scholarly journals RNA Interference with Measles Virus N, P, and L mRNAs Efficiently Prevents and with Matrix Protein mRNA Enhances Viral Transcription

2006 ◽  
Vol 80 (12) ◽  
pp. 5951-5957 ◽  
Author(s):  
Thorsten Reuter ◽  
Benedikt Weissbrich ◽  
Sibylle Schneider-Schaulies ◽  
Jürgen Schneider-Schaulies
Keyword(s):  
Rna Interference ◽  
Cell Fusion ◽  
Measles Virus ◽  
Matrix Protein ◽  
Negative Regulator ◽  
Viral Gene ◽  
Genome Replication ◽  
Genomic Rnas ◽  
Individual Gene ◽  
Cell Cell

ABSTRACT In contrast to studies with genetically modified viruses, RNA interference allows the analysis of virus infections with identical viruses and posttranscriptional ablation of individual gene functions. Using RNase III-generated multiple short interfering RNAs (siRNAs) against the six measles virus genes, we found efficient downregulation of viral gene expression in general with siRNAs against the nucleocapsid (N), phosphoprotein (P), and polymerase (L) mRNAs, the translation products of which form the ribonucleoprotein (RNP) complex. Silencing of the RNP mRNAs was highly efficient in reducing viral messenger and genomic RNAs. siRNAs against the mRNAs for the hemagglutinin (H) and fusion (F) proteins reduced the extent of cell-cell fusion. Interestingly, siRNA-mediated knockdown of the matrix (M) protein not only enhanced cell-cell fusion but also increased the levels of both mRNAs and genomic RNA by a factor of 2 to 2.5 so that the genome-to-mRNA ratio was constant. These findings indicate that M acts as a negative regulator of viral polymerase activity, affecting mRNA transcription and genome replication to the same extent.

scholarly journals Altered Interaction of the Matrix Protein with the Cytoplasmic Tail of Hemagglutinin Modulates Measles Virus Growth by Affecting Virus Assembly and Cell-Cell Fusion

2007 ◽  
Vol 81 (13) ◽  
pp. 6827-6836 ◽  
Author(s):  
Maino Tahara ◽  
Makoto Takeda ◽  
Yusuke Yanagi
Keyword(s):  
Cell Fusion ◽  
Measles Virus ◽  
Matrix Protein ◽  
Cytoplasmic Tail ◽  
Vero Cells ◽  
M Protein ◽  
Virus Growth ◽  
The Matrix ◽  
H Protein ◽  
Cell Cell

ABSTRACT Clinical isolates of measles virus (MV) use signaling lymphocyte activation molecule (SLAM) as a cellular receptor, whereas vaccine and laboratory strains may utilize the ubiquitously expressed CD46 as an additional receptor. MVs also infect, albeit inefficiently, SLAM− cells, via a SLAM- and CD46-independent pathway. Our previous study with recombinant chimeric viruses revealed that not only the receptor-binding hemagglutinin (H) but also the matrix (M) protein of the Edmonston vaccine strain can confer on an MV clinical isolate the ability to grow well in SLAM− Vero cells. Two substitutions (P64S and E89K) in the M protein which are present in many vaccine strains were found to be responsible for the efficient growth of recombinant virus in Vero cells. Here we show that the P64S and E89K substitutions allow a strong interaction of the M protein with the cytoplasmic tail of the H protein, thereby enhancing the assembly of infectious particles in Vero cells. These substitutions, however, are not necessarily advantageous for MVs, as they inhibit SLAM-dependent cell-cell fusion, thus reducing virus growth in SLAM+ B-lymphoblastoid B95a cells. When the cytoplasmic tail of the H protein is deleted, a virus with an M protein possessing the P64S and E89K substitutions no longer grows well in Vero cells yet causes cell-cell fusion and replicates efficiently in B95a cells. These results reveal a novel mechanism of adaptation and attenuation of MV in which the altered interaction of the M protein with the cytoplasmic tail of the H protein modulates MV growth in different cell types.

scholarly journals F-Actin Modulates Measles Virus Cell-Cell Fusion and Assembly by Altering the Interaction between the Matrix Protein and the Cytoplasmic Tail of Hemagglutinin

2012 ◽  
Vol 87 (4) ◽  
pp. 1974-1984 ◽  
Author(s):  
H. Wakimoto ◽  
M. Shimodo ◽  
Y. Satoh ◽  
Y. Kitagawa ◽  
K. Takeuchi ◽  
...  
Keyword(s):  
Cell Fusion ◽  
Measles Virus ◽  
Matrix Protein ◽  
Cytoplasmic Tail ◽  
The Matrix ◽  
Cell Cell

scholarly journals Previously Unrecognized Amino Acid Substitutions in the Hemagglutinin and Fusion Proteins of Measles Virus Modulate Cell-Cell Fusion, Hemadsorption, Virus Growth, and Penetration Rate

2009 ◽  
Vol 83 (17) ◽  
pp. 8713-8721 ◽  
Author(s):  
Hiromi Okada ◽  
Masae Itoh ◽  
Kyosuke Nagata ◽  
Kaoru Takeuchi
Keyword(s):  
Amino Acid ◽  
Cell Fusion ◽  
Measles Virus ◽  
Vero Cells ◽  
Amino Acid Substitutions ◽  
Wild Type ◽  
Virus Growth ◽  
F Protein ◽  
H Protein ◽  
Cell Cell

ABSTRACT Wild-type measles virus (MV) isolated in B95a cells could be adapted to Vero cells after several blind passages. In this study, we have determined the complete nucleotide sequences of the genomes of the wild type (T11wild) and its Vero cell-adapted (T11Ve-23) MV strain and identified amino acid substitutions R516G, E271K, D439E and G464W (D439E/G464W), N481Y/H495R, and Y187H/L204F in the nucleocapsid, V, fusion (F), hemagglutinin (H), and large proteins, respectively. Expression of mutated H and F proteins from cDNA revealed that the H495R substitution, in addition to N481Y, in the H protein was necessary for the wild-type H protein to use CD46 efficiently as a receptor and that the G464W substitution in the F protein was important for enhanced cell-cell fusion. Recombinant wild-type MV strains harboring the F protein with the mutations D439E/G464W [F(D439E/G464W)] and/or H(N481Y/H495R) protein revealed that both mutated F and H proteins were required for efficient syncytium formation and virus growth in Vero cells. Interestingly, a recombinant wild-type MV strain harboring the H(N481Y/H495R) protein penetrated slowly into Vero cells, while a recombinant wild-type MV strain harboring both the F(D439E/G464W) and H(N481Y/H495R) proteins penetrated efficiently into Vero cells, indicating that the F(D439E/G464W) protein compensates for the inefficient penetration of a wild-type MV strain harboring the H(N481Y/H495R) protein. Thus, the F and H proteins synergistically function to ensure efficient wild-type MV growth in Vero cells.

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

2015 ◽  
Vol 90 (6) ◽  
pp. 2849-2857 ◽  
Author(s):  
Sergey G. Guryanov ◽  
Lassi Liljeroos ◽  
Prasad Kasaragod ◽  
Tommi Kajander ◽  
Sarah J. Butcher
Keyword(s):  
Measles Virus ◽  
Matrix Protein ◽  
Hydrophobic Interactions ◽  
Rna Virus ◽  
Viral Rna ◽  
Genome Replication ◽  
Tail Domain ◽  
Human Pathogen ◽  
Nucleoprotein N ◽  
Insight Into

ABSTRACTThe enveloped negative-stranded RNA virus measles virus (MeV) is an important human pathogen. The nucleoprotein (N0) 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 N0assembly onto the NC and the activity of the polymerase are regulated by the viral phosphoprotein (P). In this study, we pulled down an N01-408fragment lacking most of its C-terminal tail domain by several affinity-tagged, N-terminal P fragments to map the N0-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 N0binding peptide, P1-48, to the C terminus of an N021-408fragment lacking both the N-terminal peptide and the C-terminal tail of N protein to reconstitute and crystallize the N0-P complex. We solved the X-ray structure of the resulting N0-P chimeric protein at a resolution of 2.7 Å. The structure reveals the molecular details of the conserved N0-P interface and explains how P chaperones N0, preventing both self-assembly of N0and 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.

Intramolecular complementation of measles virus fusion protein stability confers cell-cell fusion activity at 37 °C

FEBS Letters ◽  
2014 ◽  
Vol 589 (1) ◽  
pp. 152-158 ◽  
Author(s):  
Yuto Satoh ◽  
Mitsuhiro Hirose ◽  
Hiroko Shogaki ◽  
Hiroshi Wakimoto ◽  
Yoshinori Kitagawa ◽  
...  
Keyword(s):  
Fusion Protein ◽  
Protein Stability ◽  
Cell Fusion ◽  
Measles Virus ◽  
Fusion Activity ◽  
Virus Fusion ◽  
Cell Cell

Inhibition of Measles Virus-Induced Cell—Cell Fusion with a Monoclonal Antibody Directed Against the Haemagglutinin

1987 ◽  
Vol 1 (1) ◽  
pp. 25-34 ◽  
Author(s):  
P. DE VRIES ◽  
R.S. VAN BINNENDIJK ◽  
P. VAN DER MAREL ◽  
E.C. BEUVERY ◽  
A.L. VAN WEZEL ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
Cell Fusion ◽  
Measles Virus ◽  
Cell Cell

scholarly journals BST2/Tetherin Overexpression Modulates Morbillivirus Glycoprotein Production to Inhibit Cell–Cell Fusion

Viruses ◽  
2019 ◽  
Vol 11 (8) ◽  
pp. 692 ◽  
Author(s):  
James T. Kelly ◽  
Stacey Human ◽  
Joseph Alderman ◽  
Fatoumatta Jobe ◽  
Leanne Logan ◽  
...  
Keyword(s):  
Cell Fusion ◽  
Measles Virus ◽  
Viral Fusion ◽  
Large Reduction ◽  
Glycosyl Phosphatidylinositol ◽  
Species Specific ◽  
Syncytia Formation ◽  
Cell Cell

The measles virus (MeV), a member of the genus Morbillivirus, is an established pathogen of humans. A key feature of morbilliviruses is their ability to spread by virus–cell and cell–cell fusion. The latter process, which leads to syncytia formation in vitro and in vivo, is driven by the viral fusion (F) and haemagglutinin (H) glycoproteins. In this study, we demonstrate that MeV glycoproteins are sensitive to inhibition by bone marrow stromal antigen 2 (BST2/Tetherin/CD317) proteins. BST2 overexpression causes a large reduction in MeV syncytia expansion. Using quantitative cell–cell fusion assays, immunolabeling, and biochemistry we further demonstrate that ectopically expressed BST2 directly inhibits MeV cell–cell fusion. This restriction is mediated by the targeting of the MeV H glycoprotein, but not other MeV proteins. Using truncation mutants, we further establish that the C-terminal glycosyl-phosphatidylinositol (GPI) anchor of BST2 is required for the restriction of MeV replication in vitro and cell–cell fusion. By extending our study to the ruminant morbillivirus peste des petits ruminants virus (PPRV) and its natural host, sheep, we also confirm this is a broad and cross-species specific phenotype.

scholarly journals M protein of subacute sclerosing panencephalitis virus, synergistically with the F protein, plays a crucial role in viral neuropathogenicity

2021 ◽  
Vol 102 (10) ◽  
Author(s):  
Yuto Satoh ◽  
Kurara Higuchi ◽  
Daichi Nishikawa ◽  
Hiroshi Wakimoto ◽  
Miho Konami ◽  
...  
Keyword(s):  
Cell Fusion ◽  
Persistent Infection ◽  
High Frequency ◽  
Measles Virus ◽  
Subacute Sclerosing Panencephalitis ◽  
M Protein ◽  
Fusion Activity ◽  
F Protein ◽  
Sspe Virus ◽  
Cell Cell

Subacute sclerosing panencephalitis (SSPE) is a rare fatal neurodegenerative disease caused by a measles virus (MV) variant, SSPE virus, that accumulates mutations during long-term persistent infection of the central nervous system (CNS). Clusters of mutations identified around the matrix (M) protein in many SSPE viruses suppress productive infectious particle release and accelerate cell–cell fusion, which are features of SSPE viruses. It was reported, however, that these defects of M protein function might not be correlated directly with promotion of neurovirulence, although they might enable establishment of persistent infection. Neuropathogenicity is closely related to the character of the viral fusion (F) protein, and amino acid substitution(s) in the F protein of some SSPE viruses confers F protein hyperfusogenicity, facilitating viral propagation in the CNS through cell–cell fusion and leading to neurovirulence. The F protein of an SSPE virus Kobe-1 strain, however, displayed only moderately enhanced fusion activity and required additional mutations in the M protein for neuropathogenicity in mice. We demonstrated here the mechanism for the M protein of the Kobe-1 strain supporting the fusion activity of the F protein and cooperatively inducing neurovirulence, even though each protein, independently, has no effect on virulence. The occurrence of SSPE has been estimated recently as one in several thousand in children who acquired measles under the age of 5 years, markedly higher than reported previously. The probability of a specific mutation (or mutations) occurring in the F protein conferring hyperfusogenicity and neuropathogenicity might not be sufficient to explain the high frequency of SSPE. The induction of neurovirulence by M protein synergistically with moderately fusogenic F protein could account for the high frequency of SSPE.

scholarly journals CD9-dependent regulation of Canine distemper virus-induced cell–cell fusion segregates with the extracellular domain of the haemagglutinin

2006 ◽  
Vol 87 (6) ◽  
pp. 1635-1642 ◽  
Author(s):  
K. Singethan ◽  
E. Topfstedt ◽  
S. Schubert ◽  
W. P. Duprex ◽  
B. K. Rima ◽  
...  
Keyword(s):  
Cell Fusion ◽  
Measles Virus ◽  
Canine Distemper Virus ◽  
Transmembrane Protein ◽  
Extracellular Domain ◽  
Canine Distemper ◽  
Viral Glycoprotein ◽  
F Protein ◽  
H Protein ◽  
Cell Cell

Antibodies to CD9, a member of the tetraspan transmembrane-protein family, selectively inhibit Canine distemper virus (CDV)-induced cell–cell fusion. Neither CDV-induced virus–cell fusion nor cell–cell fusion induced by the closely related morbillivirus Measles virus (MV) is affected by anti-CD9 antibodies. As CDV does not bind CD9, an unknown, indirect mechanism is responsible for the observed inhibition of cell–cell fusion. It was investigated whether this effect was restricted to only one viral glycoprotein, either the haemagglutinin (H) or the fusion (F) protein, which form a fusion complex on the surface of virions and infected cells, or whether it is dependent on both in transient co-transfection assays. The susceptibility to CD9 antibodies segregates with the H protein of CDV. By exchanging portions of the H proteins of CDV and MV, it was determined that the complete extracellular domain, including the predicted stem structure (stem 1, barrel strand 1 and stem 2) and globular head domain, of the CDV-H protein mediates the effect. This suggests that interaction of the CDV-H protein with an unknown cellular receptor(s) is regulated by CD9, rather than F protein-mediated membrane fusion.

scholarly journals Nipah Virus Matrix Protein Influences Fusogenicity and Is Essential for Particle Infectivity and Stability

2015 ◽  
Vol 90 (5) ◽  
pp. 2514-2522 ◽  
Author(s):  
Erik Dietzel ◽  
Larissa Kolesnikova ◽  
Bevan Sawatsky ◽  
Anja Heiner ◽  
Michael Weis ◽  
...  
Keyword(s):  
Life Cycle ◽  
Cell Fusion ◽  
Matrix Protein ◽  
Fluorescent Protein ◽  
Critical Role ◽  
Nipah Virus ◽  
M Protein ◽  
The Matrix ◽  
Cell Cell

ABSTRACTNipah virus (NiV) causes fatal encephalitic infections in humans. To characterize the role of the matrix (M) protein in the viral life cycle, we generated a reverse genetics system based on NiV strain Malaysia. Using an enhanced green fluorescent protein (eGFP)-expressing M protein-deleted NiV, we observed a slightly increased cell-cell fusion, slow replication kinetics, and significantly reduced peak titers compared to the parental virus. While increased amounts of viral proteins were found in the supernatant of cells infected with M-deleted NiV, the infectivity-to-particle ratio was more than 100-fold reduced, and the particles were less thermostable and of more irregular morphology. Taken together, our data demonstrate that the M protein is not absolutely required for the production of cell-free NiV but is necessary for proper assembly and release of stable infectious NiV particles.IMPORTANCEHenipaviruses cause a severe disease with high mortality in human patients. Therefore, these viruses can be studied only in biosafety level 4 (BSL-4) laboratories, making it more challenging to characterize their life cycle. Here we investigated the role of the Nipah virus matrix protein in virus-mediated cell-cell fusion and in the formation and release of newly produced particles. We found that even though low levels of infectious viruses are produced in the absence of the matrix protein, it is required for the release of highly infectious and stable particles. Fusogenicity of matrixless viruses was slightly enhanced, further demonstrating the critical role of this protein in different steps of Nipah virus spread.

Sign in / Sign up

Export Citation Format

Share Document