scholarly journals A Chimeric Lloviu Virus Minigenome System Reveals that the Bat-Derived Filovirus Replicates More Similarly to Ebolaviruses than Marburgviruses

Cell Reports ◽  
2018 ◽  
Vol 24 (10) ◽  
pp. 2573-2580.e4 ◽  
Author(s):  
Whitney A. Manhart ◽  
Jennifer R. Pacheco ◽  
Adam J. Hume ◽  
Tessa N. Cressey ◽  
Laure R. Deflubé ◽  
...  
Keyword(s):  
Minigenome System

Development of an Isavirus minigenome system to study the function of the pocket RNA‐binding domain of the viral nucleoprotein (NP) in salmon cells

2019 ◽  
Vol 43 (2) ◽  
pp. 197-206
Author(s):  
Daniela Toro‐Ascuy ◽  
Alvaro Santibañez ◽  
Victor Peña ◽  
Carolina Beltran‐Pavez ◽  
Luis Cottet ◽  
...  
Keyword(s):  
Rna Binding ◽  
Binding Domain ◽  
Rna Binding Domain ◽  
Minigenome System

scholarly journals Oligomerization of Mumps Virus Phosphoprotein

2015 ◽  
Vol 89 (21) ◽  
pp. 11002-11010 ◽  
Author(s):  
Adrian Pickar ◽  
Andrew Elson ◽  
Yang Yang ◽  
Pei Xu ◽  
Ming Luo ◽  
...  
Keyword(s):  
Rna Synthesis ◽  
Mutational Analysis ◽  
Mumps Virus ◽  
Viral Rna ◽  
Large Protein ◽  
Terminal Domain ◽  
P Protein ◽  
Minigenome System ◽  
Oligomerization Domain

ABSTRACTThe mumps virus (MuV) genome encodes a phosphoprotein (P) that is important for viral RNA synthesis. P forms the viral RNA-dependent RNA polymerase with the large protein (L). P also interacts with the viral nucleoprotein (NP) and self-associates to form a homotetramer. The P protein consists of three domains, the N-terminal domain (PN), the oligomerization domain (PO), and the C-terminal domain (PC). While PNis known to relax the NP-bound RNA genome, the roles of POand PCare not clear. In this study, we investigated the roles of POand PCin viral RNA synthesis using mutational analysis and a minigenome system. We found that PNand PCfunctions can betrans-complemented. However, this complementation requires PO, indicating that POis essential for P function. Using thistrans-complementation system, we found that P forms parallel dimers (PNto PNand PCto PC). Furthermore, we found that residues R231, K238, K253, and K260 in POare critical for P's functions. We identified PCto be the domain that interacts with L. These results provide structure-function insights into the role of MuV P.IMPORTANCEMuV, a paramyxovirus, is an important human pathogen. The P protein of MuV is critical for viral RNA synthesis. In this work, we established a novel minigenome system that allows the domains of P to be complemented intrans. Using this system, we confirmed that MuV P forms parallel dimers. An understanding of viral RNA synthesis will allow the design of better vaccines and the development of antivirals.

scholarly journals Mapping the domains on the phosphoprotein of bovine respiratory syncytial virus required for N–P and P–L interactions using a minigenome system

2001 ◽  
Vol 82 (4) ◽  
pp. 775-779 ◽  
Author(s):  
Sunil K. Khattar ◽  
Abdul S. Yunus ◽  
Siba K. Samal
Keyword(s):  
Amino Acids ◽  
Respiratory Syncytial Virus ◽  
Binding Sites ◽  
Deletion Mutant ◽  
Bovine Respiratory Syncytial Virus ◽  
Binding Domains ◽  
P Protein ◽  
L Proteins ◽  
Syncytial Virus ◽  
Minigenome System

The interaction of bovine respiratory syncytial virus (BRSV) phosphoprotein (P) with nucleocapsid (N) and large polymerase (L) proteins was investigated using an intracellular BRSV–CAT minigenome replication system. Coimmunoprecipitation assays using P-specific antiserum revealed that the P protein can form complexes with N and L proteins. Deletion mutant analysis of the P protein was performed to identify the regions of P protein that interact with N and L proteins. The results indicate that two independent N-binding sites exist on the P protein: an internal region of 161–180 amino acids and a C-terminal region of 221–241 amino acids. The L-binding site was mapped to a region of P protein encompassing amino acids 121–160. The data suggest that N and L protein binding domains on the P protein do not overlap.

scholarly journals High-Throughput Minigenome System for Identifying Small-Molecule Inhibitors of Ebola Virus Replication

2015 ◽  
Vol 1 (8) ◽  
pp. 380-387 ◽  
Author(s):  
Megan R. Edwards ◽  
Colette Pietzsch ◽  
Thibaut Vausselin ◽  
Megan L. Shaw ◽  
Alexander Bukreyev ◽  
...  
Keyword(s):  
Virus Replication ◽  
Small Molecule ◽  
High Throughput ◽  
Ebola Virus ◽  
Small Molecule Inhibitors ◽  
Minigenome System

scholarly journals Development of a T7-Independent MARV Minigenome System

Proceedings ◽  
2020 ◽  
Vol 50 (1) ◽  
pp. 27
Author(s):  
Bert Vanmechelen ◽  
Joren Stroobants ◽  
Kurt Vermeire ◽  
Piet Maes
Keyword(s):  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Virus Disease ◽  
Ectopic Expression ◽  
Cell Types ◽  
Marburg Virus ◽  
Antiviral Compound ◽  
Wide Range ◽  
Compound Screening ◽  
Minigenome System

Marburg virus (MARV) is the only known pathogenic filovirus that does not belong to the genus Ebolavirus. It causes a severe hemorrhagic fever that is associated with a high mortality rate (>80%). The potential for filoviruses to cause devastating outbreaks, in combination with the lack of licensed therapeutics and vaccines for Marburg virus disease, illustrates the need for more MARV research. However, research involving MARV is hindered by its dependency on access to high-containment laboratories. Virus alternatives such as minigenomes have proven to be a useful tool to study virus replication and transcription at lower biosafety levels, and can be used for antiviral compound screening. All currently available MARV minigenomes are dependent on the addition of an ectopic T7 RNA polymerase that can drive minigenome expression. While this allows for high expression levels, the ectopic expression of a T7 polymerase is not feasible in all cell types, and acts as a confounding factor in compound screening assays. We have developed an alternative MARV minigenome system that is controlled by an RNA polymerase II promoter, which is natively expressed in most mammalian cell types. We show here that this novel minigenome can be used in a wide range of cell types, and can be easily amended to a 96-well format to be used for high-throughput compound screening, thereby providing a valuable alternative to previously developed MARV minigenomes.

scholarly journals The Major Attenuating Mutations of the Respiratory Syncytial Virus Vaccine Candidate cpts530/1009 Specify Temperature-Sensitive Defects in Transcription and Replication and a Non-Temperature-Sensitive Alteration in mRNA Termination

1999 ◽  
Vol 73 (6) ◽  
pp. 5176-5180 ◽  
Author(s):  
Katalin Juhasz ◽  
Brian R. Murphy ◽  
Peter L. Collins
Keyword(s):  
Respiratory Syncytial Virus ◽  
Amino Acid ◽  
Virus Vaccine ◽  
Vaccine Candidate ◽  
Rna Replication ◽  
Temperature Sensitive ◽  
Virus Growth ◽  
Syncytial Virus ◽  
Respiratory Syncytial Virus Vaccine ◽  
Minigenome System

ABSTRACT The live-attenuated respiratory syncytial virus vaccine candidatecpts530/1009 was previously shown to contain two separate amino acid changes in the L protein, mutations 530 and 1009 (Phe-521→Leu and Met-1169→Val, respectively, according to the amino acid sequence of the L protein). Each mutation independently specifies temperature-sensitive (ts) and attenuation phenotypes. In this study, we examined the effects of these mutations on transcription and RNA replication, using complete infectious recombinant virus as well as a plasmid-based minireplicon system, the latter under conditions in which effects on replication and transcription are uncoupled. In comparison with recombinant wild-type virus, the 530 and 1009 viruses were partially restricted at 37°C for RNA replication, mRNA synthesis, and virus growth. The 1009 virus was partially restricted for RNA synthesis and virus growth even at 32°C, which suggested that the 1009 mutation has a non-ts component in addition to the ts component. Interestingly, the synthesis of polycistronic readthrough mRNAs was elevated 1.6- to 3.8-fold for the 1009 virus, and this defect was non-ts. Studies with the minigenome system showed that the 530 and 1009 mutations each directly affect both replication and transcription, that the effect on replication was marginally greater than on transcription for the 530 mutation, and that the increase in readthrough mRNA associated with the 1009 mutation also was observed with the minigenome system.

Sign in / Sign up

Export Citation Format

Share Document