An infectious cDNA clone of a growth attenuated Korean isolate of MERS coronavirus KNIH002 in clade B

Kim, Minwoo; Cho, Hee; Lee, Jun Young; Park, Woo Jung; Kim, Jeong‐Min; Moon, Jae Su; Kim, Geon Woo; Lee, Wooseong; Jung, Hae Gwang; Yang, Jeong Sun; Choi, Jang Hoon; Lee, Joo Yeon; Kim, Sung Soon; Oh, Jisun

doi:10.1080/22221751.2020.1861914

Cited by 7 publications

(7 citation statements)

References 37 publications

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“…Despite there being no discernible difference in virulence among Con1 and its two derivatives Con1/MR_NS5 and Con1/MR_NS1–5, a substantial difference in growth rate between Con1/MR_NS5 (more dramatically with Con1) and Con1/MR_NS1–5 was observed in the WT C57BL/6 mice treated with MAb-5A3 antibody ( Figures 7 E–G)). In this mouse infection model mimicking a partial type I IFN signaling blocking state caused by ZIKV NS5-mediated human, but not mouse, STAT2 degradation [ 43 ], Con1 replication, as observed in Vero cells known to be defective in IFN production [ 44 , 45 ], was enhanced by MR766 NS1–5 and even by NS5 alone, as evidenced by increased viral loads (RNA genome and infectious virus levels) on 4 and 7 dpi.…”

Section: Resultsmentioning

confidence: 99%

Influence of Zika virus 3′-end sequence and nonstructural protein evolution on the viral replication competence and virulence

Jung

Cho

Kim

et al. 2022

Emerging Microbes & Infections

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Zika virus (ZIKV) has been circulating in human networks over 70 years since its first appearance in Africa, yet little is known about whether the viral 3′-terminal sequence and nonstructural (NS) protein diverged genetically from ancient ZIKV have different effects on viral replication and virulence in currently prevailing Asian lineage ZIKV. Here we show, by a reverse genetics approach using an infectious cDNA clone for a consensus sequence (Con1) of ZIKV, which represents Asian ZIKV strains, and another clone derived from the MR766 strain isolated in Uganda, Africa in 1947, that the 3′-end sequence –UUUCU-3′ homogeneously present in MR766 genome and the –GUCU-3′ sequence strictly conserved in Asian ZIKV isolates are functionally equivalent in viral replication and gene expression. By gene swapping experiments using the two infectious cDNA clones, we show that the NS1–5 proteins of MR766 enhance replication competence of ZIKV Con1. The Con1, which was less virulent than MR766, acquired severe bilateral hindlimb paralysis when its NS1–5 genes were replaced by the counterparts of MR766 in type I interferon receptor (IFNAR1)-deficient A129 mice. Moreover, MR766 NS5 RNA-dependent RNA polymerase (RdRp) alone also rendered the Con1 virulent, despite there being no difference in RdRp activity between MR766 and Con1 NS5 proteins. By contrast, the Con1 derivatives expressing MR766 Nsps, like Con1, did not develop severe disease in wild-type mice treated with an IFNAR1 blocking antibody. Together, our findings uncover an unprecedented role for ZIKV NS proteins in determining viral pathogenicity in immunocompromised hosts.

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Section: Resultsmentioning

confidence: 99%

Influence of Zika virus 3′-end sequence and nonstructural protein evolution on the viral replication competence and virulence

Jung

Cho

Kim

et al. 2022

Emerging Microbes & Infections

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“…They are suitable for cloning large DNA fragments in synthetic low-copy number plasmids that stabilize large DNA insertions, thus preventing the expression of potentially toxic regions associated with their genome [38]. Infectious DNA clones based on BAC plasmids have been established for various RNA viruses, including coronavirus [39][40][41][42], influenza virus [43,44], and flavivirus [33,45] In this investigation, a reverse genetic system was developed to generate synthetic viruses from genetically stable full-length infectious JEV3 cDNA. The CMV promoter was introduced upstream of the JEV3 genomic cDNA to transcribe genomic RNA and the HDVr sequence was added to the 3 ′ end to maintain the natural 3 ′ terminus of JEV (Figure 1).…”

Section: Discussionmentioning

confidence: 99%

Insights into the Pathogenesis and Development of Recombinant Japanese Encephalitis Virus Genotype 3 as a Vaccine

Park,

Lee,

Jun

et al. 2024

Vaccines

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Japanese encephalitis virus (JEV), a flavivirus transmitted by mosquitoes, has caused epidemics and severe neurological diseases in Asian countries. In this study, we developed a cDNA infectious clone, pBAC JYJEV3, of the JEV genotype 3 strain (EF571853.1) using a bacterial artificial chromosome (BAC) vector. The constructed infectious clone was transfected into Vero cells, where it exhibited infectivity and induced cytopathic effects akin to those of the parent virus. Confocal microscopy confirmed the expression of the JEV envelope protein. Comparative analysis of growth kinetics revealed similar replication dynamics between the parental and recombinant viruses, with peak titers observed 72 h post-infection (hpi). Furthermore, plaque assays demonstrated comparable plaque sizes and morphologies between the viruses. Cryo-electron microscopy confirmed the production of recombinant virus particles with a morphology identical to that of the parent virus. Immunization studies in mice using inactivated parental and recombinant viruses revealed robust IgG responses, with neutralizing antibody production increasing over time. These results showcase the successful generation and characterization of a recombinant JEV3 virus and provide a platform for further investigations into JEV pathogenesis and vaccine development.

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“…A nanoluciferase (Nluc)-expressing live, chimeric SARS-CoV-2 carrying the Omicron BA.5 S gene (derived from the BA.5 strain NCCP 43426) was generated using a full-length cDNA of the G-clade SARS-CoV-2 YS006 23 that was cloned in a bacterial artificial chromosomal plasmid to obtain pBAC-SARS-CoV-2/YS006(BA.5_S)_Nluc, in which the ORF7 was replaced by the Nluc gene, as previously described 50 . The recombinant chimeric virus, rYS006(BA.5_S)_Nluc, was rescued by co-culturing of BHK-21 cells transfected with pBAC-SARS-CoV-2/YS006(BA.5_S)_Nluc with VeroE6/TMPRSS2 cells at 6 h post-transfection, as previously described 50 . The co-cultured cells were incubated until clear signs of cytopathic effects (CPE) were observed.…”

Section: Methodsmentioning

confidence: 99%

Enhanced Omicron subvariant cross-neutralization efficacy of a monovalent SARS-CoV-2 BA.4/5 mRNA vaccine encoding a noncleaved, nonfusogenic spike antigen

Seo,

Jung,

Woo

et al. 2023

Preprint

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The rapid emergence of diverse SARS-CoV-2 variants, notably the Omicron variant, poses challenges to vaccine development. Here, we present a noncleaved, nonfusogenic spike (S) protein eliciting robust B- and T-cell immune responses against Omicron BA.5. The antigen incorporates the R685S and R815A mutations, effectively preventing the shedding of the S1 subunit and eliminating fusogenic activity of the resulting S antigen, termed S(SA). Through reverse genetic analysis, we found that the noncleaved form S protein with the R685S mutation enhances ACE2-dependent viral entry in vitro compared to the wild-type S protein, without increasing the virulence of the mutant virus in mice. The mRNA vaccine encoding the Omicron BA.4/5 S(SA) antigen conferred protective immunity in mice following two doses of 1 μg Ψ-UTP- or UTP-incorporated mRNA vaccines. Despite a roughly 6-fold reduction in neutralizing potency, both mRNA vaccines exhibited broad neutralizing efficacy against Omicron subvariants, including the XBB lineage variants XBB.1.5 and XBB.1.16.

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An infectious cDNA clone of a growth attenuated Korean isolate of MERS coronavirus KNIH002 in clade B

Cited by 7 publications

References 37 publications

Influence of Zika virus 3′-end sequence and nonstructural protein evolution on the viral replication competence and virulence

Influence of Zika virus 3′-end sequence and nonstructural protein evolution on the viral replication competence and virulence

Insights into the Pathogenesis and Development of Recombinant Japanese Encephalitis Virus Genotype 3 as a Vaccine

Enhanced Omicron subvariant cross-neutralization efficacy of a monovalent SARS-CoV-2 BA.4/5 mRNA vaccine encoding a noncleaved, nonfusogenic spike antigen

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