Paramyxovirus RNA Synthesis and the Requirement for Hexamer Genome Length: the Rule of Six Revisited

Kolakofsky, Daniel; Pelet, Thierry; Garcin, Dominique; Hausmann, Stéphane; Curran, Joseph; Roux, Laurent

doi:10.1128/jvi.72.2.891-899.1998

Cited by 326 publications

(135 citation statements)

References 38 publications

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“…Nucleotide sequencing also demonstrated that a 126-nucleotide gag fragment remained after the 1.416 kb sequence was deleted, and as expected, the genome of the deletion mutant conformed to the paramyxovirus rule-of-six (Kolakofsky et al, 1998), which states that the total number of nucleotides in the genomic RNA must be a multiple of 6 to replicate efficiently. Because this truncated gag gene contained the target amplicon sequence for the gag mRNA-specific RT-qPCR assay, abundant mRNA signal was observed even as the mutant virus emerged and became a dominant species in the infected ferrets.…”

Section: Discussionsupporting

confidence: 69%

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Development of a duplex real-time RT-qPCR assay to monitor genome replication, gene expression and gene insert stability during in vivo replication of a prototype live attenuated canine distemper virus vector encoding SIV gag

Coleman

Wright

Wallace

et al. 2015

Journal of Virological Methods

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Advancement of new vaccines based on live viral vectors requires sensitive assays to analyze in vivo replication, gene expression and genetic stability. In this study, attenuated canine distemper virus (CDV) was used as a vaccine delivery vector and duplex 2-step quantitative real-time RT-PCR (RT-qPCR) assays specific for genomic RNA (gRNA) or mRNA have been developed that concurrently quantify coding sequences for the CDV nucleocapsid protein (N) and a foreign vaccine antigen (SIV Gag). These amplicons, which had detection limits of about 10 copies per PCR reaction, were used to show that abdominal cavity lymphoid tissues were a primary site of CDV vector replication in infected ferrets, and importantly, CDV gRNA or mRNA was undetectable in brain tissue. In addition, the gRNA duplex assay was adapted for monitoring foreign gene insert genetic stability during in vivo replication by analyzing the ratio of CDV N and SIV gag genomic RNA copies over the course of vector infection. This measurement was found to be a sensitive probe for assessing the in vivo genetic stability of the foreign gene insert.

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Section: Discussionsupporting

confidence: 69%

“…1A). Following gene insertion, the overall genome length followed the paramyxovirus rule-of-six (Kolakofsky et al, 1998).…”

Section: Cell Culture and Virusesmentioning

confidence: 99%

Development of a duplex real-time RT-qPCR assay to monitor genome replication, gene expression and gene insert stability during in vivo replication of a prototype live attenuated canine distemper virus vector encoding SIV gag

Coleman

Wright

Wallace

et al. 2015

Journal of Virological Methods

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“…1A, C). Both new chimeric constructs conformed to the "rule of six" (i.e., each genome length was an even multiple of six nucleotides), and both were designed so that the hexamer spacing of transcription signals was conserved (i.e., the first and last genes, N and L, initiate at the second hexamer nucleotide while all other genes initiate at the first hexamer nucleotide) (Kolakofsky et al, 1998).…”

Section: Functional Replacement Of the Hpiv3 Hn And F Proteins By Ebomentioning

confidence: 99%

Chimeric human parainfluenza virus bearing the Ebola virus glycoprotein as the sole surface protein is immunogenic and highly protective against Ebola virus challenge

et al. 2009

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We generated a new live-attenuated vaccine against Ebola virus (EBOV) based on a chimeric virus HPIV3/ΔF-HN/EboGP that contains the EBOV glycoprotein (GP) as the sole transmembrane envelope protein combined with the internal proteins of human parainfluenza virus type 3 (HPIV3). Electron microscopy analysis of the virus particles showed that they have an envelope and surface spikes resembling those of EBOV and a particle size and shape resembling those of HPIV3. When HPIV3/ΔF-HN/EboGP was inoculated via apical surface of an in vitro model of human ciliated airway epithelium, the virus was released from the apical surface; when applied to basolateral surface, the virus infected basolateral cells but did not spread through the tissue. Following intranasal (IN) inoculation of guinea pigs, scattered infected cells were detected in the lungs by immunohistochemistry, but infectious HPIV3/ΔF-HN/EboGP could not be recovered from the lungs, blood, or other tissues. Despite the attenuation, the virus was highly immunogenic, and a single IN dose completely protected the animals against a highly lethal intraperitoneal challenge of guinea pig-adapted EBOV.

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“…1. In order to maintain the genome length as a multiple of six -a requirement for efficient NDV replication [46] -additional nucleotides were introduced at the GE region. A seven-nucleotide Kozak sequence [47] for efficient translation on the upstream of ATG, two XbaI restriction enzyme sites on the upstream and downstream of gene were also introduced burgdorferi antigens.…”

Section: Generation Of Recombinant Ndvs Expressing B Burgdorferi Ospmentioning

confidence: 99%

“…Thus, the inserted gene would be expressed as a separate mRNA. Extra nucleotides were inserted at the end of the ospC or bmpA ORF transcription cassette to maintain the rule of 6 [46]. For efficient translation, a Kozak sequence [47] was also inserted before the start codon.…”

Section: Construction Of Recombinant Ndvs Expressing B Burgdorferi Amentioning

confidence: 99%

A host-restricted viral vector for antigen-specific immunization against Lyme disease pathogen

Xiao

Kumar

Yang

et al. 2011

Vaccine

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Newcastle disease virus (NDV) is an avian virus that is attenuated in primates and is a potential vaccine vector for human use. We evaluated NDV as a vector for expressing selected antigens of the Lyme disease pathogen Borrelia burgdorferi. A series of recombinant NDVs were generated that expressed intracellular or extracellular forms of two Borrelia burgdorferi antigens: namely, the basic membrane protein A (BmpA) and the outer surface protein C (OspC). Expression of the intracellular and extracellular forms of these antigens was confirmed in cultured chicken cells. C3H or Balb/C mice that were immunized intranasally with the NDV vectors mounted vigorous serum antibody responses against the NDV vector, but failed to mount a robust response against either the intracellular or extracellular forms of BmpA or OspC. In contrast, a single immunization of hamsters with the NDV vectors via the intranasal, intramuscular, or intraperitoneal route resulted in rapid and rigorous antibody responses against the intracellular or extracellular forms of BmpA and OspC. When groups of hamsters were separately inoculated with various NDV vectors and challenged with B. burgdorferi (108 cells/animal), immunization with vector expressing either intracellular or extracellular BmpA was associated with a significant reduction of the pathogen load in the joints. Taken together, our studies highlighted the importance of NDV as vaccine vector that can be used for simple yet effective immunization of hosts against bacterial infections including Lyme disease.

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Paramyxovirus RNA Synthesis and the Requirement for Hexamer Genome Length: the Rule of Six Revisited

Cited by 326 publications

References 38 publications

Development of a duplex real-time RT-qPCR assay to monitor genome replication, gene expression and gene insert stability during in vivo replication of a prototype live attenuated canine distemper virus vector encoding SIV gag

Development of a duplex real-time RT-qPCR assay to monitor genome replication, gene expression and gene insert stability during in vivo replication of a prototype live attenuated canine distemper virus vector encoding SIV gag

Chimeric human parainfluenza virus bearing the Ebola virus glycoprotein as the sole surface protein is immunogenic and highly protective against Ebola virus challenge

A host-restricted viral vector for antigen-specific immunization against Lyme disease pathogen

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