Unravelling the complexities of respiratory syncytial virus RNA synthesis

Cowton, Vanessa M.; McGivern, David R.; Fearns, Rachel

doi:10.1099/vir.0.81786-0

Cited by 106 publications

(112 citation statements)

References 133 publications

(165 reference statements)

Supporting

Mentioning

Contrasting

Unclassified

Order By: Relevance

“…3E). Our data showed that N mono copurified with GST-P and GST-P but not with GST-P [1][2][3][4][5][6][7][8][9][10], GST-P [1][2][3][4][5][6][7][8][9][10][11][12][13][14][15][16][17][18][19][20], GST-P , or GST-P . These results revealed that an N mono -specific binding site is located between amino acids 1 and 29 of P. It is noteworthy that residues 1 to 10, if not sufficient to interact with N mono , are required for the interaction.…”

Section: Resultsmentioning

confidence: 99%

Identification and Characterization of the Binding Site of the Respiratory Syncytial Virus Phosphoprotein to RNA-Free Nucleoprotein

Galloux

Gabiane

Sourimant

et al. 2015

J Virol

113

View full text Add to dashboard Cite

The RNA genome of respiratory syncytial virus (RSV) is constitutively encapsidated by the viral nucleoprotein N, thus forming a helical nucleocapsid. Polymerization of N along the genomic and antigenomic RNAs is concomitant to replication and requires the preservation of an unassembled monomeric nucleoprotein pool. To this end, and by analogy with Paramyxoviridae and Rhabdoviridae, it is expected that the viral phosphoprotein P acts as a chaperone protein, forming a soluble complex with the RNA-free form of N (N 0 -P complex). Here, we have engineered a mutant form of N that is monomeric, is unable to bind RNA, still interacts with P, and could thus mimic the N 0 monomer. We used this N mutant, designated N mono , as a substitute for N 0 in order to characterize the P regions involved in the N 0 -P complex formation. Using a series of P fragments, we determined by glutathione S-transferase (GST) pulldown assays that the N and C termini of P are able to interact with N mono . We analyzed the functional role of amino-terminal residues of P by site-directed mutagenesis, using an RSV polymerase activity assay based on a human RSV minireplicon, and found that several residues were critical for viral RNA synthesis. Using GST pulldown and surface plasmon resonance assays, we showed that these critical residues are involved in the interaction between P[1-40] peptide and N mono in vitro. Finally, we showed that overexpression of the peptide P[1-29] can inhibit the polymerase activity in the context of the RSV minireplicon, thus demonstrating that targeting the N 0 -P interaction could constitute a potential antiviral strategy. IMPORTANCERespiratory syncytial virus (RSV) is the leading cause of lower respiratory tract illness in infants. Since no vaccine or efficient antiviral treatment is available against RSV, it is essential to better understand how the viral machinery functions in order to develop new antiviral strategies. RSV phosphoprotein P, the main RNA polymerase cofactor, is believed to function as a chaperon protein, maintaining N as a nonassembled, RNA-free protein (N 0 ) competent for RNA encapsidation. In this paper, we provide the first evidence, to our knowledge, that the N terminus of P contains a domain that binds specifically to this RNA-free form of N. We further show that overexpression of a small peptide spanning this region of P can inhibit viral RNA synthesis. These findings extend our understanding of the function of RSV RNA polymerase and point to a new target for the development of drugs against this virus. T he human respiratory syncytial virus (HRSV) is the leading cause of severe respiratory tract infections in newborn children worldwide (1). HRSV infects close to 100% of infants within the first 2 years of life and is the main cause of bronchiolitis. It is also recognized as a significant cause of severe respiratory infections in the elderly. The virus belongs to the Mononegavirales order and constitutes the prototype virus of the Pneumovirus genus of the Paramyxoviridae family. As f...

show abstract

Section: Resultsmentioning

confidence: 99%

Identification and Characterization of the Binding Site of the Respiratory Syncytial Virus Phosphoprotein to RNA-Free Nucleoprotein

Galloux

Gabiane

Sourimant

et al. 2015

J Virol

113

View full text Add to dashboard Cite

show abstract

“…The genomic replication promoters of Rhabdoviridae and Pneumovirinae are short monopartite elements located entirely within the leader [33,49]. By contrast, the genomic replication promoters of the Paramyxovirinae subfamily are bipartite, consisting of a first promoter element located within the leader and a second promoter element contained within the nontranslated region of the first gene.…”

Section: Filovirus Replicationmentioning

confidence: 99%

“…The fourth nucleocapsid protein of respiratory syncytial virus, M2-1, acts in a later stage of transcription as a processivity factor [49]. mRNA editing-There are two overlapping ORFs within the EBOV GP gene and neither of these are able to encode the virion-associated GP.…”

Section: Filovirus Transcriptionmentioning

confidence: 99%

Filovirus Replication and Transcription

2007

View full text Add to dashboard Cite

The highly pathogenic filoviruses, Marburg and Ebola virus, belong to the nonsegmented negative-sense RNA viruses of the order Mononegavirales. The mode of replication and transcription is similar for these viruses. On one hand, the negative-sense RNA genome serves as a template for replication, to generate progeny genomes, and, on the other hand, for transcription, to produce mRNAs. Despite the similarities in the replication/transcription strategy, filoviruses have evolved structural and functional properties that are unique among the nonsegmented negativesense RNA viruses. Moreover, there are also striking differences in the replication and transcription mechanisms of Marburg and Ebola virus. This includes nucleocapsid formation, the structure of the genomic replication promoter, the protein requirement for transcription and the use of mRNA editing. In this article, the current knowledge of the replication and transcription strategy of Marburg and Ebola virus is reviewed, with focus on the observed differences.

show abstract

“…Detailed understanding of RSV biology is made difficult by the intimate coupling of transcription and replication activities. Furthermore, the RSV polymerase is large and complex, containing multiple domains and enzymatic activities that allow it to function and be regulated as a transcriptase (including mRNA capping activity) and replicase (6)(7)(8).…”

mentioning

confidence: 99%

The Interferon Type I/III Response to Respiratory Syncytial Virus Infection in Airway Epithelial Cells Can Be Attenuated or Amplified by Antiviral Treatment

McCutcheon

Jordan

Mawhorter

et al. 2016

J Virol

View full text Add to dashboard Cite

Human respiratory syncytial virus (RSV) is a single-stranded RNA virus that causes acute, and occasionally fatal, lower respiratory illness in young infants, the elderly, and immunocompromised patients. Therapeutic interventions able to cut short viral replication and quickly return the airways to normal function are needed. An understanding of antiviral activities and their effects on host defense mechanisms is important for the design of safe and effective therapy. We targeted functionally and temporally distinct steps within the viral life cycle using small-molecule RSV inhibitors and studied their antiviral activities and their effects on innate interferon responses of airway epithelial cells in vitro. H uman respiratory syncytial virus (RSV) is a single-stranded, negative-sense RNA virus belonging to the family Paramyxoviridae. Premature and very young infants are at the highest risk of having severe lower respiratory tract infections and, later, a higher incidence of chronic asthma (1). Older adults with chronic lung or heart disease and individuals with suppressed immune systems also often require medical care after RSV infection. The lack of long-lasting protection after primary infection contributes to the capacity of RSV to cause yearly outbreaks and has challenged vaccine development (2). Synagis, a monoclonal antibody (MAb) directed against the RSV fusion protein, has shown prophylactic utility, but its cost and intramuscular route of administration have limited its use to hospitalized, high-risk infants Ͻ2 years of age (2). The only therapeutic intervention for RSV infection currently available to patients is the use of ribavirin (3), but its use is also limited because of poor efficacy and teratogenicity and the requirement of an aerosol and/or intravenous (i.v.) mode of administration in hospital settings (4). New therapeutic interventions able to lower the viral load, decrease transmission, and prevent lower respiratory complications are needed.RSV infection is initiated by attachment of the viral G protein to the surface of airway epithelial (AE) cells (5,6). Following attachment, the viral F protein mediates fusion of the viral and cellular membranes, allowing the RSV ribonucleic protein (RNP) complex, comprised of the viral RNA genome encapsulated with nucleoprotein (N) and associated with the phosphoprotein (P) and RNA-dependent RNA polymerase (L), to enter the cytoplasm. The RNP complex performs viral transcription to produce capped and polyadenylated mRNAs and genome replication. Detailed understanding of RSV biology is made difficult by the intimate coupling of transcription and replication activities. Furthermore, the RSV polymerase is large and complex, containing multiple domains and enzymatic activities that allow it to function and be

show abstract

Unravelling the complexities of respiratory syncytial virus RNA synthesis

Cited by 106 publications

References 133 publications

Identification and Characterization of the Binding Site of the Respiratory Syncytial Virus Phosphoprotein to RNA-Free Nucleoprotein

Identification and Characterization of the Binding Site of the Respiratory Syncytial Virus Phosphoprotein to RNA-Free Nucleoprotein

Filovirus Replication and Transcription

The Interferon Type I/III Response to Respiratory Syncytial Virus Infection in Airway Epithelial Cells Can Be Attenuated or Amplified by Antiviral Treatment

Contact Info

Product

Resources

About