Altered ATP Function of a Vesicular Stomatitis Virus Mutant Detected by Kinetic Analysis of the Transcriptase Using Phosphorylated Ribavirin

Fernandez-Larsson, Roberto

doi:10.1099/0022-1317-70-10-2791

Cited by 7 publications

(4 citation statements)

References 26 publications

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“…The polR mutation caused a nearly twofold increase in N mRNA synthesis relative to leader synthesis in vitro. This result confirms and extends earlier and more indirect evidence for this effect (12,24,25). More surprising, however, were the numbers obtained for N/leader molar ratios, i.e., 1.0 Ϯ 0.1 for wild-type virus and 1.9 Ϯ 0.1 for polR1 virus.…”

Section: Discussionsupporting

confidence: 92%

“…Our earlier studies established that polR viruses synthesize a normal complement of mRNAs in vitro in addition to readthrough transcripts of the leader-N gene junction. These studies, as well as those of others, suggested that polR products also contain a smaller proportion of leader molecules than do wild-type products (12,24,25). A possible change in the ratio of leader to N gene transcription as a result of the polR mutation might bear important implications for the current 3Ј end entry model of transcription in nonsegmented negative-strand viruses.…”

Section: Assay Of Leader N and Readthrough Transcript Synthesis By mentioning

confidence: 57%

“…Although equimolar synthesis of leader and N gene transcripts, as seen here with wild-type virus, is compatible with the leader-first model, there are good reasons for suggesting that internal initiation could well take place in this case also. What causes excess N mRNA synthesis in polR1 is a template-associated N protein mutation, not a change in the RNA sequence anywhere near a polymerase entry site (6,12,24). The creation of a new entry site simply as a result of this protein mutation is highly unlikely.…”

Section: Discussionmentioning

confidence: 99%

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Initiation of vesicular stomatitis virus mutant polR1 transcription internally at the N gene in vitro

Chuang

Perrault

1997

J Virol

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The vesicular stomatitis virus (VSV) polymerase is thought to initiate transcription of its genome by first copying a small leader RNA complementary to the 3 end of the template. The polR VSV mutants, in contrast to wild-type virus, frequently read through the leader termination site during transcription in vitro. To shed light on polymerase termination and reinitiation events at the crucial leader-N gene junction, we employed RNase protection assays to precisely measure molar ratios of leader, N, and readthrough transcript accumulation in vitro. Wild-type virus synthesized essentially equimolar amounts of leader and N transcripts, but, unexpectedly, the polR1 mutant yielded about twice as much N mRNA as leader (ratio of 1.9 ؎ 0.1). Primer extension assays ruled out an increase in abortive N transcript synthesis for polR1. Transcription entailed multiple rounds of synthesis, with transcript ratios remaining the same after 0.5 or 2 h of synthesis, ruling out a significant contribution from polymerases "pre-positioned" at the N gene. No significant degradation of either leader or N transcripts was observed after incubating purified products with virions. Our data lead us to conclude that transcription can initiate internally at the N gene, at least in the case of polR1 VSV. We propose, however, that productive internal initiation of transcription is a fundamental property of the VSV polymerase and that of related viruses. A model postulating two distinct polymerase complexes, one for leader synthesis and one for internal initiation, is presented.

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Section: Discussionsupporting

confidence: 92%

Section: Assay Of Leader N and Readthrough Transcript Synthesis By mentioning

confidence: 57%

Section: Discussionmentioning

confidence: 99%

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Initiation of vesicular stomatitis virus mutant polR1 transcription internally at the N gene in vitro

Chuang

Perrault

1997

J Virol

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“…Moreover, RBV was found to control host immunological responses by inhibiting the production of interleukin-10 by regulatory T cells ( Kobayashi et al, 2012 ). The blocking mechanism of ribavirin is believed to target the capping ( Goswami et al, 1979 ) and translation efficiency ( Toltzis and Huang, 1986 ) of viral mRNA, along with the direct reduction of the viral polymerase’s activity ( Wray et al, 1985 ; Fernandez-Larsson and Patterson, 1989 ). Ribavirin acts by interfering with the accuracy and effectiveness of polymerase replication.…”

Section: Recent Therapeutic Advances Against Hantavirusmentioning

confidence: 99%

Hantavirus: an overview and advancements in therapeutic approaches for infection

Afzal,

Ali,

Batool

et al. 2023

Front. Microbiol.

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Hantaviruses are a significant and emerging global public health threat, impacting more than 200,000 individuals worldwide each year. The single-stranded RNA viruses belong to the Hantaviridae family and are responsible for causing two acute febrile diseases in humans: Hantavirus pulmonary syndrome (HPS) and hemorrhagic fever with renal syndrome (HFRS). Currently, there are no licensed treatments or vaccines available globally for HTNV infection. Various candidate drugs have shown efficacy in increasing survival rates during the early stages of HTNV infection. Some of these drugs include lactoferrin, ribavirin, ETAR, favipiravir and vandetanib. Immunotherapy utilizing neutralizing antibodies (NAbs) generated from Hantavirus convalescent patients show efficacy against HTNV. Monoclonal antibodies such as MIB22 and JL16 have demonstrated effectiveness in protecting against HTNV infection. The development of vaccines and antivirals, used independently and/or in combination, is critical for elucidating hantaviral infections and the impact on public health. RNA interference (RNAi) arised as an emerging antiviral therapy, is a highly specific degrades RNA, with post-transcriptional mechanism using eukaryotic cells platform. That has demonstrated efficacy against a wide range of viruses, both in vitro and in vivo. Recent antiviral methods involve using small interfering RNA (siRNA) and other, immune-based therapies to target specific gene segments (S, M, or L) of the Hantavirus. This therapeutic approach enhances viral RNA clearance through the RNA interference process in Vero E6 cells or human lung microvascular endothelial cells. However, the use of siRNAs faces challenges due to their low biological stability and limited in vivo targeting ability. Despite their successful inhibition of Hantavirus replication in host cells, their antiviral efficacy may be hindered. In the current review, we focus on advances in therapeutic strategies, as antiviral medications, immune-based therapies and vaccine candidates aimed at enhancing the body’s ability to control the progression of Hantavirus infections, with the potential to reduce the risk of severe disease.

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Isolation and Characterization of Vesicular Stomatitis Virus PolR Revertants: Polymerase Readthrough of the Leader–N Gene Junction Is Linked to an ATP-Dependent Function

1997

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The switch from transcription to replication of the VSV genome is coupled to assembly of nascent chains and involves an unspecified change in the P-L polymerase complex when it reaches the leader-N gene junction. PoIR VSV mutants, in contrast to wild-type virus, read through this first gene junction at high frequency without concurrent assembly, and they show altered ATP requirements for transcription in vitro. The mutation(s) responsible for the poIR phenotype segregates to the N-RNA template fraction. We report here that both poIR1 and poIR2 mutants display severe growth restriction in mouse L cells but not in BHK cells. Four of six poIR1 revertant viruses, originating from rare plaques on L cells, showed wild-type characteristics for growth, readthrough of leader-N gene junction, and ATP utilization, while two showed partial and quantitatively parallel coreversion of all properties. Sequence analysis of N and P genes of poIR mutants and revertants provided proof that a single mutation in the N protein, Arg179 to His, is responsible for the poIR phenotype. PoIR1, but not poIR2, also displayed a phenotypically silent GA-to-GG change in the N-P intergenic dinucleotide sequence Five of six revertants retained the poIR1 N protein mutation and showed no change in their P gene. We conclude that the L protein likely contains second-site suppressors of the poIR phenotype, and we propose that the switch from transcription to replication is modulated by an ATP-dependent interaction between the template-associated N protein and the L subunit of the P L polymerase complex.

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Altered ATP Function of a Vesicular Stomatitis Virus Mutant Detected by Kinetic Analysis of the Transcriptase Using Phosphorylated Ribavirin

Cited by 7 publications

References 26 publications

Initiation of vesicular stomatitis virus mutant polR1 transcription internally at the N gene in vitro

Initiation of vesicular stomatitis virus mutant polR1 transcription internally at the N gene in vitro

Hantavirus: an overview and advancements in therapeutic approaches for infection

Isolation and Characterization of Vesicular Stomatitis Virus PolR Revertants: Polymerase Readthrough of the Leader–N Gene Junction Is Linked to an ATP-Dependent Function

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