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1.
Nat Commun ; 12(1): 793, 2021 02 04.
Artigo em Inglês | MEDLINE | ID: mdl-33542240

RESUMO

Adenosine-to-inosine (A-to-I) editing of eukaryotic cellular RNAs is essential for protection against auto-immune disorders. Editing is carried out by ADAR1, whose innate immune response-specific cytoplasmic isoform possesses a Z-DNA binding domain (Zα) of unknown function. Zα also binds to CpG repeats in RNA, which are a hallmark of Z-RNA formation. Unexpectedly, Zα has been predicted - and in some cases even shown - to bind to specific regions within mRNA and rRNA devoid of such repeats. Here, we use NMR, circular dichroism, and other biophysical approaches to demonstrate and characterize the binding of Zα to mRNA and rRNA fragments. Our results reveal a broad range of RNA sequences that bind to Zα and adopt Z-RNA conformations. Binding is accompanied by destabilization of neighboring A-form regions which is similar in character to what has been observed for B-Z-DNA junctions. The binding of Zα to non-CpG sequences is specific, cooperative and occurs with an affinity in the low micromolar range. This work allows us to propose a model for how Zα could influence the RNA binding specificity of ADAR1.


Assuntos
Adenosina Desaminase/metabolismo , Elementos Alu/genética , Domínios Proteicos , RNA Ribossômico/metabolismo , Proteínas de Ligação a RNA/metabolismo , Adenosina Desaminase/genética , Adenosina Desaminase/isolamento & purificação , Adenosina Desaminase/ultraestrutura , Dicroísmo Circular , Imunidade Inata , Ressonância Magnética Nuclear Biomolecular , Conformação de Ácido Nucleico , Motivo de Reconhecimento de RNA , RNA Ribossômico/genética , RNA Ribossômico/imunologia , RNA Ribossômico/ultraestrutura , Proteínas de Ligação a RNA/genética , Proteínas de Ligação a RNA/isolamento & purificação , Proteínas de Ligação a RNA/ultraestrutura , Proteínas Recombinantes/genética , Proteínas Recombinantes/isolamento & purificação , Proteínas Recombinantes/metabolismo , Proteínas Recombinantes/ultraestrutura
2.
BMC Bioinformatics ; 21(Suppl 18): 578, 2020 Dec 30.
Artigo em Inglês | MEDLINE | ID: mdl-33375933

RESUMO

BACKGROUND: As the number of RNA-seq datasets that become available to explore transcriptome diversity increases, so does the need for easy-to-use comprehensive computational workflows. Many available tools facilitate analyses of one of the two major mechanisms of transcriptome diversity, namely, differential expression of isoforms due to alternative splicing, while the second major mechanism-RNA editing due to post-transcriptional changes of individual nucleotides-remains under-appreciated. Both these mechanisms play an essential role in physiological and diseases processes, including cancer and neurological disorders. However, elucidation of RNA editing events at transcriptome-wide level requires increasingly complex computational tools, in turn resulting in a steep entrance barrier for labs who are interested in high-throughput variant calling applications on a large scale but lack the manpower and/or computational expertise. RESULTS: Here we present an easy-to-use, fully automated, computational pipeline (Automated Isoform Diversity Detector, AIDD) that contains open source tools for various tasks needed to map transcriptome diversity, including RNA editing events. To facilitate reproducibility and avoid system dependencies, the pipeline is contained within a pre-configured VirtualBox environment. The analytical tasks and format conversions are accomplished via a set of automated scripts that enable the user to go from a set of raw data, such as fastq files, to publication-ready results and figures in one step. A publicly available dataset of Zika virus-infected neural progenitor cells is used to illustrate AIDD's capabilities. CONCLUSIONS: AIDD pipeline offers a user-friendly interface for comprehensive and reproducible RNA-seq analyses. Among unique features of AIDD are its ability to infer RNA editing patterns, including ADAR editing, and inclusion of Guttman scale patterns for time series analysis of such editing landscapes. AIDD-based results show importance of diversity of ADAR isoforms, key RNA editing enzymes linked with the innate immune system and viral infections. These findings offer insights into the potential role of ADAR editing dysregulation in the disease mechanisms, including those of congenital Zika syndrome. Because of its automated all-inclusive features, AIDD pipeline enables even a novice user to easily explore common mechanisms of transcriptome diversity, including RNA editing landscapes.


Assuntos
Software , Transcriptoma , Adenosina Desaminase/genética , Adenosina Desaminase/metabolismo , Expressão Gênica , Ontologia Genética , Humanos , Análise de Componente Principal , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Edição de RNA , RNA-Seq , Células-Tronco/citologia , Células-Tronco/metabolismo , Células-Tronco/virologia , Zika virus/fisiologia
3.
Zhonghua Yi Xue Yi Chuan Xue Za Zhi ; 37(11): 1233-1235, 2020 Nov 10.
Artigo em Chinês | MEDLINE | ID: mdl-33179227

RESUMO

OBJECTIVE: To detect variants of ADAR1 gene in two Chinese pedigrees affected with dyschromatosis symmetrica hereditaria (DSH). METHODS: Clinical data and peripheral blood samples of the pedigrees were collected. All exons of the ADAR1 gene were amplified by PCR and subjected to Sanger sequencing. Suspected pathogenic variants were validated among other members of the pedigrees and 100 unrelated healthy controls. RESULTS: For pedigree 1, Sanger sequencing has identified a heterozygous missense variant c.3002G>C (p.Asp968His) in exon 11 of the ADAR1 gene in the proband and his father. For pedigree 2, a novel nonsense variant c.3145C>T (p.Gln1049Ter) was identified in exon 12 of the ADAR1 gene in the proband and his son, which were previously unreported and absent among the healthy controls. CONCLUSION: The c.3002G>C (p.Asp968His) and c.3145C>T (p.Gln1049Ter)variants of the ADAR1 gene probably underlay the DSH in the two pedigrees.


Assuntos
Adenosina Desaminase/genética , Transtornos da Pigmentação/congênito , Proteínas de Ligação a RNA/genética , Humanos , Mutação , Linhagem , Transtornos da Pigmentação/genética
4.
Nat Commun ; 11(1): 5130, 2020 10 12.
Artigo em Inglês | MEDLINE | ID: mdl-33046702

RESUMO

Adenosine Deaminases that act on RNA (ADARs) are enzymes that catalyze adenosine to inosine conversion in dsRNA, a common form of RNA editing. Mutations in the human ADAR1 gene are known to cause disease and recent studies have identified ADAR1 as a potential therapeutic target for a subset of cancers. However, efforts to define the mechanistic effects for disease associated ADAR1 mutations and the rational design of ADAR1 inhibitors are limited by a lack of structural information. Here, we describe the combination of high throughput mutagenesis screening studies, biochemical characterization and Rosetta-based structure modeling to identify unique features of ADAR1. Importantly, these studies reveal a previously unknown zinc-binding site on the surface of the ADAR1 deaminase domain which is important for ADAR1 editing activity. Furthermore, we present structural models that explain known properties of this enzyme and make predictions about the role of specific residues in a surface loop unique to ADAR1.


Assuntos
Adenosina Desaminase/química , Adenosina Desaminase/genética , Proteínas de Ligação a RNA/química , Proteínas de Ligação a RNA/genética , Adenosina Desaminase/metabolismo , Sítios de Ligação , Humanos , Mutagênese , Mutação , Domínios Proteicos , Proteínas de Ligação a RNA/metabolismo , Zinco/química , Zinco/metabolismo
5.
PLoS Pathog ; 16(9): e1008842, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32898178

RESUMO

Signaling through retinoic acid inducible gene I (RIG-I) like receptors (RLRs) is tightly regulated, with activation occurring upon sensing of viral nucleic acids, and suppression mediated by negative regulators. Under homeostatic conditions aberrant activation of melanoma differentiation-associated protein-5 (MDA5) is prevented through editing of endogenous dsRNA by RNA editing enzyme Adenosine Deaminase Acting on RNA (ADAR1). In addition, ADAR1 is postulated to play pro-viral and antiviral roles during viral infections that are dependent or independent of RNA editing activity. Here, we investigated the importance of ADAR1 isoforms in modulating influenza A virus (IAV) replication and revealed the opposing roles for ADAR1 isoforms, with the nuclear p110 isoform restricting versus the cytoplasmic p150 isoform promoting IAV replication. Importantly, we demonstrate that p150 is critical for preventing sustained RIG-I signaling, as p150 deficient cells showed increased IFN-ß expression and apoptosis during IAV infection, independent of RNA editing activity. Taken together, the p150 isoform of ADAR1 is important for preventing sustained RIG-I induced IFN-ß expression and apoptosis during viral infection.


Assuntos
Adenosina Desaminase/metabolismo , Apoptose , Proteína DEAD-box 58/metabolismo , Vírus da Influenza A/fisiologia , Influenza Humana/metabolismo , Proteínas de Ligação a RNA/metabolismo , Transdução de Sinais , Replicação Viral , Células A549 , Adenosina Desaminase/genética , Proteína DEAD-box 58/genética , Células HEK293 , Humanos , Influenza Humana/genética , Isoenzimas/genética , Isoenzimas/metabolismo , Proteínas de Ligação a RNA/genética
6.
Science ; 369(6503): 566-571, 2020 07 31.
Artigo em Inglês | MEDLINE | ID: mdl-32732424

RESUMO

CRISPR-Cas-guided base editors convert A•T to G•C, or C•G to T•A, in cellular DNA for precision genome editing. To understand the molecular basis for DNA adenosine deamination by adenine base editors (ABEs), we determined a 3.2-angstrom resolution cryo-electron microscopy structure of ABE8e in a substrate-bound state in which the deaminase domain engages DNA exposed within the CRISPR-Cas9 R-loop complex. Kinetic and structural data suggest that ABE8e catalyzes DNA deamination up to ~1100-fold faster than earlier ABEs because of mutations that stabilize DNA substrates in a constrained, transfer RNA-like conformation. Furthermore, ABE8e's accelerated DNA deamination suggests a previously unobserved transient DNA melting that may occur during double-stranded DNA surveillance by CRISPR-Cas9. These results explain ABE8e-mediated base-editing outcomes and inform the future design of base editors.


Assuntos
Adenina/química , Adenosina Desaminase/química , Proteína 9 Associada à CRISPR/química , Sistemas CRISPR-Cas , DNA/química , Proteínas de Escherichia coli/química , Edição de Genes , Adenosina Desaminase/genética , Proteína 9 Associada à CRISPR/genética , Microscopia Crioeletrônica , Desaminação , Proteínas de Escherichia coli/genética
7.
DNA Cell Biol ; 39(10): 1862-1871, 2020 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-32845709

RESUMO

Sepsis is a life-threatening disorder and leads to organ dysfunction and death. Therefore, searching for more alternative biomarkers is of great significance for sepsis assessment and surveillance. In our study, the gene expression profiles of 163 samples from healthy controls and septic patients were analyzed and 8 gene co-expression modules were identified by constructing weighted gene co-expression network. The blue and yellow modules showed close correlations with the phenotypic trait "days postsepsis." Besides, differentially expressed genes (DEGs) over time in septic patients were screened using Short Time-series Expression Miner (STEM) program. The intersection of genes in the blue and yellow modules and DEGs, which were significantly enriched in "HTLV-1 infection" pathway, was analyzed with protein-protein interaction network. The logistic regression model based on these eight mRNAs was constructed to determine the type of the sample reliably. Eight vital genes CECR1, ANXA2, ELANE, CTSG, AZU1, PRTN3, LYZ, and DEFA4 presented high scores and may be associated with sepsis, which provided candidate biomarkers for sepsis.


Assuntos
Sepse/genética , Transcriptoma , Adenosina Desaminase/genética , Adenosina Desaminase/metabolismo , Anexina A2/genética , Anexina A2/metabolismo , Biomarcadores/metabolismo , Catepsina G/genética , Catepsina G/metabolismo , Redes Reguladoras de Genes , Humanos , Peptídeos e Proteínas de Sinalização Intercelular/genética , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Elastase de Leucócito/genética , Elastase de Leucócito/metabolismo , Mieloblastina/genética , Mieloblastina/metabolismo , Sepse/metabolismo , Sepse/patologia , Análise de Sobrevida , alfa-Defensinas/genética , alfa-Defensinas/metabolismo
8.
Science ; 369(6507): 1077-1084, 2020 08 28.
Artigo em Inglês | MEDLINE | ID: mdl-32855333

RESUMO

Bacteria and archaea are frequently attacked by viruses and other mobile genetic elements and rely on dedicated antiviral defense systems, such as restriction endonucleases and CRISPR, to survive. The enormous diversity of viruses suggests that more types of defense systems exist than are currently known. By systematic defense gene prediction and heterologous reconstitution, here we discover 29 widespread antiviral gene cassettes, collectively present in 32% of all sequenced bacterial and archaeal genomes, that mediate protection against specific bacteriophages. These systems incorporate enzymatic activities not previously implicated in antiviral defense, including RNA editing and retron satellite DNA synthesis. In addition, we computationally predict a diverse set of other putative defense genes that remain to be characterized. These results highlight an immense array of molecular functions that microbes use against viruses.


Assuntos
Adenosina Desaminase/química , Archaea/virologia , Vírus de Archaea/imunologia , Bactérias/virologia , Bacteriófagos/imunologia , Sistemas CRISPR-Cas , Edição de RNA , Adenosina Desaminase/classificação , Adenosina Desaminase/genética , Archaea/enzimologia , Proteínas Arqueais , Bactérias/enzimologia , Proteínas de Bactérias , Genes Arqueais , Genes Bacterianos , Domínios Proteicos
11.
Nat Biotechnol ; 38(7): 865-869, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32483365

RESUMO

We describe base editors that combine both cytosine and adenine base-editing functions. A codon-optimized fusion of the cytosine deaminase PmCDA1, the adenosine deaminase TadA and a Cas9 nickase (Target-ACEmax) showed a high median simultaneous C-to-T and A-to-G editing activity at 47 genomic targets. On-target as well as DNA and RNA off-target activities of Target-ACEmax were similar to those of existing single-function base editors.


Assuntos
Proteína 9 Associada à CRISPR/genética , Sistemas CRISPR-Cas/genética , DNA/genética , Edição de Genes , Adenina/metabolismo , Adenosina Desaminase/genética , Citosina/metabolismo , Desoxirribonuclease I/genética , Genoma Humano/genética , Glicoproteínas/genética , Guanina/metabolismo , Células HEK293 , Humanos , Mutação/genética , Proteínas Nucleares/genética , RNA/genética
12.
Sci Adv ; 6(25): eabb5813, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32596474

RESUMO

The COVID-19 outbreak has become a global health risk, and understanding the response of the host to the SARS-CoV-2 virus will help to combat the disease. RNA editing by host deaminases is an innate restriction process to counter virus infection, but it is not yet known whether this process operates against coronaviruses. Here, we analyze RNA sequences from bronchoalveolar lavage fluids obtained from coronavirus-infected patients. We identify nucleotide changes that may be signatures of RNA editing: adenosine-to-inosine changes from ADAR deaminases and cytosine-to-uracil changes from APOBEC deaminases. Mutational analysis of genomes from different strains of Coronaviridae from human hosts reveals mutational patterns consistent with those observed in the transcriptomic data. However, the reduced ADAR signature in these data raises the possibility that ADARs might be more effective than APOBECs in restricting viral propagation. Our results thus suggest that both APOBECs and ADARs are involved in coronavirus genome editing, a process that may shape the fate of both virus and patient.


Assuntos
Betacoronavirus/genética , Betacoronavirus/metabolismo , Infecções por Coronavirus/genética , Interações Hospedeiro-Patógeno/genética , Pneumonia Viral/genética , Edição de RNA/genética , Transcriptoma , Desaminases APOBEC/genética , Desaminases APOBEC/metabolismo , Adenosina Desaminase/genética , Adenosina Desaminase/metabolismo , Sequência de Bases/genética , Líquido da Lavagem Broncoalveolar/virologia , Infecções por Coronavirus/virologia , Genoma Viral/genética , Humanos , Taxa de Mutação , Nucleotídeos/genética , Nucleotídeos/metabolismo , Pandemias , Pneumonia Viral/virologia , RNA Viral/genética , Replicação Viral/genética
13.
Nucleic Acids Res ; 48(14): 7958-7972, 2020 08 20.
Artigo em Inglês | MEDLINE | ID: mdl-32597966

RESUMO

Adenosine deaminases acting on RNA (ADARs) are enzymes that convert adenosine to inosine in duplex RNA, a modification that exhibits a multitude of effects on RNA structure and function. Recent studies have identified ADAR1 as a potential cancer therapeutic target. ADARs are also important in the development of directed RNA editing therapeutics. A comprehensive understanding of the molecular mechanism of the ADAR reaction will advance efforts to develop ADAR inhibitors and new tools for directed RNA editing. Here we report the X-ray crystal structure of a fragment of human ADAR2 comprising its deaminase domain and double stranded RNA binding domain 2 (dsRBD2) bound to an RNA duplex as an asymmetric homodimer. We identified a highly conserved ADAR dimerization interface and validated the importance of these sequence elements on dimer formation via gel mobility shift assays and size exclusion chromatography. We also show that mutation in the dimerization interface inhibits editing in an RNA substrate-dependent manner for both ADAR1 and ADAR2.


Assuntos
Adenosina Desaminase/química , Adenosina Desaminase/metabolismo , Edição de RNA , RNA de Cadeia Dupla/metabolismo , Proteínas de Ligação a RNA/química , Proteínas de Ligação a RNA/metabolismo , Adenosina Desaminase/genética , Cristalografia por Raios X , Humanos , Modelos Moleculares , Mutação , Ligação Proteica , Domínios Proteicos , Multimerização Proteica , RNA de Cadeia Dupla/química , Proteínas de Ligação a RNA/genética
14.
Ocul Immunol Inflamm ; 28(5): 735-738, 2020 Jul 03.
Artigo em Inglês | MEDLINE | ID: mdl-32589459

RESUMO

PURPOSE: The spike proteins of SARS-CoV-2 interact with ACE2 or basigin/CD147 receptors, regulating human-to-human transmissions of COVID-19 together with serine protease TMPRSS2. The expression of these receptors on the ocular surface is unknown. MATERIAL AND METHODS: Gene expression of SARS-CoV-2 receptors was investigated in conjunctival epithelial cell samples and in ex-vivo cornea samples using microarray or transcriptome sequencing. RESULTS: ACE2 is expressed in conjunctival samples at a low level, while BSG and TMPRSS2 are expressed at intermediate levels in both conjunctiva and cornea. Other receptors such as ANPEP, AGTR2 are expressed at low level in the conjunctiva. Two RNA editing enzymes involved in antiviral responses, APOBEC3A, and ADAR-1 were also highly expressed. CONCLUSIONS: The ocular surface may represent an entry point for the SARS-CoV-2 in the human body. The conjunctiva and the cornea can adopt antiviral countermeasures which may explain the low prevalence of eye involvement.


Assuntos
Betacoronavirus/fisiologia , Túnica Conjuntiva/metabolismo , Córnea/metabolismo , Infecções por Coronavirus/metabolismo , Regulação da Expressão Gênica/fisiologia , Pneumonia Viral/metabolismo , Receptores Virais/genética , Adenosina Desaminase/genética , Adolescente , Adulto , Idoso , Basigina/genética , Criança , Citidina Desaminase/genética , Perfilação da Expressão Gênica , Humanos , Pessoa de Meia-Idade , Pandemias , Peptidil Dipeptidase A/genética , Proteínas/genética , Proteínas de Ligação a RNA/genética , Serina Endopeptidases/genética , Adulto Jovem
15.
Nucleic Acids Res ; 48(11): 5849-5858, 2020 06 19.
Artigo em Inglês | MEDLINE | ID: mdl-32383740

RESUMO

Adenosine-to-inosine (A-to-I) RNA editing is a common post transcriptional modification. It has a critical role in protecting against false activation of innate immunity by endogenous double stranded RNAs and has been associated with various regulatory processes and diseases such as autoimmune and cardiovascular diseases as well as cancer. In addition, the endogenous A-to-I editing machinery has been recently harnessed for RNA engineering. The study of RNA editing in humans relies heavily on the usage of cell lines as an important and commonly-used research tool. In particular, manipulations of the editing enzymes and their targets are often developed using cell line platforms. However, RNA editing in cell lines behaves very differently than in normal and diseased tissues, and most cell lines exhibit low editing levels, requiring over-expression of the enzymes. Here, we explore the A-to-I RNA editing landscape across over 1000 human cell lines types and show that for almost every editing target of interest a suitable cell line that mimics normal tissue condition may be found. We provide CLAIRE, a searchable catalogue of RNA editing levels across cell lines available at http://srv00.recas.ba.infn.it/atlas/claire.html, to facilitate rational choice of appropriate cell lines for future work on A-to-I RNA editing.


Assuntos
Linhagem Celular Tumoral , Edição de RNA , Adenosina Desaminase/genética , Sequência de Bases , Proteínas de Transporte/genética , Estudos de Casos e Controles , Células HEK293 , Humanos , Especificidade de Órgãos , Proteínas de Ligação a RNA/genética , Reprodutibilidade dos Testes
16.
Nat Biotechnol ; 38(7): 883-891, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32433547

RESUMO

Applications of adenine base editors (ABEs) have been constrained by the limited compatibility of the deoxyadenosine deaminase component with Cas homologs other than SpCas9. We evolved the deaminase component of ABE7.10 using phage-assisted non-continuous and continuous evolution (PANCE and PACE), which resulted in ABE8e. ABE8e contains eight additional mutations that increase activity (kapp) 590-fold compared with that of ABE7.10. ABE8e offers substantially improved editing efficiencies when paired with a variety of Cas9 or Cas12 homologs. ABE8e is more processive than ABE7.10, which could benefit screening, disruption of regulatory regions and multiplex base editing applications. A modest increase in Cas9-dependent and -independent DNA off-target editing, and in transcriptome-wide RNA off-target editing can be ameliorated by the introduction of an additional mutation in the TadA-8e domain. Finally, we show that ABE8e can efficiently install natural mutations that upregulate fetal hemoglobin expression in the BCL11A enhancer or in the the HBG promoter in human cells, targets that were poorly edited with ABE7.10. ABE8e augments the effectiveness and applicability of adenine base editing.


Assuntos
Adenina/metabolismo , Sistemas CRISPR-Cas/genética , DNA/genética , RNA/genética , Adenosina Desaminase/genética , Bacteriófagos/genética , Edição de Genes , Células HEK293 , Humanos , Mutagênese/genética , Mutação/genética
17.
Toxicol Lett ; 331: 22-32, 2020 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-32439581

RESUMO

Adenosine-to-inosine (A-to-I) RNA editing, catalyzed by adenosine deaminase acting on RNA (ADAR) enzymes, is the most frequent type of post-transcriptional nucleotide conversion in humans. It is known that innate abnormalities of A-to-I RNA editing are associated with the risk of certain diseases, such as amyotrophic lateral sclerosis. Extrinsic factors that modulate ADAR-mediated RNA editing remain to be clarified. In this study, we investigated the possibility that cigarette smoking may influence the expression of ADAR and that the changes may be biologically significant. Treatment of human lung adenocarcinoma A549 cells with cigarette smoke extract (CSE) induced a significant 50% decrease in ADAR1 protein levels. Since the decrease was counteracted by cotreatment with chloroquine, the CSE-dependent decrease in the ADAR1 protein levels may be due to the activation of autophagy. In addition to the in vitro study, we performed an in vivo study in mice and found a decrease in pulmonary Adar1 protein expression induced by cigarette smoking. Then, we investigated the biological significance of decreased ADAR1 expression. We found that knockdown of ADAR1 in A549 cells by siRNA resulted in an increase in the levels of protein carbonyl, a marker of oxidative stress. Moreover, knockdown of ADAR1 triggered a decrease in super oxide dismutase activity and heme oxygenase-1 expression, suggesting that ADAR1 plays a role to suppress oxidative stress. In conclusion, we show that ADAR1 expression is decreased by cigarette smoking and is a factor that contributes to the enhanced intracellular oxidative stress caused by cigarette smoking.


Assuntos
Adenosina Desaminase/genética , Estresse Oxidativo/efeitos dos fármacos , Edição de RNA/efeitos dos fármacos , Proteínas de Ligação a RNA/genética , Fumaça/efeitos adversos , Produtos do Tabaco , Células A549 , Técnicas de Cultura de Células , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/genética , Humanos , Estresse Oxidativo/genética
18.
Cancer Invest ; 38(6): 356-364, 2020 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-32468861

RESUMO

Pleural effusion adenosine deaminase (ADA) levels are elevated in various diseases. We investigated whether pleural effusion ADA levels differ among patients with malignant pleural mesothelioma (MPM), lung cancer (LC), and benign diseases, including tuberculous pleurisy. We examined 329 patients from February 2002 to July 2013. There were 131 MPM cases with ADA levels of 32.29 IU/L; 117 LC cases with ADA levels of 21.12 IU/L; 54 benign disease cases with ADA levels of 20.98 IU/L. A significant difference existed in pleural effusion ADA levels between MPM and benign disease patients. Pleural effusion ADA levels were significantly higher in MPM patients.


Assuntos
Adenosina Desaminase/genética , Neoplasias Pulmonares/diagnóstico , Mesotelioma/diagnóstico , Neoplasias/diagnóstico , Neoplasias Pleurais/diagnóstico , Adulto , Idoso , Idoso de 80 Anos ou mais , Feminino , Humanos , Neoplasias Pulmonares/diagnóstico por imagem , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patologia , Masculino , Mesotelioma/diagnóstico por imagem , Mesotelioma/genética , Mesotelioma/patologia , Pessoa de Meia-Idade , Mycobacterium tuberculosis/isolamento & purificação , Mycobacterium tuberculosis/patogenicidade , Neoplasias/diagnóstico por imagem , Neoplasias/genética , Neoplasias/patologia , Derrame Pleural Maligno/diagnóstico , Derrame Pleural Maligno/diagnóstico por imagem , Derrame Pleural Maligno/genética , Derrame Pleural Maligno/patologia , Neoplasias Pleurais/diagnóstico por imagem , Neoplasias Pleurais/genética , Neoplasias Pleurais/patologia , Toracoscopia , Tuberculose Pleural/diagnóstico , Tuberculose Pleural/genética , Tuberculose Pleural/microbiologia , Tuberculose Pleural/patologia
19.
Nat Commun ; 11(1): 2026, 2020 04 24.
Artigo em Inglês | MEDLINE | ID: mdl-32332729

RESUMO

The cell-context dependency for RNA binding proteins (RBPs) mediated control of stem cell fate remains to be defined. Here we adapt the HyperTRIBE method using an RBP fused to a Drosophila RNA editing enzyme (ADAR) to globally map the mRNA targets of the RBP MSI2 in mammalian adult normal and malignant stem cells. We reveal a unique MUSASHI-2 (MSI2) mRNA binding network in hematopoietic stem cells that changes during transition to multipotent progenitors. Additionally, we discover a significant increase in RNA binding activity of MSI2 in leukemic stem cells compared with normal hematopoietic stem and progenitor cells, resulting in selective regulation of MSI2's oncogenic targets. This provides a basis for MSI2 increased dependency in leukemia cells compared to normal cells. Moreover, our study provides a way to measure RBP function in rare cells and suggests that RBPs can achieve differential binding activity during cell state transition independent of gene expression.


Assuntos
Diferenciação Celular/genética , Células-Tronco Hematopoéticas/patologia , Leucemia/genética , Células-Tronco Neoplásicas/patologia , Proteínas de Ligação a RNA/metabolismo , Adenosina Desaminase/genética , Animais , Sítios de Ligação/genética , Modelos Animais de Doenças , Proteínas de Drosophila/genética , Regulação Leucêmica da Expressão Gênica , Redes Reguladoras de Genes , Células HEK293 , Humanos , Leucemia/sangue , Leucemia/patologia , Camundongos , Camundongos Knockout , Cultura Primária de Células , RNA Mensageiro/metabolismo , Proteínas de Ligação a RNA/genética , RNA-Seq , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo
20.
Iran J Allergy Asthma Immunol ; 19(1): 94-101, 2020 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-32245326

RESUMO

Severe combined immunodeficiency (SCID) comprises a heterogeneous group of genetic disorders caused by early defects in the development and function of T cells. Other lymphocyte lineages (B and/or natural killer cells) are variably affected. With a worldwide frequency of approximately 1:50,000 live births, SCID may result from diverse mutations in over 16 genes. Whole-exome sequencing (WES) provides an opportunity for parallel screening of all those genes. This approach is also useful for genetic diagnosis in parents whose infant expired before genetic testing. Here, we describe a heterozygous novel non-frameshift deletion (c.587_598del p.196_199del) in the adenosine deaminase (ADA) gene identified by WES in healthy parents of an expired child with SCID. The mutation was subsequently confirmed to be homozygous in the deceased baby whose left-over blood sample volume was insufficient for direct WES analysis. In conclusion, we here describe a novel mutation in ADA, a well-known SCID gene.


Assuntos
Adenosina Desaminase/deficiência , Adenosina Desaminase/genética , Agamaglobulinemia/genética , Imunodeficiência Combinada Severa/genética , Feminino , Humanos , Lactente , Irã (Geográfico) , Masculino , Mutação , Linhagem , Sequenciamento Completo do Exoma
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