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PLoS Pathogens May 2017Cap-snatching was first discovered in influenza virus. Structures of the involved domains of the influenza virus polymerase, namely the endonuclease in the PA subunit...
Cap-snatching was first discovered in influenza virus. Structures of the involved domains of the influenza virus polymerase, namely the endonuclease in the PA subunit and the cap-binding domain in the PB2 subunit, have been solved. Cap-snatching endonucleases have also been demonstrated at the very N-terminus of the L proteins of mammarena-, orthobunya-, and hantaviruses. However, a cap-binding domain has not been identified in an arena- or bunyavirus L protein so far. We solved the structure of the 326 C-terminal residues of the L protein of California Academy of Sciences virus (CASV), a reptarenavirus, by X-ray crystallography. The individual domains of this 37-kDa fragment (L-Cterm) as well as the domain arrangement are structurally similar to the cap-binding and adjacent domains of influenza virus polymerase PB2 subunit, despite the absence of sequence homology, suggesting a common evolutionary origin. This enabled identification of a region in CASV L-Cterm with similarity to a cap-binding site; however, the typical sandwich of two aromatic residues was missing. Consistent with this, cap-binding to CASV L-Cterm could not be detected biochemically. In addition, we solved the crystal structure of the corresponding endonuclease in the N-terminus of CASV L protein. It shows a typical endonuclease fold with an active site configuration that is essentially identical to that of known mammarenavirus endonuclease structures. In conclusion, we provide evidence for a presumably functional cap-snatching endonuclease in the N-terminus and a degenerate cap-binding domain in the C-terminus of a reptarenavirus L protein. Implications of these findings for the cap-snatching mechanism in arenaviruses are discussed.
Topics: Arenaviridae; Arenaviridae Infections; Crystallography, X-Ray; Endonucleases; Models, Molecular; Protein Conformation; Protein Domains; RNA Caps; Viral Proteins
PubMed: 28505175
DOI: 10.1371/journal.ppat.1006400 -
The Journal of General Virology Jan 2014Arenaviruses can cause fatal human haemorrhagic fever (HF) diseases for which vaccines and therapies are extremely limited. Both the New World (NW) and Old World (OW)... (Comparative Study)
Comparative Study Review
Arenaviruses can cause fatal human haemorrhagic fever (HF) diseases for which vaccines and therapies are extremely limited. Both the New World (NW) and Old World (OW) groups of arenaviruses contain HF-causing pathogens. Although these two groups share many similarities, important differences with regard to pathogenicity and molecular mechanisms of virus infection exist. These closely related pathogens share many characteristics, including genome structure, viral assembly, natural host selection and the ability to interfere with innate immune signalling. However, members of the NW and OW viruses appear to use different receptors for cellular entry, as well as different mechanisms of virus internalization. General differences in disease signs and symptoms and pathological lesions in patients infected with either NW or OW arenaviruses are also noted and discussed herein. Whilst both the OW Lassa virus (LASV) and the NW Junin virus (JUNV) can cause disruption of the vascular endothelium, which is an important pathological feature of HF, the immune responses to these related pathogens seem to be quite distinct. Whereas LASV infection results in an overall generalized immune suppression, patients infected with JUNV seem to develop a cytokine storm. Additionally, the type of immune response required for recovery and clearance of the virus is different between NW and OW infections. These differences may be important to allow the viruses to evade host immune detection. Understanding these differences will aid the development of new vaccines and treatment strategies against deadly HF viral infections.
Topics: Animals; Arenaviridae Infections; Arenaviruses, New World; Arenaviruses, Old World; Hemorrhagic Fevers, Viral; Humans
PubMed: 24068704
DOI: 10.1099/vir.0.057000-0 -
Current Topics in Microbiology and... 2002
Review
Topics: Animals; Arenaviridae Infections; Arenavirus; Guinea Pigs; Humans; Mice; Molecular Epidemiology
PubMed: 11987810
DOI: No ID Found -
Antiviral Research Aug 2011Reverse-genetics systems are powerful tools enabling researchers to study the replication cycle of RNA viruses, including filoviruses and other hemorrhagic fever... (Review)
Review
Minigenomes, transcription and replication competent virus-like particles and beyond: reverse genetics systems for filoviruses and other negative stranded hemorrhagic fever viruses.
Reverse-genetics systems are powerful tools enabling researchers to study the replication cycle of RNA viruses, including filoviruses and other hemorrhagic fever viruses, as well as to discover new antivirals. They include full-length clone systems as well as a number of life cycle modeling systems. Full-length clone systems allow for the generation of infectious, recombinant viruses, and thus are an important tool for studying the virus replication cycle in its entirety. In contrast, life cycle modeling systems such as minigenome and transcription and replication competent virus-like particle systems can be used to simulate and dissect parts of the virus life cycle outside of containment facilities. Minigenome systems are used to model viral genome replication and transcription, whereas transcription and replication competent virus-like particle systems also model morphogenesis and budding as well as infection of target cells. As such, these modeling systems have tremendous potential to further the discovery and screening of new antivirals targeting hemorrhagic fever viruses. This review provides an overview of currently established reverse genetics systems for hemorrhagic fever-causing negative-sense RNA viruses, with a particular emphasis on filoviruses, and the potential application of these systems for antiviral research.
Topics: Antiviral Agents; Arenaviridae; Bunyaviridae; DNA, Complementary; Filoviridae; Genes, Reporter; Genome, Viral; Ribonucleoproteins; Transcription, Genetic; Transfection; Viral Proteins; Virus Internalization; Virus Release; Virus Replication
PubMed: 21699921
DOI: 10.1016/j.antiviral.2011.06.003 -
Intervirology 1984
Review
Topics: Animals; Antibodies, Viral; Antibody Formation; Arenaviridae; Humans; Immunity, Cellular; Immunity, Innate; Killer Cells, Natural; RNA, Viral; Viral Proteins; Virus Replication
PubMed: 6389424
DOI: 10.1159/000149543 -
BMC Microbiology Apr 2024Despite repeated spillover transmission and their potential to cause significant morbidity and mortality in human hosts, the New World mammarenaviruses remain largely... (Review)
Review
Despite repeated spillover transmission and their potential to cause significant morbidity and mortality in human hosts, the New World mammarenaviruses remain largely understudied. These viruses are endemic to South America, with animal reservoir hosts covering large geographic areas and whose transmission ecology and spillover potential are driven in part by land use change and agriculture that put humans in regular contact with zoonotic hosts.We compiled published studies about Guanarito virus, Junin virus, Machupo virus, Chapare virus, Sabia virus, and Lymphocytic Choriomeningitis virus to review the state of knowledge about the viral hemorrhagic fevers caused by New World mammarenaviruses. We summarize what is known about rodent reservoirs, the conditions of spillover transmission for each of these pathogens, and the characteristics of human populations at greatest risk for hemorrhagic fever diseases. We also review the implications of repeated outbreaks and biosecurity concerns where these diseases are endemic, and steps that countries can take to strengthen surveillance and increase capacity of local healthcare systems. While there are unique risks posed by each of these six viruses, their ecological and epidemiological similarities suggest common steps to mitigate spillover transmission and better contain future outbreaks.
Topics: Animals; Humans; Arenaviridae; Arenaviruses, New World; South America
PubMed: 38575867
DOI: 10.1186/s12866-024-03257-w -
Scientific Reports Oct 2021Viruses need cells for their replication and, therefore, ways to hijack cellular functions. Mitochondria play fundamental roles within the cell in metabolism, immunity...
Viruses need cells for their replication and, therefore, ways to hijack cellular functions. Mitochondria play fundamental roles within the cell in metabolism, immunity and regulation of homeostasis due to which some viruses aim to alter mitochondrial functions. Herein we show that the nucleoprotein (NP) of arenaviruses enters the mitochondria of infected cells, affecting the mitochondrial morphology. Reptarenaviruses cause boid inclusion body disease (BIBD) that is characterized, especially in boas, by the formation of cytoplasmic inclusion bodies (IBs) comprising reptarenavirus NP within the infected cells. We initiated this study after observing electron-dense material reminiscent of IBs within the mitochondria of reptarenavirus infected boid cell cultures in an ultrastructural study. We employed immuno-electron microscopy to confirm that the mitochondrial inclusions indeed contain reptarenavirus NP. Mutations to a putative N-terminal mitochondrial targeting signal (MTS), identified via software predictions in both mamm- and reptarenavirus NPs, did not affect the mitochondrial localization of NP, suggesting that it occurs independently of MTS. In support of MTS-independent translocation, we did not detect cleavage of the putative MTSs of arenavirus NPs in reptilian or mammalian cells. Furthermore, in vitro translated NPs could not enter isolated mitochondria, suggesting that the translocation requires cellular factors or conditions. Our findings suggest that MTS-independent mitochondrial translocation of NP is a shared feature among arenaviruses. We speculate that by targeting the mitochondria arenaviruses aim to alter mitochondrial metabolism and homeostasis or affect the cellular defense.
Topics: Animals; Arenaviridae; Boidae; Chlorocebus aethiops; Inclusion Bodies, Viral; Mitochondria; Nucleoproteins; Vero Cells
PubMed: 34702948
DOI: 10.1038/s41598-021-99887-5 -
Nature Communications Jan 2022Five New World mammarenaviruses (NWMs) cause life-threatening hemorrhagic fever (HF). Cellular entry by these viruses is mediated by human transferrin receptor 1...
Five New World mammarenaviruses (NWMs) cause life-threatening hemorrhagic fever (HF). Cellular entry by these viruses is mediated by human transferrin receptor 1 (hTfR1). Here, we demonstrate that an antibody (ch128.1/IgG1) which binds the apical domain of hTfR1, potently inhibits infection of attenuated and pathogenic NWMs in vitro. Computational docking of the antibody Fab crystal structure onto the known structure of hTfR1 shows an overlapping receptor-binding region shared by the Fab and the viral envelope glycoprotein GP1 subunit that binds hTfR1, and we demonstrate competitive inhibition of NWM GP1 binding by ch128.1/IgG1 as the principal mechanism of action. Importantly, ch128.1/IgG1 protects hTfR1-expressing transgenic mice against lethal NWM challenge. Additionally, the antibody is well-tolerated and only partially reduces ferritin uptake. Our findings provide the basis for the development of a novel, host receptor-targeted antibody therapeutic broadly applicable to the treatment of HF of NWM etiology.
Topics: A549 Cells; Animals; Antibodies, Monoclonal; Antigens, CD; Arenaviridae; Chlorocebus aethiops; Hemorrhagic Fever, American; Host-Pathogen Interactions; Humans; Junin virus; Mice, Inbred C57BL; Mice, Transgenic; Molecular Docking Simulation; Protein Binding; Receptors, Transferrin; Vero Cells; Viral Envelope Proteins; Mice
PubMed: 35091550
DOI: 10.1038/s41467-021-27949-3 -
Viruses Jul 2022Replication-competent reporter-expressing viruses are crucial tools in molecular virology with applications that range from antiviral screening to live-cell imaging of...
Replication-competent reporter-expressing viruses are crucial tools in molecular virology with applications that range from antiviral screening to live-cell imaging of protein spatiotemporal dynamics. However, there is currently little information available regarding viable strategies to develop reporter-expressing arenaviruses. To address this, we used Tacaribe virus (TCRV), an apathogenic BSL2 arenavirus, to assess the feasibility of different reporter expression approaches. We first generated trisegmented TCRV viruses with either the glycoprotein (GP) or nucleoprotein (NP) replaced by a reporter (GFP, mCherry, or nanoluciferase). These viruses were all viable, but showed marked differences in brightness and attenuation. Next, we generated terminal fusions with each of the TCRV proteins (i.e., NP, GP, polymerase (L), matrix protein (Z)) either with or without a T2A self-cleavage site. We tested both the function of the reporter-fused proteins alone, and the viability of corresponding recombinant TCRVs. We successfully rescued viruses with both direct and cleavable reporter fusions at the C-terminus of Z, as well as cleavable N-terminal fusions with NP. These viruses all displayed detectable reporter activity, but were also moderately attenuated. Finally, reporter proteins were inserted into a flexible hinge region within L. These viruses were also viable and showed moderate attenuation; however, reporter expression was only detectable for the luminescent virus. These strategies provide an exciting range of new tools for research into the molecular biology of TCRV that can likely also be adapted to other arenaviruses.
Topics: Arenaviridae; Arenavirus; Arenaviruses, New World; Nucleoproteins; Virus Replication
PubMed: 35891543
DOI: 10.3390/v14071563 -
Microbiology Spectrum Aug 2022Mammarenaviruses establish a persistent infection in their rodent and bat hosts, and the evidence suggests that reptarenaviruses and hartmaniviruses found in captive...
Mammarenaviruses establish a persistent infection in their rodent and bat hosts, and the evidence suggests that reptarenaviruses and hartmaniviruses found in captive snakes act similarly. In snakes, reptarenaviruses cause boid inclusion body disease (BIBD), which is often associated with secondary infections. Snakes with BIBD usually carry more than a single pair of reptarenavirus S and L segments and occasionally demonstrate hartmanivirus coinfection. Here, we reported the generation of cell lines persistently infected with a single or two reptarenavirus(es) and a cell line with persistent reptarenavirus-hartmanivirus coinfection. By RT-PCR we demonstrated that the amount of viral RNA within the persistently infected cells remains at levels similar to those observed following initial infection. Using antibodies against the glycoproteins (GPs) and nucleoprotein (NP) of reptarenaviruses, we studied the levels of viral protein in cells passaged 10 times after the original inoculation and observed that the expression of GPs declines dramatically during persistent infection, unlike the expression of NP. Immunofluorescence (IF) staining served to demonstrate differences in the distribution of NP within the persistently infected compared to freshly infected cells. IF staining of cells inoculated with the viruses secreted from the persistently infected cell lines produced similar NP staining compared to cells infected with a traditionally passaged virus, suggesting that the altered NP expression pattern of persistently infected cells does not relate to changes in the virus. The cell cultures described herein can serve as tools for studying the coinfection and superinfection interplay between reptarenaviruses and studying the BIBD pathogenesis mechanisms. Mammarenaviruses cause a persistent infection in their natural rodent and bat hosts. Reptarenaviruses cause boid inclusion body disease (BIBD) in constrictor snakes, but it is unclear whether snakes are the natural host of these viruses. In this study, we showed that reptarenaviruses established a persistent infection in cultured Boa constrictor cells and that the persistently infected cells continued to produce infectious virus. Our results showed that persistent infection results from subsequent passaging of cells inoculated with a single reptarenavirus, two reptarenaviruses, or even when inoculating the cells with reptarenavirus and hartmanivirus (another arenavirus genus). The results further suggested that coinfection would not result in overt competition between the different reptarenaviruses, thus helping to explain the frequent reptarenavirus coinfections in snakes with BIBD. The established cell culture models of persistent infection could help to elucidate the role of coinfection and superinfection and potential immunosuppression as the pathogenic mechanisms behind BIBD.
Topics: Animals; Arenaviridae; Boidae; Cell Line; Chiroptera; Coinfection; Superinfection
PubMed: 35862992
DOI: 10.1128/spectrum.01585-22