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1.
Viruses ; 13(7)2021 07 12.
Artigo em Inglês | MEDLINE | ID: mdl-34372555

RESUMO

Viroplasms are cytoplasmic, membraneless structures assembled in rotavirus (RV)-infected cells, which are intricately involved in viral replication. Two virus-encoded, non-structural proteins, NSP2 and NSP5, are the main drivers of viroplasm formation. The structures (as far as is known) and functions of these proteins are described. Recent studies using plasmid-only-based reverse genetics have significantly contributed to elucidation of the crucial roles of these proteins in RV replication. Thus, it has been recognized that viroplasms resemble liquid-like protein-RNA condensates that may be formed via liquid-liquid phase separation (LLPS) of NSP2 and NSP5 at the early stages of infection. Interactions between the RNA chaperone NSP2 and the multivalent, intrinsically disordered protein NSP5 result in their condensation (protein droplet formation), which plays a central role in viroplasm assembly. These droplets may provide a unique molecular environment for the establishment of inter-molecular contacts between the RV (+)ssRNA transcripts, followed by their assortment and equimolar packaging. Future efforts to improve our understanding of RV replication and genome assortment in viroplasms should focus on their complex molecular composition, which changes dynamically throughout the RV replication cycle, to support distinct stages of virion assembly.


Assuntos
Rotavirus/genética , Rotavirus/metabolismo , Compartimentos de Replicação Viral/metabolismo , Animais , Proteínas do Capsídeo/genética , Citoplasma/virologia , Citosol/metabolismo , Humanos , Fosforilação , Proteínas de Ligação a RNA/metabolismo , Infecções por Rotavirus/virologia , Proteínas não Estruturais Virais/metabolismo , Compartimentos de Replicação Viral/fisiologia , Montagem de Vírus , Replicação Viral/genética
2.
Front Immunol ; 12: 680891, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34194434

RESUMO

The network of tunneling nanotubes (TNTs) represents the filamentous (F)-actin rich tubular structure which is connected to the cytoplasm of the adjacent and or distant cells to mediate efficient cell-to-cell communication. They are long cytoplasmic bridges with an extraordinary ability to perform diverse array of function ranging from maintaining cellular physiology and cell survival to promoting immune surveillance. Ironically, TNTs are now widely documented to promote the spread of various pathogens including viruses either during early or late phase of their lifecycle. In addition, TNTs have also been associated with multiple pathologies in a complex multicellular environment. While the recent work from multiple laboratories has elucidated the role of TNTs in cellular communication and maintenance of homeostasis, this review focuses on their exploitation by the diverse group of viruses such as retroviruses, herpesviruses, influenza A, human metapneumovirus and SARS CoV-2 to promote viral entry, virus trafficking and cell-to-cell spread. The later process may aggravate disease severity and the associated complications due to widespread dissemination of the viruses to multiple organ system as observed in current coronavirus disease 2019 (COVID-19) patients. In addition, the TNT-mediated intracellular spread can be protective to the viruses from the circulating immune surveillance and possible neutralization activity present in the extracellular matrix. This review further highlights the relevance of TNTs in ocular and cardiac tissues including neurodegenerative diseases, chemotherapeutic resistance, and cancer pathogenesis. Taken together, we suggest that effective therapies should consider precise targeting of TNTs in several diseases including virus infections.


Assuntos
COVID-19/etiologia , Citoplasma/ultraestrutura , Citoplasma/virologia , Nanotubos/virologia , Doenças Neurodegenerativas/etiologia , Viroses/etiologia , Animais , COVID-19/virologia , Comunicação Celular , Humanos
3.
Arch Virol ; 166(7): 1921-1930, 2021 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-33905022

RESUMO

Positive-stranded RNA viruses usually remodel the host endomembrane system to form virus-induced intracellular vesicles for replication during infections. The genus Potyvirus of the family Potyviridae represents the largest number of positive single-stranded RNA viruses, and its members cause great damage to crop production worldwide. Although potyviruses have a wide host range, each potyvirus infects a relatively limited number of host species. Phylogenesis and host range analysis can divide potyviruses into monocot-infecting and dicot-infecting groups, suggesting that they differ in their infection mechanisms, probably during replication. Comprehensive studies on the model dicot-infecting turnip mosaic virus have shown that the 6K2-induced replication vesicles are derived from the endoplasmic reticulum (ER) and subsequently target chloroplasts for viral genome replication. However, the replication site of monocot-infecting potyviruses is unknown. In this study, we show that the precursor 6K2-VPg-Pro polyproteins of dicot-infecting potyviruses and monocot-infecting potyviruses cluster phylogenetically in two separate groups. With a typical gramineae-infecting potyvirus-sugarcane mosaic virus (SCMV)-we found that replicative double-stranded RNA (dsRNA) forms aggregates in the cytoplasm but does not associate with chloroplasts. SCMV 6K2-VPg-Pro-induced vesicles colocalize with replicative dsRNA. Moreover, SCMV 6K2-VPg-Pro-induced structures target multiple intracellular organelles, including the ER, Golgi apparatus, mitochondria, and peroxisomes, and have no evident association with chloroplasts.


Assuntos
Potyvirus/genética , RNA Viral/genética , Replicação Viral/genética , Brassica napus/virologia , Cloroplastos/virologia , Produtos Agrícolas/virologia , Citoplasma/virologia , Retículo Endoplasmático/virologia , Genômica/métodos , Interações Hospedeiro-Patógeno/genética , Doenças das Plantas/virologia , Poaceae/virologia , Proteínas Virais/genética
4.
Cell Microbiol ; 23(8): e13328, 2021 08.
Artigo em Inglês | MEDLINE | ID: mdl-33740320

RESUMO

Annulate lamellae (AL) have been observed many times over the years on electron micrographs of rapidly dividing cells, but little is known about these unusual organelles consisting of stacked sheets of endoplasmic reticulum-derived membranes with nuclear pore complexes (NPCs). Evidence is growing for a role of AL in viral infection. AL have been observed early in the life cycles of the hepatitis C virus (HCV) and, more recently, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), suggesting a specific induction of mechanisms potentially useful to these pathogens. Like other positive-strand RNA viruses, these viruses induce host cells membranes rearrangements. The NPCs of AL could potentially mediate exchanges between these partially sealed compartments and the cytoplasm. AL may also be involved in regulating Ca2+ homeostasis or cell cycle control. They were recently observed in cells infected with Theileria annulata, an intracellular protozoan parasite inducing cell proliferation. Further studies are required to clarify their role in intracellular pathogen/host-cell interactions.


Assuntos
Interações Hospedeiro-Patógeno/fisiologia , Organelas/microbiologia , Organelas/parasitologia , Animais , COVID-19 , Citoplasma/virologia , Retículo Endoplasmático/microbiologia , Retículo Endoplasmático/parasitologia , Retículo Endoplasmático/ultraestrutura , Retículo Endoplasmático/virologia , Humanos , Organelas/ultraestrutura , Organelas/virologia , SARS-CoV-2/fisiologia
5.
mBio ; 12(1)2021 02 09.
Artigo em Inglês | MEDLINE | ID: mdl-33563815

RESUMO

Hepatitis B virus (HBV) core protein (Cp) can be found in the nucleus and cytoplasm of infected hepatocytes; however, it preferentially segregates to a specific compartment correlating with disease status. Regulation of this intracellular partitioning of Cp remains obscure. In this paper, we report that cellular compartments are filled and vacated by Cp in a time- and concentration-dependent manner in both transfections and infections. At early times after transfection, Cp, in a dimeric state, preferentially localizes to the nucleolus. Later, the nucleolar compartment is emptied and Cp progresses to being predominantly nuclear, with a large fraction of the protein in an assembled state. Nuclear localization is followed by cell-wide distribution, and then Cp becomes exclusively cytoplasmic. The same trend in Cp movement is seen during an infection. Putative nucleolar retention signals have been identified and appear to be structure dependent. Export of Cp from the nucleus involves the CRM1 exportin. Time-dependent flux can be recapitulated by modifying Cp concentration, suggesting transitions are regulated by reaching a threshold concentration.IMPORTANCE HBV is an endemic virus. More than 250 million people suffer from chronic HBV infection and about 800,000 die from HBV-associated disease each year. HBV is a pararetrovirus; in an infected cell, viral DNA in the nucleus is the template for viral RNA that is packaged in nascent viral capsids in the cytoplasm. Inside those capsids, while resident in cytoplasm, the linear viral RNA is reverse transcribed to form the circular double-stranded DNA (dsDNA) of the mature virus. The HBV core (or capsid) protein plays a role in almost every step of the viral life cycle. Here, we show the core protein appears to follow a programmed, sequential localization from cytoplasmic translation then into the nucleolus, to the nucleus, and back to the cytoplasm. Localization is primarily a function of time, core protein concentration, and assembly. This has important implications for our understanding of the mechanisms of antivirals that target HBV capsid assembly.


Assuntos
Nucléolo Celular/virologia , Núcleo Celular/virologia , Citoplasma/virologia , Vírus da Hepatite B/genética , Proteínas do Core Viral/genética , Proteínas do Core Viral/metabolismo , Proteínas Virais/fisiologia , Antivirais/farmacologia , Citoplasma/fisiologia , Citosol/virologia , DNA Viral/metabolismo , Vírus da Hepatite B/química , Vírus da Hepatite B/efeitos dos fármacos , Humanos , Proteínas Virais/genética , Montagem de Vírus , Replicação Viral
6.
Viruses ; 13(2)2021 01 31.
Artigo em Inglês | MEDLINE | ID: mdl-33572517

RESUMO

West Nile virus (WNV) nonstructural protein 3 (NS3) harbors the viral triphosphatase and helicase for viral RNA synthesis and, together with NS2B, constitutes the protease responsible for polyprotein processing. NS3 is a soluble protein, but it is localized to specialized compartments at the rough endoplasmic reticulum (RER), where its enzymatic functions are essential for virus replication. However, the mechanistic details behind the recruitment of NS3 from the cytoplasm to the RER have not yet been fully elucidated. In this study, we employed immunofluorescence and biochemical assays to demonstrate that NS3, when expressed individually and when cleaved from the viral polyprotein, is localized exclusively to the cytoplasm. Furthermore, NS3 appeared to be peripherally recruited to the RER and proteolytically active when NS2B was provided in trans. Thus, we provide evidence for a potential additional role for NS2B in not only serving as the cofactor for the NS3 protease, but also in recruiting NS3 from the cytoplasm to the RER for proper enzymatic activity. Results from our study suggest that targeting the interaction between NS2B and NS3 in disrupting the NS3 ER localization may be an attractive avenue for antiviral drug discovery.


Assuntos
Proteínas não Estruturais Virais/metabolismo , Replicação Viral , Febre do Nilo Ocidental/virologia , Vírus do Nilo Ocidental/fisiologia , Citoplasma/virologia , Retículo Endoplasmático Rugoso/virologia , Humanos , Transporte Proteico , RNA Helicases/genética , RNA Helicases/metabolismo , Serina Endopeptidases/genética , Serina Endopeptidases/metabolismo , Proteínas não Estruturais Virais/genética , Vírus do Nilo Ocidental/enzimologia , Vírus do Nilo Ocidental/genética
7.
Viruses ; 13(2)2021 Jan 20.
Artigo em Inglês | MEDLINE | ID: mdl-33498458

RESUMO

Giant viruses are widespread in the biosphere and play important roles in biogeochemical cycling and host genome evolution. Also known as nucleo-cytoplasmic large DNA viruses (NCLDVs), these eukaryotic viruses harbor the largest and most complex viral genomes known. Studies have shown that NCLDVs are frequently abundant in metagenomic datasets, and that sequences derived from these viruses can also be found endogenized in diverse eukaryotic genomes. The accurate detection of sequences derived from NCLDVs is therefore of great importance, but this task is challenging owing to both the high level of sequence divergence between NCLDV families and the extraordinarily high diversity of genes encoded in their genomes, including some encoding for metabolic or translation-related functions that are typically found only in cellular lineages. Here, we present ViralRecall, a bioinformatic tool for the identification of NCLDV signatures in 'omic data. This tool leverages a library of giant virus orthologous groups (GVOGs) to identify sequences that bear signatures of NCLDVs. We demonstrate that this tool can effectively identify NCLDV sequences with high sensitivity and specificity. Moreover, we show that it can be useful both for removing contaminating sequences in metagenome-assembled viral genomes as well as the identification of eukaryotic genomic loci that derived from NCLDV. ViralRecall is written in Python 3.5 and is freely available on GitHub: https://github.com/faylward/viralrecall.


Assuntos
Biologia Computacional/métodos , Citoplasma/virologia , Células Eucarióticas/virologia , Vírus Gigantes/classificação , Metagenômica , Sequência de Aminoácidos , Biodiversidade , Biblioteca Gênica , Genoma Viral , Vírus Gigantes/genética , Filogenia
8.
Curr Issues Mol Biol ; 41: 125-170, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-32764158

RESUMO

During viral replication, herpesviruses utilize a unique strategy, termed nuclear egress, to translocate capsids from the nucleus into the cytoplasm. This initial budding step transfers a newly formed capsid from within the nucleus, too large to fit through nuclear pores, through the inner nuclear membrane to the perinuclear space. The perinuclear enveloped virion must then fuse with the outer nuclear membrane to be released into the cytoplasm for further maturation, undergoing budding once again at the trans-Golgi network or early endosomes, and ultimately exit the cell non-lytically to spread infection. This first budding process is mediated by two conserved viral proteins, UL31 and UL34, that form a heterodimer called the nuclear egress complex (NEC). This review focuses on what we know about how the NEC mediates capsid transport to the perinuclear space, including steps prior to and after this budding event. Additionally, we discuss the involvement of other viral proteins in this process and how NEC-mediated budding may be regulated during infection.


Assuntos
Núcleo Celular/metabolismo , Infecções por Herpesviridae/metabolismo , Infecções por Herpesviridae/virologia , Herpesviridae/metabolismo , Membrana Nuclear/metabolismo , Capsídeo/metabolismo , Núcleo Celular/virologia , Citoplasma/metabolismo , Citoplasma/virologia , Humanos , Proteínas Virais/metabolismo , Vírion/metabolismo
9.
Curr Issues Mol Biol ; 41: 171-220, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-32807747

RESUMO

Herpesviruses virions are large and complex structures that deliver their genetic content to nuclei upon entering cells. This property is not unusual as many other viruses including the adenoviruses, orthomyxoviruses, papillomaviruses, polyomaviruses, and retroviruses, do likewise. However, the means by which viruses in the alphaherpesvirinae subfamily accomplish this fundamental stage of the infectious cycle is tied to their defining ability to efficiently invade the nervous system. Fusion of the viral envelope with a cell membrane results in the deposition of the capsid, along with an assortment of tegument proteins, into the cytosol. Establishment of infection requires that the capsid traverse the cytosol, dock at a nuclear pore, and inject its genome into the nucleoplasm. Accumulating evidence indicates that the capsid is not the effector of this delivery process, but is instead shepherded by tegument proteins that remain capsid bound. At the same time, tegument proteins that are released from the capsid upon entry act to increase the susceptibility of the cell to the ensuing infection. Mucosal epithelial cells and neurons are both susceptible to alphaherpesvirus infection and, together, provide the niche to which these viruses have adapted. Although much has been revealed about the functions of de novo expressed tegument proteins during the late stages of assembly and egress, this review will specifically address the roles of tegument proteins brought into the cell with the incoming virion, and our current understanding of alphaherpesvirus genome delivery to nuclei.


Assuntos
Alphaherpesvirinae/genética , Alphaherpesvirinae/patogenicidade , Citoplasma/virologia , Genoma Viral/genética , Infecções por Herpesviridae/virologia , Animais , Proteínas do Capsídeo/genética , Núcleo Celular/virologia , Humanos , Vírion/genética , Montagem de Vírus/genética , Internalização do Vírus
10.
EMBO J ; 40(1): e105247, 2021 01 04.
Artigo em Inglês | MEDLINE | ID: mdl-33270250

RESUMO

In order to replicate, human immunodeficiency virus (HIV-1) reverse-transcribes its RNA genome into DNA, which subsequently integrates into host cell chromosomes. These two key events of the viral life cycle are commonly viewed as separate not only in time, but also in cellular space, since reverse transcription (RT) is thought to be completed in the cytoplasm before nuclear import and integration. However, the spatiotemporal organization of the early viral replication cycle in macrophages, the natural non-dividing target cells that constitute reservoirs of HIV-1 and an obstacle to curing AIDS, remains unclear. Here, we demonstrate that infected macrophages display large nuclear foci of viral DNA (vDNA) and viral RNA, in which multiple viral genomes cluster together. These clusters form in the absence of chromosomal integration, sequester the paraspeckle protein CPSF6, and localize to nuclear speckles. Surprisingly, these viral RNA clusters consist mostly of genomic, incoming RNA, both in cells where reverse transcription is pharmacologically suppressed and in untreated cells. We demonstrate that following temporary inhibition, reverse transcription can resume in the nucleus and lead to vDNA accumulation in these clusters. We further show that nuclear reverse transcription can result in transcription-competent viral DNA. These findings change our understanding of the early HIV-1 replication cycle and may have implications for addressing HIV-1 persistence.


Assuntos
Núcleo Celular/virologia , Genoma Viral/genética , HIV-1/genética , Macrófagos/virologia , Transcrição Reversa/genética , Transporte Ativo do Núcleo Celular/genética , Linhagem Celular , Análise por Conglomerados , Citoplasma/virologia , DNA Viral/genética , Células HEK293 , Infecções por HIV/virologia , Humanos , RNA Viral/genética , Células THP-1 , Replicação Viral/genética
11.
Virus Res ; 291: 198193, 2021 01 02.
Artigo em Inglês | MEDLINE | ID: mdl-33053412

RESUMO

Rotavirus (RV) replication occurs in cytoplasmic compartments, known as viroplasms, that are composed of viral and cellular proteins. Viroplasm formation requires RV nonstructural proteins NSP2 and NSP5 and cellular lipid droplets (LDs); however, the mechanisms required for viroplasm assembly remain largely unknown. We previously identified two conformationally-distinct forms of NSP2 (dNSP2, vNSP2) found in RV-infected cells that interact differentially with hypo- and hyperphosphorylated NSP5, respectively, and indicate a coordinated phosphorylation-dependent mechanism regulating viroplasm assembly. We also reported that phosphorylation of dNSP2 on serine 313 by the cellular kinase CK1α triggers the localization of vNSP2 to sites of viroplasm assembly and its association with hyperphosphorylated NSP5. To directly evaluate the role of CK1α-mediated NSP2 phosphorylation on viroplasm formation, we used a recently published plasmid-based reverse genetics method to generate a recombinant rotavirus (rRV) with a phosphomimetic NSP2 mutation (rRV NSP2 S313D). The rRV NSP2 S313D virus is significantly delayed in viroplasm formation, virus replication, and interferes with wild type RV replication during co-infection. The rRV NSP2 S313A virus was not rescued. Taking advantage of the delay in viroplasm formation, the NSP2 S313D phosphomimetic mutant was used as a tool to observe very early events in viroplasm assembly. We show that (1) viroplasm assembly correlates with NSP5 hyperphosphorylation, and (2) that vNSP2 S313D co-localizes with RV-induced LDs without NSP5, suggesting that vNSP2 phospho-S313 is sufficient for interacting with LDs and may be the virus factor required for RV-induced LD formation. Further studies with the rRV NSP2 S313D virus are expected to reveal new aspects of viroplasm and LD initiation and assembly.


Assuntos
Citoplasma/virologia , Plasmídeos/genética , Genética Reversa/métodos , Rotavirus/genética , Rotavirus/fisiologia , Humanos , Fosforilação , Infecções por Rotavirus/virologia , Replicação Viral
12.
J Biol Chem ; 296: 100236, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33380421

RESUMO

Herpesviruses are large and complex viruses that have a long history of coevolution with their host species. One important factor in the virus-host interaction is the alteration of intracellular morphology during viral replication with critical implications for viral assembly. However, the details of this remodeling event are not well understood, in part because insufficient tools are available to deconstruct this highly heterogeneous process. To provide an accurate and reliable method of investigating the spatiotemporal dynamics of virus-induced changes to cellular architecture, we constructed a dual-fluorescent reporter virus that enabled us to classify four distinct stages in the infection cycle of herpes simplex virus-1 at the single cell level. This timestamping method can accurately track the infection cycle across a wide range of multiplicities of infection. We used high-resolution fluorescence microscopy analysis of cellular structures in live and fixed cells in concert with our reporter virus to generate a detailed and chronological overview of the spatial and temporal reorganization during viral replication. The highly orchestrated and striking relocation of many organelles around the compartments of secondary envelopment during transition from early to late gene expression suggests that the reshaping of these compartments is essential for virus assembly. We furthermore find that accumulation of HSV-1 capsids in the cytoplasm is accompanied by fragmentation of the Golgi apparatus with potential impact on the late steps of viral assembly. We anticipate that in the future similar tools can be systematically applied for the systems-level analysis of intracellular morphology during replication of other viruses.


Assuntos
Complexo de Golgi/genética , Herpesvirus Humano 1/genética , Microscopia de Fluorescência , Replicação Viral/genética , Animais , Capsídeo/ultraestrutura , Chlorocebus aethiops , Citoplasma/genética , Citoplasma/ultraestrutura , Citoplasma/virologia , Genes Reporter/genética , Complexo de Golgi/ultraestrutura , Complexo de Golgi/virologia , Herpesvirus Humano 1/ultraestrutura , Humanos , Análise de Célula Única , Análise Espaço-Temporal , Células Vero , Montagem de Vírus/genética
13.
Curr Opin Virol ; 47: 25-31, 2021 04.
Artigo em Inglês | MEDLINE | ID: mdl-33383355

RESUMO

Positive-strand RNA virus genome replication takes place on intracellular membranes that separate the reduced cytosol from the oxidized extracellular/luminal milieu. Ongoing studies of these membrane-bounded genome replication complexes have revealed underlying common principles in their structure, assembly and functionalization, including transmembrane features and redox dependencies. Among these, members of the alphavirus, flavivirus, and picornavirus supergroups all encode membrane-permeabilizing viroporins required for efficient RNA replication. For flaviviruses and particularly alphavirus supergroup members, these viroporins are linked to activating viral RNA capping and potentially other later-stage RNA replication functions, and to local transmembrane release of oxidizing potential to trigger these changes in cytoplasmic RNA replication complexes. Further exploration of these emerging shared principles could spur development of broad-spectrum antivirals.


Assuntos
Genoma Viral/fisiologia , Membranas Intracelulares/virologia , Oxirredução , Vírus de RNA de Cadeia Positiva/fisiologia , Replicação Viral/fisiologia , Citoplasma/metabolismo , Citoplasma/virologia , Membranas Intracelulares/metabolismo , Vírus de RNA de Cadeia Positiva/classificação , Capuzes de RNA/metabolismo , RNA Viral/biossíntese , Proteínas Virais/metabolismo , Compartimentos de Replicação Viral/metabolismo
14.
Nucleic Acids Res ; 49(2): 621-635, 2021 01 25.
Artigo em Inglês | MEDLINE | ID: mdl-33337475

RESUMO

The integration of retroviral reverse transcripts into the chromatin of the cells that they infect is required for virus replication. Retroviral integration has far-reaching consequences, from perpetuating deadly human diseases to molding metazoan evolution. The lentivirus human immunodeficiency virus 1 (HIV-1), which is the causative agent of the AIDS pandemic, efficiently infects interphase cells due to the active nuclear import of its preintegration complex (PIC). To enable integration, the PIC must navigate the densely-packed nuclear environment where the genome is organized into different chromatin states of varying accessibility in accordance with cellular needs. The HIV-1 capsid protein interacts with specific host factors to facilitate PIC nuclear import, while additional interactions of viral integrase, the enzyme responsible for viral DNA integration, with cellular nuclear proteins and nucleobases guide integration to specific chromosomal sites. HIV-1 integration favors transcriptionally active chromatin such as speckle-associated domains and disfavors heterochromatin including lamina-associated domains. In this review, we describe virus-host interactions that facilitate HIV-1 PIC nuclear import and integration site targeting, highlighting commonalities among factors that participate in both of these steps. We moreover discuss how the nuclear landscape influences HIV-1 integration site selection as well as the establishment of active versus latent virus infection.


Assuntos
HIV-1/fisiologia , Interações Hospedeiro-Patógeno , Proteínas do Vírus da Imunodeficiência Humana/metabolismo , Integração Viral , Transporte Ativo do Núcleo Celular , Proteínas Adaptadoras de Transdução de Sinal/deficiência , Proteínas Adaptadoras de Transdução de Sinal/fisiologia , Proteínas do Capsídeo/metabolismo , Núcleo Celular/metabolismo , Núcleo Celular/virologia , Cromatina/genética , Cromatina/metabolismo , Citoplasma/metabolismo , Citoplasma/virologia , Proteínas do Citoesqueleto/metabolismo , Transcriptase Reversa do HIV/fisiologia , HIV-1/enzimologia , HIV-1/genética , Proteínas do Vírus da Imunodeficiência Humana/genética , Humanos , Interfase , Modelos Moleculares , Complexos Multiproteicos/metabolismo , Poro Nuclear/metabolismo , Proteínas Nucleares/metabolismo , Conformação Proteica , Domínios Proteicos , Fatores de Transcrição/deficiência , Fatores de Transcrição/fisiologia , Integração Viral/genética , Integração Viral/fisiologia , Latência Viral , Replicação Viral , Fatores de Poliadenilação e Clivagem de mRNA/deficiência , Fatores de Poliadenilação e Clivagem de mRNA/fisiologia
15.
Front Immunol ; 11: 543444, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-33329509

RESUMO

Host antiviral factor interferon-induced transmembrane proteins (IFITMs) are a kind of small-molecule transmembrane proteins induced by interferon. Their broad-spectrum antiviral activity and unique ability to inhibit viral invasion have made them a hot molecule in antiviral research in recent years. Since the first demonstration of their natural ability to resist viral infection in 1996, IFITMs have been reported to limit a variety of viral infections, including some major pathogens that seriously endanger human health and social stability, such as influenza A, Ebol, severe acute respiratory syndrome, AIDS, and Zika viruses, etc. Studies show that IFITMs mainly exert antiviral activity during virus entry, specifically interfering with the fusion of the envelope and the endosome membrane or forming fusion micropores to block the virus from entering the cytoplasm. However, their specific mechanism is still unclear. This article mainly reviews the research progress in the structure, evolution, function, and mechanism of IFITMs, which may provide a theoretical basis for clarifying the molecular mechanism of interaction between the molecules and viruses and the research and development of new antiviral drugs based on IFITMs.


Assuntos
Citoplasma/imunologia , Imunidade Inata , Proteínas de Membrana/imunologia , Viroses/imunologia , Internalização do Vírus , Vírus/imunologia , Animais , Antivirais/uso terapêutico , Citoplasma/virologia , Humanos , Viroses/tratamento farmacológico
16.
mBio ; 11(6)2020 11 10.
Artigo em Inglês | MEDLINE | ID: mdl-33172997

RESUMO

HIV-1 full-length RNA (referred to as HIV-1 RNA here) serves as the viral genome in virions and as a template for Gag/Gag-Pol translation. We previously showed that HIV-1 RNA, which is exported via the CRM1 pathway, travels in the cytoplasm mainly through diffusion. A recent report suggested that the export pathway used by retroviral RNA could affect its cytoplasmic transport mechanism and localization. HIV-1 RNA export is directed by the viral protein Rev and the cis-acting element, Rev response element (RRE). When Rev/RRE is replaced with the constitutive transport element (CTE) from Mason-Pfizer monkey virus (MPMV), HIV-1 RNA is exported through the NXF1 pathway. To determine the effects of the export pathway on HIV-1 RNA, we tracked individual RNAs and found that the vast majority of cytoplasmic HIV-1 RNAs travel by diffusion regardless of the export pathway. However, CTE-containing HIV-1 RNA diffuses at a rate slower than that of RRE-containing HIV-1 RNA. Using in situ hybridization, we analyzed the subcellular localizations of HIV-1 RNAs in cells expressing a CTE-containing and an RRE-containing provirus. We found that these two types of HIV-1 RNAs have similar subcellular distributions. HIV-1 RNA exported through the NXF1 pathway was suggested to cluster near centrosomes. To investigate this possibility, we measured the distances between individual RNAs to the centrosomes and found that HIV-1 RNAs exported through different pathways do not exhibit significantly different distances to centrosomes. Therefore, HIV-1 RNAs exported through CRM1 and NXF1 pathways use the same RNA transport mechanism and exhibit similar cytoplasmic distributions.IMPORTANCE The unspliced HIV-1 full-length RNA (HIV-1 RNA) is packaged into virions as the genome and is translated to generate viral structural proteins and enzymes. To serve these functions, HIV-1 RNA must be exported from the nucleus to the cytoplasm. It was recently suggested that export pathways used by HIV-1 RNA could affect its cytoplasmic transport mechanisms and distribution. In the current report, we examined the HIV-1 RNA transport mechanism by following the movement of individual RNAs and identifying the distribution of RNA using in situ hybridization. Our results showed that whether exported by the CRM1 or NXF1 pathway, HIV-1 RNAs mainly use diffusion for cytoplasmic travel. Furthermore, HIV-1 RNAs exported using the CRM1 or NXF1 pathway are well mixed in the cytoplasm and do not display export pathway-specific clustering near centrosomes. Thus, the export pathways used by HIV-1 RNAs do not alter the cytoplasmic transport mechanisms or distribution.


Assuntos
Citoplasma/virologia , Infecções por HIV/virologia , HIV-1/metabolismo , RNA Viral/metabolismo , Transporte Ativo do Núcleo Celular , Núcleo Celular/metabolismo , Núcleo Celular/virologia , Citoplasma/metabolismo , Produtos do Gene rev/genética , Produtos do Gene rev/metabolismo , Infecções por HIV/metabolismo , HIV-1/genética , Humanos , Transporte de RNA , RNA Viral/genética
18.
Nat Commun ; 11(1): 6059, 2020 11 27.
Artigo em Inglês | MEDLINE | ID: mdl-33247099

RESUMO

Severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) was first detected in late December 2019 and has spread worldwide. Coronaviruses are enveloped, positive sense, single-stranded RNA viruses and employ a complicated pattern of virus genome length RNA replication as well as transcription of genome length and leader containing subgenomic RNAs. Although not fully understood, both replication and transcription are thought to take place in so-called double-membrane vesicles in the cytoplasm of infected cells. Here we show detection of SARS-CoV-2 subgenomic RNAs in diagnostic samples up to 17 days after initial detection of infection and provide evidence for their nuclease resistance and protection by cellular membranes suggesting that detection of subgenomic RNAs in such samples may not be a suitable indicator of active coronavirus replication/infection.


Assuntos
Teste para COVID-19/métodos , COVID-19/diagnóstico , Genoma Viral , RNA Viral/isolamento & purificação , SARS-CoV-2/genética , Replicação Viral , Adulto , COVID-19/virologia , Citoplasma/virologia , Feminino , Sequenciamento de Nucleotídeos em Larga Escala , Interações Hospedeiro-Patógeno/genética , Humanos , Masculino , Pessoa de Meia-Idade , Nasofaringe/citologia , Nasofaringe/virologia , Orofaringe/citologia , Orofaringe/virologia , Reação em Cadeia da Polimerase , SARS-CoV-2/isolamento & purificação , SARS-CoV-2/patogenicidade , Fatores de Tempo , Adulto Jovem
19.
Arch Virol ; 165(12): 2953-2959, 2020 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-33040310

RESUMO

Chrysanthemum virus B encodes a multifunctional p12 protein that acts as a transcriptional activator in the nucleus and as a suppressor of RNA silencing in the cytoplasm. Here, we investigated the impact of p12 on accumulation of major classes of small RNAs (sRNAs). The results show dramatic changes in the sRNA profiles characterised by an overall reduction in sRNA accumulation, changes in the pattern of size distribution of canonical siRNAs and in the ratio between sense and antisense strands, lower abundance of siRNAs with a U residue at the 5'-terminus, and changes in the expression of certain miRNAs, most of which were downregulated.


Assuntos
Carlavirus/genética , MicroRNAs/genética , Interferência de RNA , RNA de Plantas/genética , RNA Interferente Pequeno/genética , Chrysanthemum/genética , Chrysanthemum/virologia , Citoplasma/virologia
20.
PLoS One ; 15(10): e0241212, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-33095800

RESUMO

Hepatitis B virus (HBV) is a human pathogen of global concern, while a high diversity of viruses related to HBV have been discovered in other animals during the last decade. Recently, the novel mammalian hepadnavirus, tentatively named domestic cat hepadnavirus (DCH), was detected in an immunocompromised cat. Herein, a collection of 209 cat sera and 15 hepato-diseased cats were screened for DCH using PCR, resulting in 12.4% and 20% positivity in the tested sera and necropsied cats, respectively. Among the DCH-positive sera, a significantly high level of co-detection with retroviral infection was found, with the highest proportion being co-detection with feline immunodeficiency virus (FIV). Full-length genome characterization of DCH revealed the genetic diversity between the nine Thai DCH sequences obtained, and that they phylogenetically formed three distinct monophyletic clades. A putative DCH recombinant strain was found, suggesting a possible role of recombination in DCH evolution. Additionally, quantitative PCR was used to determine the viral copy number in various organs of the DCH-moribund cats, while the pathological findings were compared to the viral localization in hepatocytes, adjacent to areas of hepatic fibrosis, by immunohistochemical (IHC) and western blot analysis. In addition to the liver, positive-DCH immunoreactivity was found in various other organs, including kidneys, lung, heart, intestine, brain, and lymph nodes, providing evidence of systemic infection. Ultrastructure of infected cells revealed electron-dense particles in the nucleus and cytoplasm of hepatocytes, bronchial epithelial cells, and fibroblasts. We propose the intracellular development mechanism of this virus. Although the definitive roles of pathogenicity of DCH remains undetermined, a contributory role of the virus associated with systemic diseases is possible.


Assuntos
Coinfecção/veterinária , Síndrome de Imunodeficiência Adquirida Felina/virologia , Infecções por Hepadnaviridae/veterinária , Hepadnaviridae/genética , Animais de Estimação/virologia , Animais , Brônquios/citologia , Brônquios/virologia , Gatos , Coinfecção/virologia , Citoplasma/virologia , Células Epiteliais/citologia , Células Epiteliais/ultraestrutura , Células Epiteliais/virologia , Síndrome de Imunodeficiência Adquirida Felina/sangue , Feminino , Fibroblastos/citologia , Fibroblastos/virologia , Variação Genética , Genoma Viral/genética , Hepadnaviridae/isolamento & purificação , Infecções por Hepadnaviridae/virologia , Hepatócitos/citologia , Hepatócitos/ultraestrutura , Hepatócitos/virologia , Vírus da Imunodeficiência Felina/isolamento & purificação , Masculino , Microscopia Eletrônica de Transmissão , Filogenia , Recombinação Genética , Mucosa Respiratória/citologia , Mucosa Respiratória/virologia , Tailândia , Replicação Viral , Eliminação de Partículas Virais
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