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      Molecular characterisation of RIG-I-like helicases in the black flying fox, Pteropus alecto

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          Highlights

          ► Molecular characterisation of RIG-I-like helicases in the fruit bat Pteropus alecto. ► Bat RLHs are structurally conserved with those in other vertebrates. ► Bat RLHs are ubiquitously expressed and enriched in immune tissues. ► Treatment of bat kidney cells with poly I:C produces an fibroblast-like IFN response. ► Poly I:C induces upregulation of bat RLHs, indicating they are also ISGs.

          Abstract

          The RIG-I like helicases, RIG-I, mda5 and LGP2 are an evolutionarily conserved family of cytosolic pattern recognition receptors important in the recognition of viral RNA, and responsible for the innate induction of interferons and proinflammatory cytokines upon viral infection. Bats are natural reservoir hosts to a variety of RNA viruses that cause significant morbidity and mortality in other species; however the mechanisms responsible for the control of viral replication in bats are not understood. This report describes the molecular cloning and expression analysis of RIG-I, mda5 and LGP2 genes in the fruit bat Pteropus alecto, and is the first description of RIG-I like helicases from any species of bat. Our results demonstrate that P. alecto RIG-I, mda5 and LGP2 have similar primary structures and tissue expression patterns to their counterparts in humans and other mammals. Stimulation of bat kidney cells with synthetic dsRNA (poly I:C) induced high levels of interferon β and rapid upregulation of all three helicases. These findings reveal that the cytoplasmic virus sensing machinery is present and intact in P. alecto. This study provides the foundation for further investigations into the interactions between bat RIG-I-like helicases and viruses to elucidate the mechanisms responsible for the asymptomatic nature of viral infections in bats.

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          Most cited references33

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          Severe acute respiratory syndrome coronavirus-like virus in Chinese horseshoe bats.

          Although the finding of severe acute respiratory syndrome coronavirus (SARS-CoV) in caged palm civets from live animal markets in China has provided evidence for interspecies transmission in the genesis of the SARS epidemic, subsequent studies suggested that the civet may have served only as an amplification host for SARS-CoV. In a surveillance study for CoV in noncaged animals from the wild areas of the Hong Kong Special Administration Region, we identified a CoV closely related to SARS-CoV (bat-SARS-CoV) from 23 (39%) of 59 anal swabs of wild Chinese horseshoe bats (Rhinolophus sinicus) by using RT-PCR. Sequencing and analysis of three bat-SARS-CoV genomes from samples collected at different dates showed that bat-SARS-CoV is closely related to SARS-CoV from humans and civets. Phylogenetic analysis showed that bat-SARS-CoV formed a distinct cluster with SARS-CoV as group 2b CoV, distantly related to known group 2 CoV. Most differences between the bat-SARS-CoV and SARS-CoV genomes were observed in the spike genes, ORF 3 and ORF 8, which are the regions where most variations also were observed between human and civet SARS-CoV genomes. In addition, the presence of a 29-bp insertion in ORF 8 of bat-SARS-CoV genome, not in most human SARS-CoV genomes, suggests that it has a common ancestor with civet SARS-CoV. Antibody against recombinant bat-SARS-CoV nucleocapsid protein was detected in 84% of Chinese horseshoe bats by using an enzyme immunoassay. Neutralizing antibody to human SARS-CoV also was detected in bats with lower viral loads. Precautions should be exercised in the handling of these animals.
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            Shared and unique functions of the DExD/H-box helicases RIG-I, MDA5, and LGP2 in antiviral innate immunity.

            The cellular protein retinoic acid-inducible gene I (RIG-I) senses intracellular viral infection and triggers a signal for innate antiviral responses including the production of type I IFN. RIG-I contains a domain that belongs to a DExD/H-box helicase family and exhibits an N-terminal caspase recruitment domain (CARD) homology. There are three genes encoding RIG-I-related proteins in human and mouse genomes. Melanoma differentiation associated gene 5 (MDA5), which consists of CARD and a helicase domain, functions as a positive regulator, similarly to RIG-I. Both proteins sense viral RNA with a helicase domain and transmit a signal downstream by CARD; thus, these proteins share overlapping functions. Another protein, LGP2, lacks the CARD homology and functions as a negative regulator by interfering with the recognition of viral RNA by RIG-I and MDA5. The nonstructural protein 3/4A protein of hepatitis C virus blocks the signaling by RIG-I and MDA5; however, the V protein of the Sendai virus selectively abrogates the MDA5 function. These results highlight ingenious mechanisms for initiating antiviral innate immune responses and the action of virus-encoded inhibitors.
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              Isolation of Hendra virus from pteropid bats: a natural reservoir of Hendra virus.

              Since it was first described in Australia in 1994, Hendra virus (HeV) has caused two outbreaks of fatal disease in horses and humans, and an isolated fatal horse case. Our preliminary studies revealed a high prevalence of neutralizing antibodies to HeV in bats of the genus PTEROPUS:, but it was unclear whether this was due to infection with HeV or a related virus. We developed the hypothesis that HeV excretion from bats might be related to the birthing process and we targeted the reproductive tract for virus isolation. Three virus isolates were obtained from the uterine fluid and a pool of foetal lung and liver from one grey-headed flying-fox (Pteropus poliocephalus), and from the foetal lung of one black flying-fox (P. alecto). Antigenically, these isolates appeared to be closely related to HeV, returning positive results on immunofluorescent antibody staining and constant-serum varying-virus neutralization tests. Using an HeV-specific oligonucleotide primer pair, genomic sequences of the isolates were amplified. Sequencing of 200 nucleotides in the matrix gene identified that these three isolates were identical to HeV. Isolations were confirmed after RNA extracted from original material was positive for HeV RNA when screened on an HeV Taqman assay. The isolation of HeV from pteropid bats corroborates our earlier serological and epidemiological evidence that they are a natural reservoir host of the virus.
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                Author and article information

                Contributors
                Journal
                Dev Comp Immunol
                Dev. Comp. Immunol
                Developmental and Comparative Immunology
                Elsevier Science
                0145-305X
                1879-0089
                3 December 2011
                April 2012
                3 December 2011
                : 36
                : 4
                : 657-664
                Affiliations
                [a ]CSIRO Livestock Industries, Australian Animal Health Laboratory, 5 Portarlington Rd, East Geelong, Victoria 3220, Australia
                [b ]Center for Evolutionary and Theoretical Immunology, Department of Biology, The University of New Mexico, Albuquerque, New Mexico 87131, USA
                [c ]State Key Laboratory of Virology, Wuhan Institute of Virology, Chinese Academy of Sciences, Wuhan, China
                Author notes
                [* ]Corresponding authors at: CSIRO Livestock Industries, Australian Animal Health Laboratory, 5 Portarlington Rd, East Geelong, Victoria 3220, Australia. Tel.: +613 52275052; fax: +613 52275555 (M.L. Baker), tel.: +613 52275026; fax: +613 52275555 (C. Cowled). chris.cowled@ 123456csiro.au michelle.baker@ 123456csiro.au
                Article
                S0145-305X(11)00308-9
                10.1016/j.dci.2011.11.008
                7103216
                22166340
                cc08a4ed-62bc-47a9-a752-aaa004f81774
                Crown copyright © 2011 Published by Elsevier Ltd. All rights reserved.

                Since January 2020 Elsevier has created a COVID-19 resource centre with free information in English and Mandarin on the novel coronavirus COVID-19. The COVID-19 resource centre is hosted on Elsevier Connect, the company's public news and information website. Elsevier hereby grants permission to make all its COVID-19-related research that is available on the COVID-19 resource centre - including this research content - immediately available in PubMed Central and other publicly funded repositories, such as the WHO COVID database with rights for unrestricted research re-use and analyses in any form or by any means with acknowledgement of the original source. These permissions are granted for free by Elsevier for as long as the COVID-19 resource centre remains active.

                History
                : 20 July 2011
                : 16 November 2011
                : 16 November 2011
                Categories
                Article

                Developmental biology
                innate immunity,viral infection,fruit bat,pteropus alecto,rig-i,mda5,lgp2
                Developmental biology
                innate immunity, viral infection, fruit bat, pteropus alecto, rig-i, mda5, lgp2

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