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1.
Nat Commun ; 11(1): 5117, 2020 10 09.
Artigo em Inglês | MEDLINE | ID: mdl-33037203

RESUMO

Exposure of gastric epithelial cells to the bacterial carcinogen Helicobacter pylori causes DNA double strand breaks. Here, we show that H. pylori-induced DNA damage occurs co-transcriptionally in S-phase cells that activate NF-κB signaling upon innate immune recognition of the lipopolysaccharide biosynthetic intermediate ß-ADP-heptose by the ALPK1/TIFA signaling pathway. DNA damage depends on the bi-functional RfaE enzyme and the Cag pathogenicity island of H. pylori, is accompanied by replication fork stalling and can be observed also in primary cells derived from gastric organoids. Importantly, H. pylori-induced replication stress and DNA damage depend on the presence of co-transcriptional RNA/DNA hybrids (R-loops) that form in infected cells during S-phase as a consequence of ß-ADP-heptose/ ALPK1/TIFA/NF-κB signaling. H. pylori resides in close proximity to S-phase cells in the gastric mucosa of gastritis patients. Taken together, our results link bacterial infection and NF-κB-driven innate immune responses to R-loop-dependent replication stress and DNA damage.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Helicobacter pylori/patogenicidade , NF-kappa B/metabolismo , Proteínas Quinases/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas de Bactérias/metabolismo , Linhagem Celular Tumoral , DNA/química , DNA/genética , Dano ao DNA , Replicação do DNA/efeitos dos fármacos , Floxuridina , Glicosiltransferases/metabolismo , Infecções por Helicobacter/metabolismo , Infecções por Helicobacter/microbiologia , Helicobacter pylori/metabolismo , Interações Hospedeiro-Patógeno/fisiologia , Humanos , Lipopolissacarídeos/metabolismo , Mutação , NF-kappa B/genética , Proteínas Quinases/genética , Espécies Reativas de Oxigênio/metabolismo , Neoplasias Gástricas/genética , Neoplasias Gástricas/microbiologia , Neoplasias Gástricas/patologia
2.
Adv Genet ; 106: 1-4, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-33081918

RESUMO

This current volume is, in many ways, a 2020 update to the important 1999-2000 compendium by Sir Fred Hoyle and Professor N. Chandra Wickramasinghe's "Astronomical Origins of life: Steps towards Panspermia." The emerging new paradigm of biology that connects life on Earth with the wider cosmos is covered in considerable depth showing that terrestrial biological evolution is best understood as a cosmically derived habitat and an interconnected genetic system. The various chapters here discuss all aspects of this interconnectedness, particularly relevant now in this time of the coronavirus pandemic (COVID-19) as the human race reacts to the many microbes and viral pathogens that arrive regularly from space.


Assuntos
Evolução Biológica , Meio Ambiente , Exobiologia , Interações Hospedeiro-Patógeno/fisiologia , Betacoronavirus/fisiologia , Infecções por Coronavirus/epidemiologia , Infecções por Coronavirus/virologia , Doença/etiologia , Ecossistema , Meio Ambiente Extraterreno , Humanos , Modelos Biológicos , Origem da Vida , Pandemias , Pneumonia Viral/epidemiologia , Pneumonia Viral/virologia
3.
Adv Genet ; 106: 75-100, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-33081928

RESUMO

The origins and global spread of two recent, yet quite different, pandemic diseases is discussed and reviewed in depth: Candida auris, a eukaryotic fungal disease, and COVID-19 (SARS-CoV-2), a positive strand RNA viral respiratory disease. Both these diseases display highly distinctive patterns of sudden emergence and global spread, which are not easy to understand by conventional epidemiological analysis based on simple infection-driven human- to-human spread of an infectious disease (assumed to jump suddenly and thus genetically, from an animal reservoir). Both these enigmatic diseases make sense however under a Panspermia in-fall model and the evidence consistent with such a model is critically reviewed.


Assuntos
Evolução Biológica , Candidíase/etiologia , Doenças Transmissíveis Emergentes/etiologia , Infecções por Coronavirus/etiologia , Origem da Vida , Pneumonia Viral/etiologia , Animais , Betacoronavirus/isolamento & purificação , Betacoronavirus/fisiologia , Candida/isolamento & purificação , Candida/fisiologia , Candidíase/epidemiologia , Doenças Transmissíveis Emergentes/epidemiologia , Coronavirus/isolamento & purificação , Coronavirus/fisiologia , Infecções por Coronavirus/epidemiologia , Planeta Terra , Exobiologia , Meio Ambiente Extraterreno , Interações Hospedeiro-Patógeno/fisiologia , Humanos , Pandemias , Pneumonia Viral/epidemiologia
4.
Sci Rep ; 10(1): 16099, 2020 09 30.
Artigo em Inglês | MEDLINE | ID: mdl-32999356

RESUMO

SARS-CoV-2 is the cause of the ongoing COVID-19 pandemic. Here, we investigated the interaction of this new coronavirus with Vero cells using high resolution scanning electron microscopy. Surface morphology, the interior of infected cells and the distribution of viral particles in both environments were observed 2 and 48 h after infection. We showed areas of viral processing, details of vacuole contents, and viral interactions with the cell surface. Intercellular connections were also approached, and viral particles were adhered to these extensions suggesting direct cell-to-cell transmission of SARS-CoV-2.


Assuntos
Betacoronavirus/ultraestrutura , Infecções por Coronavirus/transmissão , Interações Hospedeiro-Patógeno/fisiologia , Pneumonia Viral/transmissão , Animais , Linhagem Celular , Chlorocebus aethiops , Infecções por Coronavirus/patologia , Humanos , Microscopia Eletrônica de Varredura , Pandemias , Pneumonia Viral/patologia , Células Vero
5.
Bioessays ; 42(11): e2000078, 2020 11.
Artigo em Inglês | MEDLINE | ID: mdl-32893352

RESUMO

Intermediate filaments (IFs) formed by vimentin are less understood than their cytoskeletal partners, microtubules and F-actin, but the unique physical properties of IFs, especially their resistance to large deformations, initially suggest a mechanical function. Indeed, vimentin IFs help regulate cell mechanics and contractility, and in crowded 3D environments they protect the nucleus during cell migration. Recently, a multitude of studies, often using genetic or proteomic screenings show that vimentin has many non-mechanical functions within and outside of cells. These include signaling roles in wound healing, lipogenesis, sterol processing, and various functions related to extracellular and cell surface vimentin. Extracellular vimentin is implicated in marking circulating tumor cells, promoting neural repair, and mediating the invasion of host cells by viruses, including SARS-CoV, or bacteria such as Listeria and Streptococcus. These findings underscore the fundamental role of vimentin in not only cell mechanics but also a range of physiological functions. Also see the video abstract here https://youtu.be/YPfoddqvz-g.


Assuntos
Filamentos Intermediários/fisiologia , Mecanotransdução Celular/fisiologia , Vimentina/fisiologia , Animais , Fenômenos Fisiológicos Bacterianos , Interações Hospedeiro-Patógeno/fisiologia , Humanos , Filamentos Intermediários/química , Fenômenos Mecânicos , Vírus da SARS/fisiologia , Vimentina/química , Internalização do Vírus
6.
PLoS One ; 15(9): e0238993, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32925983

RESUMO

The placental membranes that surround the fetus during pregnancy were suggested to contain a low abundance microbiota. Specifically, abundance of Lactobacillus, a probiotic and dominant member of the microbiome of the lower reproductive tract, has been shown to correlate with healthy, term pregnancies. We therefore sought to assess the interactions between four different Lactobacillus strains with immortalized decidualized endometrial cells (dT-HESCs), which were used as a model to represent the outermost layer of the placental membranes. Notably, we demonstrated that all four Lactobacillus strains could associate with dT-HESCs in vitro. L. crispatus was significantly more successful (p < 0.00005), with 10.6% of bacteria attaching to the host cells compared to an average of 0.8% for the remaining three strains. The four strains also varied in their ability to form biofilms. Dependent on media type, L. reuteri 6475 formed the strongest biofilms in vitro. To examine the impact on immune responses, levels of total and phosphorylated protein p38, a member of the Mitogen Activated Protein Kinase (MAPK) pathway, were examined following Lactobacillus association with dT-HESCs. Total levels of p38 were reduced to an average of 44% that of the cells without Lactobacillus (p < 0.05). While a trend towards a reduction in phosphorylated p38 was observed, this difference was not significant (p > 0.05). In addition, association with Lactobacillus did not result in increased host cell death. Collectively, these data suggest that varying types of Lactobacillus can attach to the outermost cells of the placental membranes and that these interactions do not contribute to inflammatory responses or host cell death. To our knowledge this is the first in vitro study to support the ability of Lactobacillus to interact with placental cells, which is important when considering its use as a potential probiotic within the reproductive tract.


Assuntos
Interações Hospedeiro-Patógeno/fisiologia , Lactobacillus/metabolismo , Placenta/microbiologia , Biofilmes , Linhagem Celular Tumoral , Endométrio/metabolismo , Endométrio/microbiologia , Células Epiteliais/metabolismo , Células Epiteliais/microbiologia , Feminino , Humanos , Lactobacillus/patogenicidade , Microbiota , Modelos Biológicos , Gravidez , Células Estromais/metabolismo , Células Estromais/microbiologia
7.
Nat Struct Mol Biol ; 27(10): 959-966, 2020 10.
Artigo em Inglês | MEDLINE | ID: mdl-32908316

RESUMO

The SARS-CoV-2 non-structural protein 1 (Nsp1), also referred to as the host shutoff factor, suppresses host innate immune functions. By combining cryo-electron microscopy and biochemistry, we show that SARS-CoV-2 Nsp1 binds to the human 40S subunit in ribosomal complexes, including the 43S pre-initiation complex and the non-translating 80S ribosome. The protein inserts its C-terminal domain into the mRNA channel, where it interferes with mRNA binding. We observe translation inhibition in the presence of Nsp1 in an in vitro translation system and in human cells. Based on the high-resolution structure of the 40S-Nsp1 complex, we identify residues of Nsp1 crucial for mediating translation inhibition. We further show that the full-length 5' untranslated region of the genomic viral mRNA stimulates translation in vitro, suggesting that SARS-CoV-2 combines global inhibition of translation by Nsp1 with efficient translation of the viral mRNA to allow expression of viral genes.


Assuntos
Betacoronavirus/química , Betacoronavirus/metabolismo , Biossíntese de Proteínas , RNA Mensageiro/genética , Proteínas não Estruturais Virais/química , Proteínas não Estruturais Virais/metabolismo , Regiões 5' não Traduzidas , Betacoronavirus/genética , Microscopia Crioeletrônica , Células HEK293 , Células HeLa , Interações Hospedeiro-Patógeno/fisiologia , Humanos , Modelos Moleculares , Mutação , Conformação Proteica , Domínios Proteicos , RNA Mensageiro/metabolismo , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Subunidades Ribossômicas Menores de Eucariotos/genética , Subunidades Ribossômicas Menores de Eucariotos/metabolismo , Proteínas não Estruturais Virais/genética
8.
Nat Commun ; 11(1): 4775, 2020 09 22.
Artigo em Inglês | MEDLINE | ID: mdl-32963221

RESUMO

Enterovirus 71 (EV71) poses serious threats to human health, particularly in Southeast Asia, and no drugs or vaccines are available. Previous work identified the stem loop II structure of the EV71 internal ribosomal entry site as vital to viral translation and a potential target. After screening an RNA-biased library using a peptide-displacement assay, we identify DMA-135 as a dose-dependent inhibitor of viral translation and replication with no significant toxicity in cell-based studies. Structural, biophysical, and biochemical characterization support an allosteric mechanism in which DMA-135 induces a conformational change in the RNA structure that stabilizes a ternary complex with the AUF1 protein, thus repressing translation. This mechanism is supported by pull-down experiments in cell culture. These detailed studies establish enterovirus RNA structures as promising drug targets while revealing an approach and mechanism of action that should be broadly applicable to functional RNA targeting.


Assuntos
Enterovirus Humano A/genética , Enterovirus Humano A/fisiologia , Infecções por Enterovirus/metabolismo , Interações Hospedeiro-Patógeno/fisiologia , Sítios Internos de Entrada Ribossomal/fisiologia , Replicação Viral/fisiologia , Regiões 5' não Traduzidas , Linhagem Celular , Infecções por Enterovirus/virologia , Regulação Viral da Expressão Gênica , Ribonucleoproteína Nuclear Heterogênea D0/metabolismo , Humanos , Ligantes , Modelos Moleculares , Ligação Proteica , RNA Viral/química , Proteínas Virais/metabolismo
9.
PLoS Comput Biol ; 16(9): e1007470, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32941445

RESUMO

Human T-lymphotropic virus type-1 (HTLV-1) persists within hosts via infectious spread (de novo infection) and mitotic spread (infected cell proliferation), creating a population structure of multiple clones (infected cell populations with identical genomic proviral integration sites). The relative contributions of infectious and mitotic spread to HTLV-1 persistence are unknown, and will determine the efficacy of different approaches to treatment. The prevailing view is that infectious spread is negligible in HTLV-1 persistence beyond early infection. However, in light of recent high-throughput data on the abundance of HTLV-1 clones, and recent estimates of HTLV-1 clonal diversity that are substantially higher than previously thought (typically between 104 and 105 HTLV-1+ T cell clones in the body of an asymptomatic carrier or patient with HTLV-1-associated myelopathy/tropical spastic paraparesis), ongoing infectious spread during chronic infection remains possible. We estimate the ratio of infectious to mitotic spread using a hybrid model of deterministic and stochastic processes, fitted to previously published HTLV-1 clonal diversity estimates. We investigate the robustness of our estimates using three alternative estimators. We find that, contrary to previous belief, infectious spread persists during chronic infection, even after HTLV-1 proviral load has reached its set point, and we estimate that between 100 and 200 new HTLV-1 clones are created and killed every day. We find broad agreement between all estimators. The risk of HTLV-1-associated malignancy and inflammatory disease is strongly correlated with proviral load, which in turn is correlated with the number of HTLV-1-infected clones, which are created by de novo infection. Our results therefore imply that suppression of de novo infection may reduce the risk of malignant transformation.


Assuntos
Infecções por HTLV-I , Interações Hospedeiro-Patógeno , Vírus Linfotrópico T Tipo 1 Humano , Linfócitos T CD4-Positivos/virologia , Infecções por HTLV-I/fisiopatologia , Infecções por HTLV-I/virologia , Interações Hospedeiro-Patógeno/genética , Interações Hospedeiro-Patógeno/fisiologia , Vírus Linfotrópico T Tipo 1 Humano/classificação , Vírus Linfotrópico T Tipo 1 Humano/genética , Vírus Linfotrópico T Tipo 1 Humano/patogenicidade , Humanos , Mitose/genética , Mitose/fisiologia , Modelos Biológicos , Provírus/genética , Provírus/patogenicidade , Carga Viral/genética , Integração Viral/genética
10.
PLoS Pathog ; 16(9): e1008927, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32997711

RESUMO

Viruses cleave cellular proteins to remodel the host proteome. The study of these cleavages has revealed mechanisms of immune evasion, resource exploitation, and pathogenesis. However, the full extent of virus-induced proteolysis in infected cells is unknown, mainly because until recently the technology for a global view of proteolysis within cells was lacking. Here, we report the first comprehensive catalog of proteins cleaved upon enterovirus infection and identify the sites within proteins where the cleavages occur. We employed multiple strategies to confirm protein cleavages and assigned them to one of the two enteroviral proteases. Detailed characterization of one substrate, LSM14A, a p body protein with a role in antiviral immunity, showed that cleavage of this protein disrupts its antiviral function. This study yields a new depth of information about the host interface with a group of viruses that are both important biological tools and significant agents of disease.


Assuntos
Cisteína Endopeptidases/metabolismo , Infecções por Enterovirus/virologia , Enterovirus/patogenicidade , Replicação Viral/fisiologia , Antivirais/metabolismo , Enterovirus/metabolismo , Interações Hospedeiro-Patógeno/fisiologia , Humanos , Proteólise , Proteínas Virais/metabolismo
11.
Nat Commun ; 11(1): 3896, 2020 08 04.
Artigo em Inglês | MEDLINE | ID: mdl-32753727

RESUMO

The outbreak of Zika virus (ZIKV) in 2016 created worldwide health emergency which demand urgent research efforts on understanding the virus biology and developing therapeutic strategies. Here, we present a time-resolved chemical proteomic strategy to track the early-stage entry of ZIKV into host cells. ZIKV was labeled on its surface with a chemical probe, which carries a photocrosslinker to covalently link virus-interacting proteins in living cells on UV exposure at different time points, and a biotin tag for subsequent enrichment and mass spectrometric identification of the receptor or other host proteins critical for virus internalization. We identified Neural Cell Adhesion Molecule (NCAM1) as a potential ZIKV receptor and further validated it through overexpression, knockout, and inhibition of NCAM1 in Vero cells and human glioblastoma cells U-251 MG. Collectively, the strategy can serve as a universal tool to map virus entry pathways and uncover key interacting proteins.


Assuntos
Moléculas de Adesão de Célula Nervosa/metabolismo , Proteômica , Receptores Virais/metabolismo , Internalização do Vírus , Replicação Viral/fisiologia , Zika virus/fisiologia , Animais , Antígeno CD56/genética , Antígeno CD56/metabolismo , Linhagem Celular Tumoral , Chlorocebus aethiops , Técnicas de Inativação de Genes , Glioblastoma , Células HEK293 , Interações Hospedeiro-Patógeno/fisiologia , Humanos , Moléculas de Adesão de Célula Nervosa/genética , Células Vero , Proteínas Virais/metabolismo , Ligação Viral , Infecção por Zika virus/virologia
12.
Methods Mol Biol ; 2203: 187-204, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32833213

RESUMO

Biotin-based proximity labeling circumvents major pitfalls of classical biochemical approaches to identify protein-protein interactions. It consists of enzyme-catalyzed biotin tags ubiquitously apposed on proteins located in close proximity of the labeling enzyme, followed by affinity purification and identification of biotinylated proteins by mass spectrometry. Here we outline the methods by which the molecular microenvironment of the coronavirus replicase/transcriptase complex (RTC), i.e., proteins located within a close perimeter of the RTC, can be determined by different proximity labeling approaches using BirAR118G (BioID), TurboID, and APEX2. These factors represent a molecular signature of coronavirus RTCs and likely contribute to the viral life cycle, thereby constituting attractive targets for the development of antiviral intervention strategies.


Assuntos
Coronavirus/patogenicidade , Enzimas/genética , Interações Hospedeiro-Patógeno/fisiologia , Proteômica/métodos , Proteínas Virais/metabolismo , Animais , Ascorbato Peroxidases/genética , Biotinilação , Carbono-Nitrogênio Ligases/genética , Linhagem Celular , Coronavirus/genética , Enzimas/metabolismo , Proteínas de Escherichia coli/genética , Imunofluorescência , Microrganismos Geneticamente Modificados , Proteínas Repressoras/genética , Proteínas Virais/química , Proteínas Virais/genética
13.
Methods Mol Biol ; 2203: 205-221, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32833214

RESUMO

We have developed a screening system using the yeast Saccharomyces cerevisiae to identify eukaryotic genes involved in the replication of mammalian viruses. Yeast come with various advantages, but in the context of coronavirus research and the system outlined here, they are simple and easy to work with and can be used at biosafety level 2. The system involves inducible expression of individual viral proteins and identification of detrimental phenotypes in the yeast. Yeast knockout and overexpression libraries can then be used for genome-wide screening of host proteins that provide a suppressor phenotype. From the yeast hits, a narrowed list of candidate genes can be produced to investigate for roles in viral replication. Since the system only requires expression of viral proteins, it can be used for any current or emerging virus, regardless of biocontainment requirements and ability to culture the virus. In this chapter, we will outline the protocols that can be used to take advantage of S. cerevisiae as a tool to advance understanding of how viruses interact with eukaryotic cells.


Assuntos
Coronavirus/fisiologia , Coronavirus/patogenicidade , Interações Hospedeiro-Patógeno/fisiologia , Saccharomyces cerevisiae/genética , Plasmídeos , Proteínas Virais/genética , Proteínas Virais/isolamento & purificação , Replicação Viral
14.
Methods Mol Biol ; 2203: 241-261, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32833217

RESUMO

Coronavirus entry encompasses the initial steps of infection, from virion attachment to genome release. Advances in fluorescent labeling of viral and cellular components and confocal imaging enable broad spectrum studies on this process. Here, we describe methods for visualization of coronavirus entry into immortalized cell lines and 3D tissue culture models.


Assuntos
Coronavirus/patogenicidade , Interações Hospedeiro-Patógeno/fisiologia , Microscopia Confocal/métodos , Linhagem Celular , Coronavirus/isolamento & purificação , Meios de Cultura/química , Endocitose , Humanos , Proteínas do Nucleocapsídeo/metabolismo , Ácidos Tri-Iodobenzoicos/química , Internalização do Vírus
15.
Nat Commun ; 11(1): 4332, 2020 08 28.
Artigo em Inglês | MEDLINE | ID: mdl-32859902

RESUMO

The group of enteroviruses contains many important pathogens for humans, including poliovirus, coxsackievirus, rhinovirus, as well as newly emerging global health threats such as EV-A71 and EV-D68. Here, we describe an unbiased, system-wide and time-resolved analysis of the proteome and phosphoproteome of human cells infected with coxsackievirus B3. Of the ~3,200 proteins quantified throughout the time course, a large amount (~25%) shows a significant change, with the majority being downregulated. We find ~85% of the detected phosphosites to be significantly regulated, implying that most changes occur at the post-translational level. Kinase-motif analysis reveals temporal activation patterns of certain protein kinases, with several CDKs/MAPKs immediately active upon the infection, and basophilic kinases, ATM, and ATR engaging later. Through bioinformatics analysis and dedicated experiments, we identify mTORC1 signalling as a major regulation network during enterovirus infection. We demonstrate that inhibition of mTORC1 activates TFEB, which increases expression of lysosomal and autophagosomal genes, and that TFEB activation facilitates the release of virions in extracellular vesicles via secretory autophagy. Our study provides a rich framework for a system-level understanding of enterovirus-induced perturbations at the protein and signalling pathway levels, forming a base for the development of pharmacological inhibitors to treat enterovirus infections.


Assuntos
Infecções por Coxsackievirus/metabolismo , Interações Hospedeiro-Patógeno/fisiologia , Proteoma/análise , Animais , Autofagia , Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos/genética , Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos/metabolismo , Linhagem Celular , Sobrevivência Celular , Enterovirus/fisiologia , Enterovirus Humano B/fisiologia , Técnicas de Inativação de Genes , Células HeLa , Humanos , Alvo Mecanístico do Complexo 1 de Rapamicina , Fosforilação , Transdução de Sinais , Proteínas Virais/metabolismo
16.
J Transl Med ; 18(1): 319, 2020 08 18.
Artigo em Inglês | MEDLINE | ID: mdl-32811513

RESUMO

In less than 20 years, three deadly coronaviruses, SARS-CoV, MERS-CoV and SARS-CoV-2, have emerged in human population causing hundreds to hundreds of thousands of deaths. Other coronaviruses are causing epizootic representing a significant threat for both domestic and wild animals. Members of this viral family have the longest genome of all RNA viruses, and express up to 29 proteins establishing complex interactions with the host proteome. Deciphering these interactions is essential to identify cellular pathways hijacked by these viruses to replicate and escape innate immunity. Virus-host interactions also provide key information to select targets for antiviral drug development. Here, we have manually curated the literature to assemble a unique dataset of 1311 coronavirus-host protein-protein interactions. Functional enrichment and network-based analyses showed coronavirus connections to RNA processing and translation, DNA damage and pathogen sensing, interferon production, and metabolic pathways. In particular, this global analysis pinpointed overlooked interactions with translation modulators (GIGYF2-EIF4E2), components of the nuclear pore, proteins involved in mitochondria homeostasis (PHB, PHB2, STOML2), and methylation pathways (MAT2A/B). Finally, interactome data provided a rational for the antiviral activity of some drugs inhibiting coronaviruses replication. Altogether, this work describing the current landscape of coronavirus-host interactions provides valuable hints for understanding the pathophysiology of coronavirus infections and developing effective antiviral therapies.


Assuntos
Infecções por Coronavirus/metabolismo , Coronavirus/metabolismo , Interações Hospedeiro-Patógeno/fisiologia , Mapas de Interação de Proteínas , Proteínas Virais/metabolismo , Animais , Betacoronavirus/fisiologia , Coronavirus/química , Infecções por Coronavirus/virologia , Bases de Dados de Proteínas , Humanos , Proteínas Mitocondriais/metabolismo , Pandemias , Pneumonia Viral/metabolismo , Pneumonia Viral/virologia , Fatores de Transcrição/metabolismo , Replicação Viral/genética
17.
Biomolecules ; 10(8)2020 08 01.
Artigo em Inglês | MEDLINE | ID: mdl-32752270

RESUMO

Posttranslational modifications of cellular proteins by covalent conjugation of ubiquitin and ubiquitin-like polypeptides regulate numerous cellular processes that are captured by viruses to promote infection, replication, and spreading. The importance of these protein modifications for the viral life cycle is underscored by the discovery that many viruses encode deconjugases that reverse their functions. The structural and functional characterization of these viral enzymes and the identification of their viral and cellular substrates is providing valuable insights into the biology of viral infections and the host's antiviral defense. Given the growing body of evidence demonstrating their key contribution to pathogenesis, the viral deconjugases are now recognized as attractive targets for the design of novel antiviral therapeutics.


Assuntos
Antivirais/farmacologia , Enzimas/metabolismo , Interações Hospedeiro-Patógeno/fisiologia , Ubiquitina/metabolismo , Proteínas Virais/metabolismo , Viroses/metabolismo , Adenoviridae/enzimologia , Coronavirus/enzimologia , Enzimas/química , Herpesviridae/enzimologia , Humanos , Processamento de Proteína Pós-Traducional , Proteínas Virais/química , Viroses/tratamento farmacológico
18.
Genes (Basel) ; 11(8)2020 08 10.
Artigo em Inglês | MEDLINE | ID: mdl-32785186

RESUMO

The coronaviruses are a large family of enveloped RNA viruses that commonly cause gastrointestinal or respiratory illnesses in the infected host. Avian coronavirus infectious bronchitis virus (IBV) is a highly contagious respiratory pathogen of chickens that can affect the kidneys and reproductive systems resulting in bird mortality and decreased reproductivity. The interferon-inducible transmembrane (IFITM) proteins are activated in response to viral infections and represent a class of cellular restriction factors that restrict the replication of many viral pathogens. Here, we characterize the relative mRNA expression of the chicken IFITM genes in response to IBV infection, in vivo, ex vivo and in vitro using the pathogenic M41-CK strain, the nephropathogenic QX strain and the nonpathogenic Beaudette strain. In vivo we demonstrate a significant upregulation of chIFITM1, 2, 3 and 5 in M41-CK- and QX-infected trachea two days post-infection. In vitro infection with Beaudette, M41-CK and QX results in a significant upregulation of chIFITM1, 2 and 3 at 24 h post-infection. We confirmed a differential innate response following infection with distinct IBV strains and believe that our data provide new insights into the possible role of chIFITMs in early IBV infection.


Assuntos
Galinhas/genética , Galinhas/virologia , Infecções por Coronavirus/veterinária , Interações Hospedeiro-Patógeno/genética , Proteínas de Membrana/genética , Animais , Infecções por Coronavirus/genética , Regulação Viral da Expressão Gênica , Interações Hospedeiro-Patógeno/fisiologia , Vírus da Bronquite Infecciosa/patogenicidade , Vírus da Bronquite Infecciosa/fisiologia , Técnicas de Cultura de Órgãos , Doenças das Aves Domésticas/etiologia , Doenças das Aves Domésticas/genética , Doenças das Aves Domésticas/virologia , Carga Viral , Tropismo Viral
19.
Proc Natl Acad Sci U S A ; 117(36): 22462-22472, 2020 09 08.
Artigo em Inglês | MEDLINE | ID: mdl-32839311

RESUMO

Huntingtin-interacting protein family members are evolutionarily conserved from yeast to humans, and they are known to be key factors in clathrin-mediated endocytosis. Here we identified the Caenorhabditis elegans protein huntingtin-interacting protein-related 1 (HIPR-1) as a host factor essential for Orsay virus infection of C. elegans Ablation of HIPR-1 resulted in a greater than 10,000-fold reduction in viral RNA, which could be rescued by ectopic expression of HIPR-1. Viral RNA replication from an endogenous transgene replicon system was not affected by lack of HIPR-1, suggesting that HIPR-1 plays a role during an early, prereplication virus life-cycle stage. Ectopic expression of HIPR-1 mutants demonstrated that neither the clathrin light chain-binding domain nor the clathrin heavy chain-binding motif were needed for virus infection, whereas the inositol phospholipid-binding and F-actin-binding domains were essential. In human cell culture, deletion of the human HIP orthologs HIP1 and HIP1R led to decreased infection by Coxsackie B3 virus. Finally, ectopic expression of a chimeric HIPR-1 harboring the human HIP1 ANTH (AP180 N-terminal homology) domain rescued Orsay infection in C. elegans, demonstrating conservation of its function through evolution. Collectively, these findings further our knowledge of cellular factors impacting viral infection in C. elegans and humans.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Proteínas de Caenorhabditis elegans/metabolismo , Proteínas de Ligação a DNA/metabolismo , Interações Hospedeiro-Patógeno , Proteínas dos Microfilamentos/metabolismo , Células A549 , Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas Adaptadoras de Transdução de Sinal/fisiologia , Animais , Caenorhabditis elegans/metabolismo , Caenorhabditis elegans/virologia , Proteínas de Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/fisiologia , Sequência Conservada/genética , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/fisiologia , Enterovirus Humano B/patogenicidade , Enterovirus Humano B/fisiologia , Feminino , Técnicas de Silenciamento de Genes , Interações Hospedeiro-Patógeno/genética , Interações Hospedeiro-Patógeno/fisiologia , Humanos , Masculino , Proteínas dos Microfilamentos/genética , Proteínas dos Microfilamentos/fisiologia , Nodaviridae/patogenicidade , Nodaviridae/fisiologia , Domínios Proteicos/genética , Replicação Viral
20.
Plant Mol Biol ; 104(4-5): 381-395, 2020 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-32803478

RESUMO

KEY MESSAGE: Host mediated silencing of COM1 gene of Colletotrichum gloeosporioides disables appressorial differentiation and effectively prevents the development of Anthracnose disease in chilli and tomato. Anthracnose disease is caused by the ascomycetes fungal species Colletotrichum, which is responsible for heavy yield losses in chilli and tomato worldwide. Conventionally, harmful pesticides are used to contain anthracnose disease with limited success. In this study, we assessed the potential of Host-Induced Gene Silencing (HIGS) approach to target the Colletotrichum gloeosporioides COM1 (CgCOM1) developmental gene involved in the fungal conidial and appressorium formation, to restrict fungal infection in chilli and tomato fruits. For this study, we have developed stable transgenic lines of chilli and tomato expressing CgCOM1-RNAi construct employing Agrobacterium-mediated transformation. Transgenic plants were characterized by molecular and gene expression analyses. Production of specific CgCOM1 siRNA in transgenic chilli and tomato RNAi lines was confirmed by stem-loop RT-PCR. Fungal challenge assays on leaves and fruits showed that the transgenic lines were resistant to anthracnose disease-causing C. gloeosporioides in comparison to wild type and empty-vector control plants. RT-qPCR analyses in transgenic lines revealed extremely low abundance of CgCOM1 transcripts in the C. gloeosporioides infected tissues, indicating near complete silencing of CgCOM1 gene expression in the pathogen. Microscopic examination of the Cg-challenged leaves of chilli-CgCOM1i lines revealed highly suppressed conidial germination, germ tube development, appressoria formation and mycelial growth of C. gloeosporioides, resulting in reduced infection of plant tissues. These results demonstrated highly efficient use of HIGS in silencing the expression of essential fungal developmental genes to inhibit the growth of pathogenic fungi, thus providing a highly precise approach to arrest the spread of disease.


Assuntos
Capsicum/microbiologia , Colletotrichum/genética , Colletotrichum/patogenicidade , Interações Hospedeiro-Patógeno/fisiologia , Lycopersicon esculentum/microbiologia , Capsicum/genética , Resistência à Doença , Frutas/genética , Frutas/microbiologia , Regulação Fúngica da Expressão Gênica , Inativação Gênica , Genes Fúngicos , Lycopersicon esculentum/genética , Filogenia , Doenças das Plantas/microbiologia , Folhas de Planta/genética , Plantas Geneticamente Modificadas , Interferência de RNA , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Esporos Fúngicos
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