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At least 19 recordsLinked to original sources
PubMed2004

Identification and partial characterization of Taastrup virus: a newly identified member species of the Mononegavirales.

We present a 8904-nt sequence of the central part of the RNA genome of a novel virus with a filovirus-like, nonidentical morphology named Taastrup virus (TV) detected in the leafhopper Psammotettix alienus. Sequence analysis identified five potential open reading frames (ORFs) and a complex pattern of homologies to various members of the Mononegavirales suggests a genome organization with the following order of genes: 3'-N-P-M-G-L-5'. Sequence analyses reveal an unusually large glycoprotein (G) containing both potential O-linked (14) and N-linked (9) glycosylation sites-a feature shared with the glycoproteins of Filoviridae and Pneumovirinae, and a nucleoprotein (N) with homology to the nucleoprotein of viral hemorrhagic septicemia virus (VHSV), a member of the Rhabdoviridae. Highly conserved domains were identified in the RNA-dependent RNA polymerase (L) between TV and other viruses within the order of Mononegavirales, and homology was found in particular with members of the Rhabdoviridae. The sequence similarities and the unique filovirus-like but nonidentical morphology unambiguously refer this newly identified virus to the order of Mononegavirales but to no family more than any, to other within the order.

Amino Acid Sequence↗
PubMed2006

RNA polymerase II-controlled expression of antigenomic RNA enhances the rescue efficacies of two different members of the Mononegavirales independently of the site of viral genome replication.

De novo generation of negative-strand RNA viruses depends on the efficient expression of antigenomic RNA (cRNA) from cDNA. To improve the rescue system of Borna disease virus (BDV), a member of the Mononegavirales with a nuclear replication phase, we evaluated different RNA polymerase (Pol) promoters for viral cRNA expression. Human and mouse Pol I promoters did not increase the recovery rate of infectious BDV from cDNA compared to the originally employed T7 RNA polymerase system. In contrast, expression of viral cRNA under the control of an RNA Pol II promoter increased the rescue efficacy by nearly 20-fold. Similarly, rescue of measles virus (MV), a member of the Mononegavirales with a cytoplasmic replication phase, was strongly improved by Pol II-controlled expression of viral cRNA. Analysis of transcription levels derived from different promoters suggested that the rescue-enhancing function of the Pol II promoter was due mainly to enhanced cRNA synthesis from the plasmid. Remarkably, correct 5'-terminal processing of Pol II-transcribed cRNA by a hammerhead ribozyme was not necessary for efficient rescue of BDV or MV. The correct 5' termini were reconstituted during replication of the artificially prolonged cRNA, indicating that the BDV and MV replicase complexes are able to recognize internal viral replication signals.

Borna disease virus↗
PubMed2002

Viral RNA-polymerases -- a predicted 2'-O-ribose methyltransferase domain shared by all Mononegavirales.

The Mononegavirales virus group comprises several major human pathogens, including measles, rabies and Ebola viruses. This article reports a computational analysis of the C-terminal region of RNA-dependent RNA-polymerases from Mononegavirales. Using a combination of sequence similarity and threading analysis, a 2'-O-ribose methyltransferase domain was identified that is involved in the capping of viral mRNAs.

Amino Acid Sequence↗
PubMed2004

Reverse genetics of mononegavirales.

"Reverse genetics" or de novo synthesis of nonsegmented negative-sense RNA viruses (Mononegavirales) from cloned cDNA has become a reliable technique to study this group of medically important viruses. Since the first generation of a negative-sense RNA virus entirely from cDNA in 1994, reverse genetics systems have been established for members of most genera of the Rhabdo-, Paramyxo-, and Filoviridae families. These systems are based on intracellular transcription of viral full-length RNAs and simultaneous expression of viral proteins required to form the typical viral ribonucleoprotein complex (RNP). These systems are powerful tools to study all aspects of the virus life cycle as well as the roles of virus proteins in virus-host interplay and pathogenicity. In addition, recombinant viruses can be designed to have specific properties that make them attractive as biotechnological tools and live vaccines.

Animals↗
PubMed1999

Host cell protein kinases in nonsegmented negative-strand virus (mononegavirales) infection.

Phosphorylation of one or more viral proteins is probably an essential step in the life cycle of every member of the nonsegmented negative-strand RNA virus (mononegavirales [MNV]) group. Since no virally encoded protein kinases have been discovered in this group, phosphorylation is effected entirely by host cell kinases. The virally encoded P proteins of the MNV are the only ones consistently phosphorylated with a stoichiometry > or =1. The P protein of vesicular stomatitis virus (VSV), and perhaps also of respiratory syncytial virus, are the only ones for which a function of phosphorylation has been established. Phosphorylation by casein kinase 2 at one or more identified sites in the VSV P protein activates transcriptional activity by promoting formation of a homotrimer, which is then capable of binding the RNA polymerase and attaching it to the N protein-RNA template. A second phosphorylation of VSV P protein by a different kinase also occurs, dependent upon prior modification by casein kinase 2, but its function is not definitely known. Phosphorylation of the other MNV P proteins may serve a different purpose. No evidence has been obtained yet for any function for phosphorylation of any other MNV protein.

Casein Kinases↗
PubMed1998

Sendai virus C proteins are categorically nonessential gene products but silencing their expression severely impairs viral replication and pathogenesis.

BACKGROUND: The P/C mRNA of Sendai virus (SeV), a prototypic member of the family Paramyxoviridae in the Mononegavirales superfamily comprising a large number of nonsegmented negative strand RNA viruses, encodes a nested set of accessory proteins, C', C, Y1 and Y2, referred to collectively as C proteins, initiating, respectively, at ACG/81 and AUGs/114, 183, 201 in the +1 frame relative to the ORF of phospho (P) protein, the smaller subunit of RNA polymerase. Among them, C is the major species expressed in infected cells at a molar ratio which is several-fold higher than the other three. However, their function has remained an enigma. It has not even been established whether or not the C proteins are essential for viral replication. Many other viruses in Mononegavirales encode C-like proteins, but their roles also remain to be defined. RESULTS: By taking advantage of a recently developed reverse genetics system to recover infectious SeV from cDNA, we created mutants in which C protein frames were variously silenced. C/C'(-) viruses which did not express C and C', but did express Y1 and Y2, were severely attenuated in replication in tissue culture cells of various species and tissues, as well as in embryonated chicken eggs. More notably, they were almost totally incapable of growing productively in--and hence nonpathogenic for mice--the natural host. Both gene expression and genome replication appeared to be impaired in C/C'(-) viruses. Additionally silencing the Y1 and Y2 expression was also possible, and a critically impaired but viable clone, the 4C(-) virus, was isolated which expressed none of the four C proteins. CONCLUSION: SeV C proteins are categorically nonessential gene products, but greatly contribute to full replication capability in vitro and are indispensable for in vivo multiplication and pathogenesis. This study represents the first comprehensive functional assessment of the accessary C protein for Mononegavirales.

Animals↗
PubMed1999

Phenotypic consequences of rearranging the P, M, and G genes of vesicular stomatitis virus.

The nonsegmented negative-strand RNA viruses (order Mononegavirales) include many important human pathogens. The order of their genes, which is highly conserved, is the major determinant of the relative levels of gene expression, since genes that are close to the single promoter site at the 3' end of the viral genome are transcribed at higher levels than those that occupy more distal positions. We manipulated an infectious cDNA clone of the prototypic vesicular stomatitis virus (VSV) to rearrange three of the five viral genes, using an approach which left the viral nucleotide sequence otherwise unaltered. The central three genes in the gene order, which encode the phosphoprotein P, the matrix protein M, and the glycoprotein G, were rearranged into all six possible orders. Viable viruses were recovered from each of the rearranged cDNAs. The recovered viruses were examined for their levels of gene expression, growth potential in cell culture, and virulence in mice. Gene rearrangement changed the expression levels of the encoded proteins in concordance with their distance from the 3' promoter. Some of the viruses with rearranged genomes replicated as well or slightly better than wild-type virus in cultured cells, while others showed decreased replication. All of the viruses were lethal for mice, although the time to symptoms and death following inoculation varied. These data show that despite the highly conserved gene order of the Mononegavirales, gene rearrangement is not lethal or necessarily even detrimental to the virus. These findings suggest that the conservation of the gene order observed among the Mononegavirales may result from immobilization of the ancestral gene order due to the lack of a mechanism for homologous recombination in this group of viruses. As a consequence, gene rearrangement should be irreversible and provide an approach for constructing viruses with novel phenotypes.

Animals↗
PubMed2004

Transcription and replication of nonsegmented negative-strand RNA viruses.

The nonsegmented negative-strand (NNS) RNA viruses of the order Mononegavirales include a wide variety of human, animal, and plant pathogens. The NNS RNA genomes of these viruses are templates for two distinct RNA synthetic processes: transcription to generate mRNAs and replication of the genome via production of a positive-sense antigenome that acts as template to generate progeny negative-strand genomes. The four virus families within the Mononegavirales all express the information encoded in their genomes by transcription of discrete subgenomic mRNAs. The key feature of transcriptional control in the NNS RNA viruses is entry of the virus-encoded RNA-dependent RNA polymerase at a single 3' proximal site followed by obligatory sequential transcription of the linear array of genes. Levels of gene expression are primarily regulated by position of each gene relative to the single promoter and also by cis-acting sequences located at the beginning and end of each gene and at the intergenic junctions. Obligatory sequential transcription dictates that termination of each upstream gene is required for initiation of downstream genes. Therefore, termination is a means to regulate expression of individual genes within the framework of a single transcriptional promoter. By engineering either whole virus genomes or subgenomic replicon derivatives, elements important for signaling transcript initiation, 5' end modification, 3' end polyadenylation, and transcription termination have been identified. Although the diverse families of NNS RNA virus use different sequences to control these processes, transcriptional termination is a common theme in controlling gene expression and overall transcriptional regulation is key in controlling the outcome of viral infection. The latest models for control of replication and transcription are discussed.

Gene Expression Regulation, Viral↗
PubMed2006

Structural disorder within the replicative complex of measles virus: functional implications.

Measles virus belongs to the Paramyxoviridae family within the Mononegavirales order. Its non-segmented, single stranded, negative sense RNA genome is encapsidated by the nucleoprotein (N) to form a helical nucleocapsid. This ribonucleoproteic complex is the substrate for both transcription and replication. The RNA-dependent RNA polymerase binds to the nucleocapsid template via its co-factor, the phosphoprotein (P). In this review, we summarize the main experimental data pointing out the abundance of structural disorder within measles virus N and P. We also describe studies indicating that structural disorder is a widespread property in the replicative complex of Paramyxoviridae and, more generally, of Mononegavirales. The functional implications of structural disorder are also discussed. Finally, we propose a model where the flexibility of the disordered N and P domains allows the formation of a tripartite complex (N degrees-P-L) during replication, followed by the delivery of N monomers to the newly synthesized genomic RNA chain.

Amino Acid Motifs↗
PubMed1994

Sequence and genome organization of Borna disease virus.

We have previously demonstrated that Borna disease virus (BDV) has a negative nonsegmented single-stranded (NNS) RNA genome that replicates in the nucleus of infected cells. Here we report for the first time the cloning and complete sequence of the BDV genome. Our results revealed that BDV has a genomic organization similar to that of other members of the Mononegavirales order. We have identified five main open reading frames (ORFs). The largest ORF, V, is located closest to the 5' end in the BDV genome and, on the basis of strong homology with other NNS-RNA virus polymerases, is a member of the L-protein family. The intercistronic regions vary in length and nucleotide composition and contain putative transcriptional start and stop signals. BDV untranslated 3' and 5' RNA sequences resemble those of other NNS-RNA viruses. Using a set of overlapping probes across the BDV genome, we identified nine in vivo synthesized species of polyadenylated subgenomic RNAs complementary to the negative-strand RNA genome, including monocistronic transcripts corresponding to ORFs I, II, and IV, as well as six polycistronic polyadenylated BDV RNAs. Interestingly, although ORFs III and V were detected within polycistronic transcripts, their corresponding monocistronic transcripts were not detected. Our data indicate that BDV is a member of the Mononegavirales, specially related to the family Rhabdoviridae. However, in contrast to the rest of the NNS-RNA animal viruses, BDV replication and transcription occur in the nucleus of infected cells. These findings suggest a possible relationship between BDV and the plant rhabdoviruses, which also replicate and transcribe in the nucleus.

Amino Acid Sequence↗
PubMed2005

Novel insights into the regulation of the viral polymerase complex of neurotropic Borna disease virus.

Borna disease virus (BDV) genetic information is encoded in a highly condensed non-segmented RNA genome of negative polarity. Replication and transcription of the genome occurs in the nucleus, enabling the virus to employ the cellular splicing machinery to process primary transcripts and to regulate expression of viral gene products. BDV establishes a non-cytolytic, persistent infection that in animals is mainly restricted to neurons of the central nervous system. Based on these unique properties, BDV represents the prototype member of the virus family Bornaviridae in the order Mononegavirales. Analysis of molecular aspects of BDV replication has long been hampered by the lack of a reverse genetics system. Only recently, artificial BDV minigenomes permitted the reconstitution of the viral polymerase complex, allowing finally the recovery of BDV from cDNA. As in other families of the Mononegavirales, the active polymerase complex of BDV is composed of the polymerase (L), the nucleoprotein (N) and the phosphoprotein (P). In addition, the viral X protein was identified as potent negative regulator of polymerase activity. Protein interaction studies combined with minireplicon assays suggested that P is a central regulatory element of BDV replication that directs the assembly of the polymerase complex. Most intriguingly, BDV obtained from cDNA with variable genomic termini suggests a novel strategy for viral replication-control. BDV seems to restrict its propagation efficacy by defined 5' terminal trimming of genomic and antigenomic RNA molecules. This review will summarize these novel findings and will discuss them in the context of BDV neurotropism and persistence.

Animals↗
PubMed1997

Analysis of the large (L) protein gene of the porcine rubulavirus LPMV: identification of possible functional domains.

The complete nucleotide sequence of the porcine rubulavirus LPMV (La Piedad Michoacan virus) large (L) protein gene was determined and analysed. The L mRNA was found to span 6,786 nucleotides, containing one single large open reading frame (ORF), putatively encoding a polypeptide of 2,251 amino acids. By aligning the amino acid sequence of the LPMV L-protein with L-protein of a number of viruses belonging to the order mononegavirale, a high degree of similarity between the LPMV L-protein and other rubula virus L-proteins was demonstrated, extending through almost the whole protein. Additionally we could identify several regions as being highly conserved among all studied viruses of the order mononegavirale. The significance of these regions are discussed.

Amino Acid Sequence↗
PubMed1997

Borna disease virus P-protein is phosphorylated by protein kinase Cepsilon and casein kinase II.

Borna disease virus (BDV) is a newly classified nonsegmented negative-strand RNA virus (order of Mononegavirales) that persistently infects specific brain regions and circuits of warm-blooded animals to cause behavioral disturbances. Viruses within the order of Mononegavirales have phosphoproteins that typically serve as transcription factors and are modulated in functional activity through phosphorylation. To identify the kinases involved in BDV phosphoprotein (BDV-P) phosphorylation, in vitro phosphorylation assays were performed using recombinant phosphoprotein produced in Escherichia coli as substrate and cytoplasmic extracts from a rat glioma cell line (C6) or rat brain extracts as sources of kinase activity. These experiments revealed that BDV-P was phosphorylated predominantly by protein kinase C (PKC) and to a lesser extent by casein kinase II. Partial purification of the PKC from rat brain extract suggested that the BDV-P phosphorylating kinase is PKCepsilon. A role for PKC phosphorylation in vivo was confirmed by using the PKC-specific inhibitor GF109203X. Furthermore, peptide mapping studies indicated that BDV-P is phosphorylated at the same sites in vitro as it is in vivo. Mutational analysis identified Ser26 and Ser28 as sites for PKC phosphorylation and Ser70 and Ser86 as sites for casein kinase II phosphorylation. The anatomic distribution of PKCepsilon in the central nervous system may have implications for BDV neurotropism and pathogenesis.

Animals↗
PubMed2005

Functional characterization of the genomic promoter of borna disease virus (BDV): implications of 3'-terminal sequence heterogeneity for BDV persistence.

Borna disease virus (BDV) is an enveloped virus with a genome organization characteristic of Mononegavirales. However, based on its unique features, BDV is considered the prototypic member of a new virus family, Bornaviridae, within the order Mononegavirales. We have described the establishment of a reverse genetics system for the rescue of BDV RNA analogues, or minigenomes, that is based on the use of polymerase I/polymerase II. Using this BDV minigenome rescue system, we have examined the functional implications of the reported sequence heterogeneity found at the 5' and 3' termini of the BDV genome and also defined the minimal BDV genomic promoter within the 3'-terminal 25 nucleotides. Our results suggest that the accumulation of RNA genome species containing truncations of one to three nucleotides at their 3' termini may contribute to modulate BDV RNA replication and gene expression during long-term persistence.

3' Untranslated Regions↗
PubMed2006

Generation of measles virus with a segmented RNA genome.

Viruses classified in the order Mononegavirales have a single nonsegmented RNA molecule as the genome and employ similar strategies for genome replication and gene expression. Infectious particles of Measles virus (MeV), a member of the family Paramyxoviridae in the order Mononegavirales, with two or three RNA genome segments (2 seg- or 3 seg-MeV) were generated using a highly efficient reverse genetics system. All RNA segments of the viruses were designed to have authentic 3' and 5' self-complementary termini, similar to those of negative-stranded RNA viruses that intrinsically have multiple RNA genome segments. The 2 seg- and 3 seg-MeV were viable and replicated well in cultured cells. 3 seg-MeV could accommodate up to six additional transcriptional units, five of which were shown to be capable of expressing foreign proteins efficiently. These data indicate that the MeV genome can be segmented, providing an experimental insight into the divergence of the negative-stranded RNA viruses with nonsegmented or segmented RNA genomes. They also illustrate a new strategy to develop mononegavirus-derived vectors harboring multiple additional transcriptional units.

Animals↗
PubMed2002

Molecular biology of Borna disease virus and persistence.

Borna disease virus (BDV) causes central nervous system (CNS) disease that is frequently manifested by behavioral abnormalities. Recent evidence indicates that the natural host range and geographic distribution of BDV is wider than originally thought BDV has been molecularly characterized as a non-segmented, negative single-stranded (NNS) RNA virus. Its genome (ca 8.9 kb), the smallest among known NNS RNA viruses, has an organization similar to that of other members of the order Mononegavirales. BDV has the property, unique among known animal NNS RNA viruses, of a nuclear site for the replication and transcription of its genome. The nucleocytoplasmic transport of BDV macromolecules is an essential component of the life cycle of BDV. An overlap of transcription units and transcriptive signals, overlap of ORFs, transcriptional readthrough and RNA splicing regulate expression of the BDV compact genome. The concurrent use of such diversity of strategies for the regulation of virus gene expression is unique among known NNS RNA viruses. Moreover, BDV appears to have also an unusual assembly process. Based on its unique genetic and biological features, BDV is considered to be the prototypic member of a new virus family, Bornaviridae, within the order Mononegavirales. Therefore, the investigation of the molecular biology of BDV may provide new insights about the biology of mononegaviruses, which include important human pathogens.

Animals↗