Initiation and regulation of paramyxovirus transcription and replication

Noton, Sarah L.; Fearns, Rachel

doi:10.1016/j.virol.2015.01.014

Cited by 114 publications

(138 citation statements)

References 96 publications

(112 reference statements)

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“…According to the current paramyxovirus models, both transcription and replication are initiated at the 3= end of the genomic RNA (30). The linear unidirectional organization of the "herringbone" NC means that the 3= and 5= ends of the NC do not present the same molecular surface due to this polarity.…”

Section: Discussionmentioning

confidence: 99%

Crystal Structure of the Measles Virus Nucleoprotein Core in Complex with an N-Terminal Region of Phosphoprotein

Guryanov

Liljeroos

Kasaragod

et al. 2016

J Virol

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The enveloped negative-stranded RNA virus measles virus (MeV) is an important human pathogen. The nucleoprotein (N 0 ) assembles with the viral RNA into helical ribonucleocapsids (NC) which are, in turn, coated by a helical layer of the matrix protein.The viral polymerase complex uses the NC as its template. The N 0 assembly onto the NC and the activity of the polymerase are regulated by the viral phosphoprotein (P). In this study, we pulled down an N 0 1-408 fragment lacking most of its C-terminal tail domain by several affinity-tagged, N-terminal P fragments to map the N 0 -binding region of P to the first 48 amino acids. We showed biochemically and using P mutants the importance of the hydrophobic interactions for the binding. We fused an N 0 binding peptide, P 1-48 , to the C terminus of an N 0 21-408 fragment lacking both the N-terminal peptide and the C-terminal tail of N protein to reconstitute and crystallize the N 0 -P complex. We solved the X-ray structure of the resulting N 0 -P chimeric protein at a resolution of 2.7 Å. The structure reveals the molecular details of the conserved N 0 -P interface and explains how P chaperones N 0 , preventing both self-assembly of N 0 and its binding to RNA. Finally, we propose a model for a preinitiation complex for RNA polymerization. IMPORTANCEMeasles virus is an important, highly contagious human pathogen. The nucleoprotein N binds only to viral genomic RNA and forms the helical ribonucleocapsid that serves as a template for viral replication. We address how N is regulated by another protein, the phosphoprotein (P), to prevent newly synthesized N from binding to cellular RNA. We describe the atomic model of an N-P complex and compare it to helical ribonucleocapsid. We thus provide insight into how P chaperones N and helps to start viral RNA synthesis. Our results provide a new insight into mechanisms of paramyxovirus replication. New data on the mechanisms of phosphoprotein chaperone action allows better understanding of virus genome replication and nucleocapsid assembly. We describe a conserved structural interface for the N-P interaction which could be a target for drug development to treat not only measles but also potentially other paramyxovirus diseases. Measles virus (MeV) belongs to the Paramyxoviridae family, which includes several other human pathogens, like respiratory syncytial (RSV), mumps, and parainfluenza viruses. It has a helical ribonucleocapsid (NC) containing a nonsegmented singlestrand RNA (ssRNA) genome wrapped around the outside the nucleoprotein (N) helix (1). The helical NC is active in both transcription and replication. During virus assembly, the matrix protein forms an additional helix covering the majority of the NC, potentially inhibiting transcription and promoting packaging into progeny virions (2). There are still only limited data on the detailed molecular interactions required to go from replication initiation to packaging of nascent RNA. The availability of N in a chaperoned, assembly-competent state with the phosphoprot...

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Section: Discussionmentioning

confidence: 99%

Crystal Structure of the Measles Virus Nucleoprotein Core in Complex with an N-Terminal Region of Phosphoprotein

Guryanov

Liljeroos

Kasaragod

et al. 2016

J Virol

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“…A more recently developed nucleoside analogue, marketed by Gilead Pharmaceuticals as Sovaldi, is now the basis of a cure for hepatitis C virus infection (350), with several more promising drugs and preparations in various stages of development. With regard to RSV therapeutic development, a robust in vitro RSV polymerase system (115,117,351) has been used to test nucleoside analogue libraries and other small-molecule inhibitors for their ability to inhibit the RSV polymerase complex. There is one nucleoside analogue named ALS-008176 (4=-chloromethyl-2=-deoxy-3=,5=-di-O-isobutyryl-2=-fluorocytidine) that was discovered using an RSV replicon readout system (352); it underwent successful clinical trials and was reported to inhibit RSV viral load by over 85% in human volunteers (102).…”

Section: Nucleoside Analogues and Small-molecule Inhibitorsmentioning

confidence: 99%

Respiratory Syncytial Virus: Infection, Detection, and New Options for Prevention and Treatment

2017

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SUMMARY Respiratory syncytial virus (RSV) infection is a significant cause of hospitalization of children in North America and one of the leading causes of death of infants less than 1 year of age worldwide, second only to malaria. Despite its global impact on human health, there are relatively few therapeutic options available to prevent or treat RSV infection. Paradoxically, there is a very large volume of information that is constantly being refined on RSV replication, the mechanisms of RSV-induced pathology, and community transmission. Compounding the burden of acute RSV infections is the exacerbation of preexisting chronic airway diseases and the chronic sequelae of RSV infection. A mechanistic link is even starting to emerge between asthma and those who suffer severe RSV infection early in childhood. In this article, we discuss developments in the understanding of RSV replication, pathogenesis, diagnostics, and therapeutics. We attempt to reconcile the large body of information on RSV and why after many clinical trials there is still no efficacious RSV vaccine and few therapeutics.

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“…The mechanism that drives the very same viral polymerase complex to either mRNA or genome synthesis, a process that ignores all cis-acting transcription start and stop signals encoded by the genome, is poorly understood. For other Mononegavirales, it is suggested that the amount of NP which encapsidates the nascent genomic and antigenomic RNA plays an important role in activating replication (11)(12)(13). It is conceivable that once replication of the viral genome is initiated, transcription and replication run in parallel.…”

Section: Importancementioning

confidence: 99%

Dynamic Phosphorylation of VP30 Is Essential for Ebola Virus Life Cycle

Biedenkopf

Lier

Becker

2016

J Virol

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Ebola virus is the causative agent of a severe fever with high fatality rates in humans and nonhuman primates. The regulation of Ebola virus transcription and replication currently is not well understood. An important factor regulating viral transcription is VP30, an Ebola virus-specific transcription factor associated with the viral nucleocapsid. Previous studies revealed that the phosphorylation status of VP30 impacts viral transcription. Together with NP, L, and the polymerase cofactor VP35, nonphosphorylated VP30 supports viral transcription. Upon VP30 phosphorylation, viral transcription ceases. Phosphorylation weakens the interaction between VP30 and the polymerase cofactor VP35 and/or the viral RNA. VP30 thereby is excluded from the viral transcription complex, simultaneously leading to increased viral replication which is supported by NP, L, and VP35 alone. Here, we use an infectious virus-like particle assay and recombinant viruses to show that the dynamic phosphorylation of VP30 is critical for the cotransport of VP30 with nucleocapsids to the sites of viral RNA synthesis, where VP30 is required to initiate primary viral transcription. We further demonstrate that a single serine residue at amino acid position 29 was sufficient to render VP30 active in primary transcription and to generate a recombinant virus with characteristics comparable to those of wild-type virus. In contrast, the rescue of a recombinant virus with a single serine at position 30 in VP30 was unsuccessful. Our results indicate critical roles for phosphorylated and dephosphorylated VP30 during the viral life cycle. IMPORTANCEThe current Ebola virus outbreak in West Africa has caused more than 28,000 cases and 11,000 fatalities. Very little is known regarding the molecular mechanisms of how the Ebola virus transcribes and replicates its genome. Previous investigations showed that the transcriptional support activity of VP30 is activated upon VP30 dephosphorylation. The current study reveals that the situation is more complex and that primary transcription as well as the rescue of recombinant Ebola virus also requires the transient phosphorylation of VP30. VP30 encodes six N-proximal serine residues that serve as phosphorylation acceptor sites. The present study shows that the dynamic phosphorylation of serine at position 29 alone is sufficient to activate primary viral transcription. Our results indicate a series of phosphorylation/dephosphorylation events that trigger binding to and release from the nucleocapsid and transcription complex to be essential for the full activity of VP30.

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Initiation and regulation of paramyxovirus transcription and replication

Cited by 114 publications

References 96 publications

Crystal Structure of the Measles Virus Nucleoprotein Core in Complex with an N-Terminal Region of Phosphoprotein

Crystal Structure of the Measles Virus Nucleoprotein Core in Complex with an N-Terminal Region of Phosphoprotein

Respiratory Syncytial Virus: Infection, Detection, and New Options for Prevention and Treatment

Dynamic Phosphorylation of VP30 Is Essential for Ebola Virus Life Cycle

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