An RNA Pseudoknot Is Required for Production of Yellow Fever Virus Subgenomic RNA by the Host Nuclease XRN1

ABSTRACT Cells and mice infected with arthropod-borne flaviviruses produce a small subgenomic RNA that is colinear with the distal part of the viral 3′-untranslated region (UTR). This small subgenomic flavivirus RNA (sfRNA) results from the incomplete degradation of the viral genome by the host 5′-3′ exonuclease XRN1. Production of the sfRNA is important for the pathogenicity of the virus. This study not only presents a detailed description of the yellow fever virus (YFV) sfRNA but, more importantly, describes for the first time the molecular characteristics of the stalling site for XRN1 in the flavivirus genome. Similar to the case for West Nile virus, the YFV sfRNA was produced by XRN1. However, in contrast to the case for other arthropod-borne flaviviruses, not one but two sfRNAs were detected in YFV-infected mammalian cells. The smaller of these two sfRNAs was not observed in infected mosquito cells. The larger sfRNA could also be produced in vitro by incubation with purified XRN1. These two YFV sfRNAs formed a 5′-nested set. The 5′ ends of the YFV sfRNAs were found to be just upstream of the previously predicted RNA pseudoknot PSK3. RNA structure probing and mutagenesis studies provided strong evidence that this pseudoknot structure was formed and served as the molecular signal to stall XRN1. The sequence involved in PSK3 formation was cloned into the Sinrep5 expression vector and shown to direct the production of an sfRNA-like RNA. These results underscore the importance of the RNA pseudoknot in stalling XRN1 and also demonstrate that it is the sole viral requirement for sfRNA production.

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Yellow Fever Virus Down-Regulates mRNA Expression of SOCS1 in the Initial Phase of Infection in Human Cell Lines

Viruses ◽

10.3390/v12080802 ◽

2020 ◽

Vol 12 (8) ◽

pp. 802

Author(s):

Michael B. Yakass ◽

David Franco ◽

Osbourne Quaye

Keyword(s):

Mrna Expression ◽

Yellow Fever ◽

Yellow Fever Virus ◽

Expression Patterns ◽

Fever Virus ◽

Effective Vaccine ◽

Interferon Β ◽

Infected Cells ◽

Insight Into

Flaviviruses are constantly evolving diverse immune evasion strategies, and the exploitation of the functions of suppressors of cytokine signalling (SOCS) and protein inhibitors of activated STATs (PIAS) to favour virus replication has been described for Dengue and Japanese encephalitis viruses but not for yellow fever virus (YFV), which is still of global importance despite the existence of an effective vaccine. Some mechanisms that YFV employs to evade host immune defence has been reported, but the expression patterns of SOCS and PIAS in infected cells is yet to be determined. Here, we show that SOCS1 is down-regulated early in YFV-infected HeLa and HEK 293T cells, while SOCS3 and SOCS5 are not significantly altered, and PIAS mRNA expression appears to follow a rise-dip pattern akin to circadian-controlled genes. We also demonstrate that YFV evades interferon-β application to produce comparable viral titres. This report provides initial insight into the in vitro expression dynamics of SOCS and PIAS upon YFV infection and a basis for further investigation into SOCS/PIAS expression and how these modulate the immune response in animal models.

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Yellow fever virus is susceptible to sofosbuvir both in vitro and in vivo

PLoS Neglected Tropical Diseases ◽

10.1371/journal.pntd.0007072 ◽

2019 ◽

Vol 13 (1) ◽

pp. e0007072 ◽

Cited By ~ 31

Author(s):

Caroline S. de Freitas ◽

Luiza M. Higa ◽

Carolina Q. Sacramento ◽

André C. Ferreira ◽

Patrícia A. Reis ◽

...

Keyword(s):

Yellow Fever ◽

Yellow Fever Virus ◽

Fever Virus

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Inhibitory effect of essential oils obtained from plants grown in Colombia on yellow fever virus replication in vitro

Annals of Clinical Microbiology and Antimicrobials ◽

10.1186/1476-0711-8-8 ◽

2009 ◽

Vol 8 (1) ◽

pp. 8 ◽

Cited By ~ 39

Author(s):

Rocio Meneses Lopez ◽

Raquel E Ocazionez ◽

Jairo R Martinez ◽

Elena E Stashenko

Keyword(s):

Essential Oils ◽

Yellow Fever ◽

Virus Replication ◽

Yellow Fever Virus ◽

Inhibitory Effect ◽

Fever Virus

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CULTURE OF YELLOW-FEVER VIRUS IN VITRO

The Lancet ◽

10.1016/s0140-6736(01)08234-4 ◽

1940 ◽

Vol 236 (6102) ◽

pp. 163-164 ◽

Cited By ~ 1

Author(s):

G.M. Findlay ◽

F.O. Maccallum

Keyword(s):

Yellow Fever ◽

Yellow Fever Virus ◽

Fever Virus

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High Fidelity of Yellow Fever Virus RNA Polymerase

Journal of Virology ◽

10.1128/jvi.78.2.1032-1038.2004 ◽

2004 ◽

Vol 78 (2) ◽

pp. 1032-1038 ◽

Cited By ~ 82

Author(s):

Konstantin V. Pugachev ◽

Farshad Guirakhoo ◽

Simeon W. Ocran ◽

Fred Mitchell ◽

Megan Parsons ◽

...

Keyword(s):

Dengue Virus ◽

Rna Polymerase ◽

Yellow Fever ◽

Error Rate ◽

Yellow Fever Virus ◽

Protein A ◽

Fever Virus ◽

Genome Replication ◽

Virus Rna

ABSTRACT Three consecutive plaque purifications of four chimeric yellow fever virus-dengue virus (ChimeriVax-DEN) vaccine candidates against dengue virus types 1 to 4 were performed. The genome of each candidate was sequenced by the consensus approach after plaque purification and additional passages in cell culture. Our data suggest that the nucleotide sequence error rate for SP6 RNA polymerase used in the in vitro transcription step to initiate virus replication was as high as 1.34 × 10−4 per copied nucleotide and that the error rate of the yellow fever virus RNA polymerase employed by the chimeras for genome replication in infected cells was as low as 1.9 × 10−7 to 2.3 × 10−7. Clustering of beneficial mutations that accumulated after multiple virus passages suggests that the N-terminal part of the prM protein, a specific site in the middle of the E protein, and the NS4B protein may be essential for nucleocapsid-envelope interaction during flavivirus assembly.

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THE ADAPTATION OF UNMODIFIED STRAINS OF YELLOW FEVER VIRUS TO CULTIVATION IN VITRO

Journal of Experimental Medicine ◽

10.1084/jem.65.6.801 ◽

1937 ◽

Vol 65 (6) ◽

pp. 801-808 ◽

Cited By ~ 19

Author(s):

Hugh H. Smith ◽

Max Theiler

Keyword(s):

Brain Tissue ◽

Yellow Fever ◽

Mouse Embryo ◽

Yellow Fever Virus ◽

Fever Virus ◽

In Vitro Cultivation ◽

Virus Content ◽

Cultivation In Vitro ◽

Embryo Brain

1. In a search for suitable tissues for the cultivation of yellow fever virus in vitro, mouse embryos were inoculated with this virus in utero. A titration for virus content of the various organs of the embryos indicated that the virus was present in the brain in greatest concentration. 2. Unmodified strains of yellow fever virus were readily adapted to cultivation in vitro in a medium consisting of minced mouse embryo brain tissue and Tyrode solution containing 10 per cent normal monkey serum. 3. After a continued cultivation in mouse embryo brain tissue cultures for twenty to twenty-five subcultures, these strains were readily adapted to cultivation in whole mouse embryo tissue medium. 4. There is evidence to indicate that a prolonged cultivation of the virus in mouse embryo brain medium increases its neurotropic properties. 5. Attempts to employ monkey tissues for in vitro cultivation of yellow fever virus gave entirely negative results.

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AT-752, a double prodrug of a guanosine nucleotide analog, inhibits yellow fever virus in a hamster model

10.1101/2021.10.25.465665 ◽

2021 ◽

Author(s):

Kai Lin ◽

Steven S Good ◽

Justin G. Julander ◽

Abbie Weight ◽

Adel Moussa

Keyword(s):

Yellow Fever ◽

Yellow Fever Virus ◽

Limit Of Detection ◽

Virus Titer ◽

Fever Virus ◽

Effective Vaccine ◽

Hamster Model ◽

Nucleotide Analog

Yellow fever virus (YFV) is a zoonotic pathogen re-emerging in parts of the world, causing a viral hemorrhagic fever associated with high mortality rates. While an effective vaccine is available, having an effective antiviral against YFV is critical against unexpected outbreaks, or when vaccination is not recommended. We have previously identified AT-281, the free base of AT-752, an orally available double prodrug of a guanosine nucleotide analog, as a potent inhibitor of YFV in vitro , with a 50% effective concentration (EC 50 ) of 0.31 µM. In hamsters infected with YFV (Jimenez strain), viremia rose about 4 log 10 -fold and serum alanine aminotransferase (ALT) 2-fold compared to sham-infected animals. Treatment with 1000 mg/kg AT-752 for 7 days, initiated 4 h prior to viral challenge, reduced viremia to below the limit of detection by day 4 post infection (pi) and returned ALT to normal levels by day 6 pi. When treatment with AT-752 was initiated 2 days pi, the virus titer and ALT dropped >2 log 10 and 53% by day 4 and 6 pi, respectively. In addition, at 21 days pi, 70 – 100% of the infected animals in the treatment groups survived compared to 0% of the untreated group (p<0.001). Moreover, in vivo formation of the active triphosphate metabolite AT-9010 was measured in the animal tissues, with the highest concentrations in liver and kidney, organs that are vulnerable to the virus. The demonstrated in vivo activity of AT-752 suggests that it is a promising compound for clinical development in the treatment of YFV infection.

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RNA interference inhibits yellow fever virus replication in vitro and in vivo

Virus Genes ◽

10.1007/s11262-009-0328-3 ◽

2009 ◽

Vol 38 (2) ◽

pp. 224-231 ◽

Cited By ~ 28

Author(s):

Carolina C. Pacca ◽

Adriana A. Severino ◽

Adriano Mondini ◽

Paula Rahal ◽

Solange G. P. D’avila ◽

...

Keyword(s):

Rna Interference ◽

Yellow Fever ◽

Virus Replication ◽

Yellow Fever Virus ◽

Fever Virus

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Exchanging the Yellow Fever Virus Envelope Proteins with Modoc Virus prM and E Proteins Results in a Chimeric Virus That Is Neuroinvasive in SCID Mice

Journal of Virology ◽

10.1128/jvi.78.14.7418-7426.2004 ◽

2004 ◽

Vol 78 (14) ◽

pp. 7418-7426 ◽

Cited By ~ 16

Author(s):

Nathalie Charlier ◽

Richard Molenkamp ◽

Pieter Leyssen ◽

Jan Paeshuyse ◽

Christian Drosten ◽

...

Keyword(s):

Yellow Fever ◽

Yellow Fever Virus ◽

Vero Cells ◽

Envelope Proteins ◽

Scid Mice ◽

Fever Virus ◽

Chimeric Virus ◽

E Proteins ◽

Virus Envelope

ABSTRACT A chimeric flavivirus infectious cDNA was constructed by exchanging the premembrane (prM) and envelope (E) genes of the yellow fever virus vaccine strain 17D (YF17D) with the corresponding genes of Modoc virus (MOD). This latter virus belongs to the cluster of the “not-known vector” flaviviruses and is, unlike YF17D, neuroinvasive in SCID mice. Replication of in vitro-transcribed RNA from this chimeric flavivirus was shown by [3H]uridine labeling and RNA analysis. Expression of the MOD prM and E proteins was monitored by radioimmunoprecipitation and revealed that the MOD proteins were correctly and efficiently produced from the chimeric precursor protein. The MOD E protein was shown to be N-linked glycosylated, whereas prM, as predicted from the genome sequence, did not contain N-linked carbohydrates. In Vero cells, the chimeric virus replicated with a similar efficiency as the parental viruses, although it formed smaller plaques than YF17D and MOD. In SCID mice that had been infected intraperitoneally with the chimeric virus, the viral load increased steadily until death. The MOD/YF virus, like MOD from which it had acquired the prM and E structural proteins, but unlike YF, proved neuroinvasive in SCID mice. Animals developed neurological symptoms about 15 days after inoculation and died shortly thereafter. The distribution of MOD/YF RNA in the brain of infected mice was similar to that observed in MOD-infected mice. The observations provide compelling evidence that the determinants of neuroinvasiveness of flaviviruses are entirely located in the envelope proteins prM and E.

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