Translational Gymnastics on the Sendai Virus P/C mRNA

Curran, Joseph; Latorre, Patrizia; Kolakofsky, Daniel

doi:10.1006/smvy.1997.0138

Cited by 20 publications

(17 citation statements)

References 32 publications

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“…However, this cannot be ruled out. Alternatively, scanning-independent initiation of internal start sites has been reported, as exemplified by the mechanism of ribosomal shunt described for the Y1, Y2, and X proteins of Sendai virus (17). The finding that the HMPV M2-1 ORF is dispensable for virus recovery and replication in vitro contrasts sharply with previous results with HRSV, where it has not been possible to recover virus in which the integrity of the M2 protein was disrupted by mutation or deletion.…”

Section: Discussionmentioning

confidence: 86%

“…These changes did not affect amino acid coding by the M2-1 ORF. The ATG-to-ACG changes were not ideal because ACG has the potential to serve as a translational start site, as exemplified by the C ORF of Sendai virus (17), but the efficiency would be reduced compared to ATG, and these were the only changes possible at those loci that did not alter amino acid coding in the M2-1 ORF. In addition, the portion of the M2-2 ORF that is downstream of the M2-1 ORF was deleted, removing 152 nucleotides (HMPV nt 5289 to 5440).…”

Section: Resultsmentioning

confidence: 99%

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Deletion of M2 Gene Open Reading Frames 1 and 2 of Human Metapneumovirus: Effects on RNA Synthesis, Attenuation, and Immunogenicity

Buchholz

Biacchesi

Pham

et al. 2005

J Virol

128

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The M2 gene of human metapneumovirus (HMPV) contains two overlapping open reading frames (ORFs), M2-1 and M2-2. The expression of separate M2-1 and M2-2 proteins from these ORFs was confirmed, and recombinant HMPVs were recovered in which expression of M2-1 and M2-2 was ablated individually or together [r⌬M2-1, r⌬M2-2, and r⌬M2(1؉2)]. Each M2 mutant virus directed efficient multicycle growth in Vero cells. The ability to recover HMPV lacking M2-1 contrasts with human respiratory syncytial virus, for which M2-1 is an essential transcription factor. Expression of the downstream HMPV M2-2 ORF was not reduced when translation of the upstream M2-1 ORF was silenced, indicating that it is initiated separately. The r⌬M2-2 mutants exhibited a two-to fivefold increase in the accumulation of mRNA, normalized to the genome template, suggesting that M2-2 has a role in regulating RNA synthesis. Replication and immunogenicity were tested in hamsters. Animals infected intranasally with r⌬M2-1 or r⌬M2(1؉2) did not have recoverable virus in the lungs or nasal turbinates on days 3 or 5 postinfection and did not develop HMPV-neutralizing serum antibodies or resistance to HMPV challenge. Thus, M2-1 appears to be essential for significant virus replication in vivo. In animals infected with r⌬M2-2, virus was recovered from only 1 of 12 animals and only in the nasal turbinates on a single day. However, all of the animals developed a high titer of HMPV-neutralizing serum antibodies and were highly protected against challenge with wild-type HMPV. The HMPV r⌬M2-2 virus is a promising and highly attenuated HMPV vaccine candidate.Human metapneumovirus (HMPV) was first identified in 2001 in The Netherlands from infants and children with acute respiratory tract disease (38) and is now recognized to be worldwide in prevalence (22,41). HMPV resembles human respiratory syncytial virus (HRSV) with regard to disease signs and the ability to infect and cause disease in infants as well as in individuals of all ages (7,18,20,29,32,39,41). The contribution of HMPV to respiratory tract disease remains to be fully defined but appears to be sufficient to warrant the development of a vaccine, especially for the pediatric population. Reverse genetic systems were recently developed for HMPV, allowing the generation of infectious virus from cDNA and providing an important tool for characterizing HMPV biology and for designing live-attenuated HMPV vaccines (5, 25).HMPV has a negative-strand RNA genome of approximately 13 kb (4, 37). It has been classified, together with avian metapneumovirus, in the Metapneumovirus genus, Pneumovirus subfamily, Paramyxovirus family, of the order Mononegavirales. The Pneumovirus subfamily also contains the genus Pneumovirus, represented by HRSV. The Metapneumovirus gene order is N-P-M-F-M2-SH-G-L. By analogy to HRSV, the predicted HMPV proteins are the following: the nucleocapsid protein N, which encapsidates the RNA genome and, together with the phosphoprotein P and the RNA polymerase protein L, forms the ribonucleoprotein co...

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Section: Discussionmentioning

confidence: 86%

Section: Resultsmentioning

confidence: 99%

Deletion of M2 Gene Open Reading Frames 1 and 2 of Human Metapneumovirus: Effects on RNA Synthesis, Attenuation, and Immunogenicity

Buchholz

Biacchesi

Pham

et al. 2005

J Virol

128

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“…rSeV-[C⌬10-15/P⌬13-18]. A rSeV containing a 21-nucleotide-deleted genome (encompassing C protein residues 9 LeuLys-Leu-Arg-Gly-Arg-Arg 15 ) is not expected to be viable (2). We therefore restricted ourselves to the six-amino-acid aa deletion of C⌬10-15 (and P⌬13-18), since C⌬10-15 also inhibits vRNA synthesis as had C wt (Fig.…”

Section: Resultsmentioning

confidence: 99%

Longer and Shorter Forms of Sendai Virus C Proteins Play Different Roles in Modulating the Cellular Antiviral Response

Garcin

Curran

Itoh

et al. 2001

J Virol

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The Sendai virus (SeV) C gene codes for a nested set of four C proteins that carry out several functions, including the modulation of viral RNA synthesis and countering of the cellular antiviral response. Using mutant C genes (and in particular a C gene with a deletion of six amino acids present only in the larger pair of C proteins) and recombinant SeV carrying these mutant C genes, we find that the nested set of C proteins carry out a nested set of functions. All of the C proteins interdict interferon (IFN) signaling to IFN-stimulated genes (ISGs) and prevent pY701-Stat1 formation. However, only the larger C proteins can induce STAT1 instability, prevent IFN from inducing an antiviral state, or prevent programmed cell death. Remarkably, interdiction of IFN signaling to ISGs and the absence of pY701-Stat1 formation did not prevent IFN-␣ from inducing an anti-Vesicular stomatitis virus (VSV) state. It is possible that IFN-␣ signaling to induce an anti-VSV state can occur independently of the well-established Jak/Stat/ISGF3 pathway and that it is this parallel pathway that is targeted by the longer C proteins.Paramyxovirus P genes are remarkable for the complexity of their genetic organization and expression. This gene, named for the phosphoprotein that is an essential component of the P/L viral RNA polymerase (vRNAP), contains additional open reading frames (ORFs) that overlap the beginning and the middle of the P protein ORF (the C and V ORFs, respectively) ( Fig. 1). The C and V overlapping ORFs are accessed by a variety of unusual mechanisms of ribosomal choice (reviewed in references 9 and 31) and mRNA editing (39, 41), respectively. The overlapping ORFs are also referred to as "accessory" genes (1) because in each case there is at least one virus within the subfamily Paramyxovirinae that does not contain (or express) the V or C genes. For Sendai virus (SeV), a nested set of 2 longer C proteins (CЈ ACG81 and C AUG114) and 2 shorter C proteins (Y1 AUG183 and Y2 AUG201 ) are expressed (Fig. 1). The two longer C proteins (along with the P protein) are initiated by scanning ribosomes, whereas the two shorter C proteins are initiated by a ribosomal shunt (31).The first property of the multifunctional C proteins noted was their ability to modulate viral RNA (vRNA) synthesis. When vRNA synthesis is reconstituted in vitro with purified N:RNA nucleocapsids and the products of the P/C and L genes (expressed in transfected cell extracts), specific ablation of C gene expression (or its replacement with mutant C F170S genes) increases vRNAP specific activity ϳ8-fold (5). The longer CЈ or C proteins are equally active in this respect, but the shorter Y proteins are inactive. The inhibitory effects of C expression are promoter specific, in that genomic promoters are considerably more sensitive to inhibition than antigenomic promoters (1, 38). Remarkably, the negative effects of C on vRNA synthesis are accompanied by an equivalent increase in promoter fidelity, in that minigenomes with promoter mutations (including those wh...

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“…Most paramyxoviruses encode a number of accessory proteins from the P gene, either through RNA editing leading to interferon antagonist V and W families, or by translating alternate reading frames from the P mRNA (55) to generate C family members. While we propose that C proteins are critical for engagement with Rab11, we suggest that in the absence of sufficient accumulation of L protein, vRNPs would be unable to interact with Rab11.…”

Section: Discussionmentioning

confidence: 99%

The Viral Polymerase Complex Mediates the Interaction of vRNPs with Recycling Endosomes During SeV Assembly

Genoyer

Kulej

Hung

et al. 2020

Preprint

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31Paramyxoviruses are negative sense single-stranded RNA viruses that comprise many important 32 human and animal pathogens, including human parainfluenza viruses. These viruses bud from 33 the plasma membrane of infected cells after the viral ribonucleoprotein complex (vRNP) is 34 transported from the cytoplasm to the cell membrane via The viral proteins that are critical for mediating this important initial step in viral assembly are 36 unknown. Here we use the model paramyxovirus, murine parainfluenza virus 1, or Sendai virus 37 (SeV), to investigate the roles of viral proteins in Rab11a-driven virion assembly. We previously 38 reported that infection with SeV containing high levels of copy-back defective viral genomes 39 (DVGs) generates heterogenous populations of cells. Cells enriched in full-length virus produce 40 viral particles containing standard or defective viral genomes, while cells enriched in DVGs do 41 not, despite high levels of defective viral genome replication. Here we take advantage of this 42 heterogenous cell phenotype to identify proteins that mediate interaction of vRNPs with Rab11a. 43We examine the role of matrix protein and nucleoprotein and determine that they are not 44 sufficient to drive interaction of vRNPs with recycling endosomes. Using a combination of mass 45 spectrometry and comparative protein abundance and localization in DVG-and FL-high cells, 46we identify viral polymerase complex components L and, specifically, its cofactor C proteins as 47Paramyxoviruses are single-stranded negative sense RNA viruses that include viruses of 65 clinical and global health significance, such as human parainfluenza viruses (HPIV). HPIV1 and 66 2 are the leading cause of croup in young children and HPIV3 is associated with bronchiolitis, 67 bronchitis, and pneumonia (1). These infections have also been associated with the development 68 or exacerbation of asthma and chronic airway disorders (2). Additionally, HPIVs are the 69 etiological agent of 7% of hospitalized pneumonia cases in children in the U.S. (3), as well as in 70Africa and Asia (4). Although HPIVs pose a significant health burden, there are no direct acting 71 antivirals or preventative vaccines. In order to identify targets for antiviral development, there is 72 a need to better understand fundamental aspects of paramyxovirus biology, including 73 mechanisms of virion assembly and particle production. 74Sendai virus (SeV), or murine parainfluenza virus 1, is closely related to human 75 parainfluenza viruses 1 and 3 with 69% homology at the RNA level between SeV and HPIV1. 76SeV has long served as a well-established model for understanding paramyxovirus replication. 77SeV genome replication occurs in the cytoplasm of infected cells (5) and virions bud from the 78 plasma membrane. The viral genomes are tightly coated in the viral nucleocapsid protein (NP) 79 with one NP molecule for every six nucleotides of genome (6). This viral RNA-protein complex 80 is referred to as the viral ribonucleoprotein (vRNP) (7, 8). Viral genomes a...

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Translational Gymnastics on the Sendai Virus P/C mRNA

Cited by 20 publications

References 32 publications

Deletion of M2 Gene Open Reading Frames 1 and 2 of Human Metapneumovirus: Effects on RNA Synthesis, Attenuation, and Immunogenicity

Deletion of M2 Gene Open Reading Frames 1 and 2 of Human Metapneumovirus: Effects on RNA Synthesis, Attenuation, and Immunogenicity

Longer and Shorter Forms of Sendai Virus C Proteins Play Different Roles in Modulating the Cellular Antiviral Response

The Viral Polymerase Complex Mediates the Interaction of vRNPs with Recycling Endosomes During SeV Assembly

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