth program. Thus, our MBD pulldown alignment model can improve methylation predictions without the need to perform additional control experiments.


Asunto(s)
Biología Computacional/métodos , Metilación de ADN/genética , ADN-Citosina Metilasas/metabolismo , ADN/metabolismo , Modelos Biológicos , Alineación de Secuencia , Algoritmos , Emparejamiento Base , Cromosomas Humanos Par 7/genética , Islas de CpG/genética , Genoma Humano , Secuenciación de Nucleótidos de Alto Rendimiento , Humanos , Dominio de Unión a CpG-Metil , Análisis de Secuencia de ADN/métodos
4.
Genome Med ; 11(1): 14, 2019 03 12.
Artículo en Inglés | MEDLINE | ID: mdl-30867038

RESUMEN

It was highlighted that in the original article [1] the Availability of data and materials section was incorrect.

5.
Genome Med ; 10(1): 17, 2018 02 27.
Artículo en Inglés | MEDLINE | ID: mdl-29486792

RESUMEN

BACKGROUND: Use of aspirin and other non-steroidal anti-inflammatory drugs (NSAIDs) has been shown to protect against tetraploidy, aneuploidy, and chromosomal alterations in the metaplastic condition Barrett's esophagus (BE) and to lower the incidence and mortality of esophageal adenocarcinoma (EA). The esophagus is exposed to both intrinsic and extrinsic mutagens resulting from gastric reflux, chronic inflammation, and exposure to environmental carcinogens such as those found in cigarettes. Here we test the hypothesis that NSAID use inhibits accumulation of point mutations/indels during somatic genomic evolution in BE. METHODS: Whole exome sequences were generated from 82 purified epithelial biopsies and paired blood samples from a cross-sectional study of 41 NSAID users and 41 non-users matched by sex, age, smoking, and continuous time using or not using NSAIDs. RESULTS: NSAID use reduced overall frequency of point mutations across the spectrum of mutation types, lowered the frequency of mutations even when adjusted for both TP53 mutation and smoking status, and decreased the prevalence of clones with high variant allele frequency. Never smokers who consistently used NSAIDs had fewer point mutations in signature 17, which is commonly found in EA. NSAID users had, on average, a 50% reduction in functional gene mutations in nine cancer-associated pathways and also had less diversity in pathway mutational burden compared to non-users. CONCLUSIONS: These results indicate NSAID use functions to limit overall mutations on which selection can act and supports a model in which specific mutant cell populations survive or expand better in the absence of NSAIDs.


Asunto(s)
Antiinflamatorios no Esteroideos/uso terapéutico , Esófago de Barrett/tratamiento farmacológico , Esófago de Barrett/genética , Exoma/genética , Mutación/genética , Variaciones en el Número de Copia de ADN/genética , Frecuencia de los Genes/genética , Humanos , Pérdida de Heterocigocidad , Mutagénesis/genética
6.
Biophys J ; 111(12): 2551-2561, 2016 Dec 20.
Artículo en Inglés | MEDLINE | ID: mdl-28002732

RESUMEN

Determining the pattern of methylation at CpG dinucleotides in a cell remains an essential component of epigenetic profiling. The correlations among methylation, gene expression, and accompanying disease have just begun to be explored. Many experiments for sensing methylation use a relatively inexpensive, high-throughput approach with a methyl-binding domain (MBD) protein that preferentially binds to methylated CpGs. Here, we characterize the cooperativity and sequence specificity of MBD2-DNA binding in a pulldown experiment revealing three potential biases in such experiments. The first is caused by steric clashes between two MBD2 proteins at mCpGs separated by 2 bp or less, which suggests that simultaneous binding at these sites is inhibited. This is confirmed by comparing input versus pulldown high-throughput sequencing data on M.SssI-treated samples, from which we also find that pulldown efficiency sharply increases for DNA fragments with four or more mCpGs. Analysis of these two data sets was again employed to investigate MBD2's sequence preferences surrounding a methylated CpG (mCpG). In comparing the distributions of bases at positions with respect to an mCpG, statistically significant preferences for certain bases were found, although the corresponding biases in pulldown efficiency were all <5%. While this suggests that mCpG sequence context can mostly be ignored in MBD2 binding, the statistical certainty exhibited by our high-throughput approach bodes well for future applications.


Asunto(s)
Islas de CpG/genética , Proteínas de Unión al ADN/metabolismo , ADN/genética , ADN/metabolismo , Secuencia de Bases , ADN/química , Metilación , Modelos Moleculares , Conformación de Ácido Nucleico , Unión Proteica , Especificidad por Sustrato
7.
Nucleic Acids Res ; 44(1): 304-14, 2016 Jan 08.
Artículo en Inglés | MEDLINE | ID: mdl-26673707

RESUMEN

Cap homeostasis is a cyclical process of decapping and recapping that maintains the cap on a subset of the cytoplasmic transcriptome. Interfering with cytoplasmic capping results in the redistribution of target transcripts from polysomes to non-translating mRNPs, where they accumulate in an uncapped but nonetheless stable form. It is generally thought that decapping is preceded by shortening of the poly(A) tail to a length that can no longer support translation. Therefore recapped target transcripts would either have to undergo cytoplasmic polyadenylation or retain a reasonably long poly(A) tail if they are to return to the translating pool. In cells that are inhibited for cytoplasmic capping there is no change in the overall distribution of poly(A) lengths or in the elution profile of oligo(dT)-bound targets. Poly(A) tail lengths were similar for target mRNAs on polysomes or in non-translating mRNPs, and the presence of polyadenylated uncapped mRNA in mRNPs was confirmed by separation into capped and uncapped pools prior to assay. Finally, in silico analysis of cytoplasmic capping targets revealed significant correlations with genes encoding transcripts with uridylated or multiply modified 3' ends, and genes possessing multiple 3'-untranslated regions (UTRs) generated by alternative cleavage and polyadenylation.


Asunto(s)
Poli A , Poliadenilación , Caperuzas de ARN , ARN Mensajero/genética , ARN Mensajero/metabolismo , Regiones no Traducidas 3' , Animales , Línea Celular , Citoplasma/metabolismo , Homeostasis , Polirribosomas/metabolismo , Biosíntesis de Proteínas , ARN no Traducido/genética , ARN no Traducido/metabolismo , Ribonucleoproteínas/metabolismo
8.
FEBS Lett ; 589(3): 279-84, 2015 Jan 30.
Artículo en Inglés | MEDLINE | ID: mdl-25541487

RESUMEN

In mammalian transcriptomes approximately 25% of 5' ends determined by Capped Analysis of Gene Expression (CAGE) map to locations within spliced exons. The current study sought to determine if the cytoplasmic capping complex participates in generating these downstream CAGE tags. 5'-RACE was used to amplify the uncapped ends of target transcripts that accumulate when cytoplasmic capping is blocked. Sequencing of these RACE products mapped the positions of uncapped ends either exactly to or just downstream of archived CAGE tags. These findings support a role for cytoplasmic capping in generating the downstream capped ends identified by CAGE.


Asunto(s)
Regulación de la Expresión Génica , Caperuzas de ARN/genética , Sitios de Empalme de ARN/genética , ARN Largo no Codificante/genética , Animales , Citoplasma/genética , Exones/genética , Genoma , ARN Mensajero/genética , Transcriptoma/genética
9.
Nucleic Acids Res ; 42(21): 13370-83, 2014 Dec 01.
Artículo en Inglés | MEDLINE | ID: mdl-25378333

RESUMEN

LepA is a paralog of EF-G found in all bacteria. Deletion of lepA confers no obvious growth defect in Escherichia coli, and the physiological role of LepA remains unknown. Here, we identify nine strains (ΔdksA, ΔmolR1, ΔrsgA, ΔtatB, ΔtonB, ΔtolR, ΔubiF, ΔubiG or ΔubiH) in which ΔlepA confers a synthetic growth phenotype. These strains are compromised for gene regulation, ribosome assembly, transport and/or respiration, indicating that LepA contributes to these functions in some way. We also use ribosome profiling to deduce the effects of LepA on translation. We find that loss of LepA alters the average ribosome density (ARD) for hundreds of mRNA coding regions in the cell, substantially reducing ARD in many cases. By contrast, only subtle and codon-specific changes in ribosome distribution along mRNA are seen. These data suggest that LepA contributes mainly to the initiation phase of translation. Consistent with this interpretation, the effect of LepA on ARD is related to the sequence of the Shine-Dalgarno region. Global perturbation of gene expression in the ΔlepA mutant likely explains most of its phenotypes.


Asunto(s)
Proteínas de Escherichia coli/fisiología , Escherichia coli/genética , Iniciación de la Cadena Peptídica Traduccional , Factores de Iniciación de Péptidos/fisiología , Factores Procarióticos de Iniciación/fisiología , Dominio Catalítico , Escherichia coli/enzimología , Proteínas de Escherichia coli/química , Proteínas de Escherichia coli/genética , Proteínas de Escherichia coli/metabolismo , GTP Fosfohidrolasas/metabolismo , Eliminación de Gen , Extensión de la Cadena Peptídica de Translación , Factores de Iniciación de Péptidos/química , Factores de Iniciación de Péptidos/genética , Factores de Iniciación de Péptidos/metabolismo , Fenotipo , Factores Procarióticos de Iniciación/química , Factores Procarióticos de Iniciación/genética , Factores Procarióticos de Iniciación/metabolismo , Estructura Terciaria de Proteína , ARN Mensajero/análisis , Ribosomas/metabolismo
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