scholarly journals Matrix Protein of Rabies Virus Is Responsible for the Assembly and Budding of Bullet-Shaped Particles and Interacts with the Transmembrane Spike Glycoprotein G

1999 ◽  
Vol 73 (1) ◽  
pp. 242-250 ◽  
Author(s):  
Teshome Mebatsion ◽  
Frank Weiland ◽  
Karl-Klaus Conzelmann
Keyword(s):  
Rabies Virus ◽  
Matrix Protein ◽  
M Protein ◽  
Virus Budding ◽  
Glycoprotein G ◽  
Severe Impairment ◽  
Rnp Complex ◽  
Long Rod ◽  
Nucleoprotein N

ABSTRACT To elucidate the functions of rhabdovirus matrix (M) protein, we determined the localization of M in rabies virus (RV) and analyzed the properties of an M-deficient RV mutant. We provide evidence that M completely covers the ribonucleoprotein (RNP) coil and keeps it in a condensed form. As determined by cosedimentation experiments, not only the M-RNP complex but also M alone was found to interact specifically with the glycoprotein G. In contrast, an interaction of G with the nucleoprotein N or M-less RNP was not observed. In the absence of M, infectious particles were mainly cell associated and the yield of cell-free infectious virus was reduced by as much as 500,000-fold, demonstrating the crucial role of M in virus budding. Supernatants from cells infected with the M-deficient RV did not contain the typical bullet-shaped rhabdovirus particles but instead contained long, rod-shaped virions, demonstrating severe impairment of the virus formation process. Complementation with M protein expressed from plasmids rescued rhabdovirus formation. These results demonstrate the pivotal role of M protein in condensing and targeting the RNP to the plasma membrane as well as in incorporation of G protein into budding virions.

scholarly journals The Deoptimization of Rabies Virus Matrix Protein Impacts Viral Transcription and Replication

Viruses ◽  
2019 ◽  
Vol 12 (1) ◽  
pp. 4 ◽  
Author(s):  
Jun Luo ◽  
Yue Zhang ◽  
Qiong Zhang ◽  
Yuting Wu ◽  
Boyue Zhang ◽  
...  
Keyword(s):  
Rabies Virus ◽  
Matrix Protein ◽  
Early Stage ◽  
Viral Proteins ◽  
Codon Pair Bias ◽  
Codon Pair ◽  
Codon Deoptimization ◽  
Cell Spread ◽  
Transcription And Replication

Rabies virus (RABV) matrix (M) protein plays several important roles during RABV infection. Although previous studies have assessed the functions of M through gene rearrangements, this interferes with the position of other viral proteins. In this study, we attenuated M expression through deoptimizing its codon usage based on codon pair bias in RABV. This strategy more objectively clarifies the role of M during virus infection. Codon-deoptimized M inhibited RABV replication during the early stages of infection, but enhanced viral titers at later stages. Codon-deoptimized M also inhibited genome synthesis at early stage of infection and increased the RABV transcription rates. Attenuated M through codon deoptimization enhanced RABV glycoprotein expression following RABV infection in neuronal cells, but had no influence on the cell-to-cell spread of RABV. In addition, codon-deoptimized M virus induced higher levels of apoptosis compared to the parental RABV. These results indicate that codon-deoptimized M increases glycoprotein expression, providing a foundation for further investigation of the role of M during RABV infection.

scholarly journals Mutations in the PPPY Motif of Vesicular Stomatitis Virus Matrix Protein Reduce Virus Budding by Inhibiting a Late Step in Virion Release

2000 ◽  
Vol 74 (21) ◽  
pp. 9818-9827 ◽  
Author(s):  
Himangi R. Jayakar ◽  
K. Gopal Murti ◽  
Michael A. Whitt
Keyword(s):  
Vesicular Stomatitis Virus ◽  
Matrix Protein ◽  
Electron Microscopic Examination ◽  
M Protein ◽  
Virus Yield ◽  
Virus Budding ◽  
Electron Microscopic ◽  
Gag Proteins ◽  
Virion Release ◽  
Vesicular Stomatitis

ABSTRACT The N terminus of the matrix (M) protein of vesicular stomatitis virus (VSV) and of other rhabdoviruses contains a highly conserved PPPY sequence (or PY motif) similar to the late (L) domains in the Gag proteins of some retroviruses. These L domains in retroviral Gag proteins are required for efficient release of virus particles. In this report, we show that mutations in the PPPY sequence of the VSV M protein reduce virus yield by blocking a late stage in virus budding. We also observed a delay in the ability of mutant viruses to cause inhibition of host gene expression compared to wild-type (WT) VSV. The effect of PY mutations on virus budding appears to be due to a block at a stage just prior to virion release, since electron microscopic examination of PPPA mutant-infected cells showed a large number of assembled virions at the plasma membrane trapped in the process of budding. Deletion of the glycoprotein (G) in addition to these mutations further reduced the virus yield to less than 1% of WT levels, and very few particles were assembled at the cell surface. This observation suggested that G protein aids in the initial stage of budding, presumably during the formation of the bud site. Overall, our results confirm that the PPPY sequence of the VSV M protein possesses L domain activity analogous to that of the retroviral Gag proteins.

scholarly journals Genome Characterization of Bird-Related Rhabdoviruses Circulating in Africa

Viruses ◽  
2021 ◽  
Vol 13 (11) ◽  
pp. 2168
Author(s):  
Dong-Sheng Luo ◽  
Zhi-Jian Zhou ◽  
Xing-Yi Ge ◽  
Hervé Bourhy ◽  
Zheng-Li Shi ◽  
...  
Keyword(s):  
Matrix Protein ◽  
Genomic Structure ◽  
Bird Species ◽  
Open Reading Frames ◽  
P Gene ◽  
Glycoprotein G ◽  
Close Relationship ◽  
1960S And 1970S ◽  
The 1960S ◽  
Nucleoprotein N

Rhabdoviridae is the most diverse family of the negative, single-stranded RNA viruses, which includes 40 ecologically different genera that infect plants, insects, reptiles, fishes, and mammals, including humans, and birds. To date, only a few bird-related rhabdoviruses among the genera Sunrhavirus, Hapavirus, and Tupavirus have been described and analyzed at the molecular level. In this study, we characterized seven additional and previously unclassified rhabdoviruses, which were isolated from various bird species collected in Africa during the 1960s and 1970s. Based on the analysis of their genome sequences obtained by next generation sequencing, we observed a classical genomic structure, with the presence of the five canonical rhabdovirus genes, i.e., nucleoprotein (N), phosphoprotein (P), matrix protein (M), glycoprotein (G), and polymerase (L). In addition, different additional open reading frames which code putative proteins of unknown function were identified, with the common presence of the C and the SH proteins, within the P gene and between the M and G genes, respectively. Genetic comparisons and phylogenetic analysis demonstrated that these seven bird-related rhabdoviruses could be considered as putative new species within the genus Sunrhavirus, where they clustered into a single group (named Clade III), a companion to two other groups that encompass mainly insect-related viruses. The results of this study shed light on the high diversity of the rhabdoviruses circulating in birds, mainly in Africa. Their close relationship with other insect-related sunrhaviruses raise questions about their potential role and impact as arboviruses that affect bird communities.

scholarly journals Interaction of Human Parainfluenza Virus Type 3 Nucleoprotein with Matrix Protein Mediates Internal Viral Protein Assembly

2015 ◽  
Vol 90 (5) ◽  
pp. 2306-2315 ◽  
Author(s):  
Guangyuan Zhang ◽  
Yi Zhong ◽  
Yali Qin ◽  
Mingzhou Chen
Keyword(s):  
Virus Type ◽  
Matrix Protein ◽  
Viral Proteins ◽  
Parainfluenza Virus ◽  
M Protein ◽  
Content Type ◽  
Virion Assembly ◽  
Nucleoprotein N ◽  
Human Parainfluenza Virus ◽  
Type 3

ABSTRACTHuman parainfluenza virus type 3 (HPIV3) belongs to theParamyxoviridaefamily. Its three internal viral proteins, the nucleoprotein (N), the phosphoprotein (P), and the polymerase (L), form the ribonucleoprotein (RNP) complex, which encapsidates the viral genome and associates with the matrix protein (M) for virion assembly. We previously showed that the M protein expressed alone is sufficient to assemble and release virus-like particles (VLPs) and a mutant with the L305A point mutation in the M protein (ML305A) has a VLP formation ability similar to that of wild-type M protein. In addition, recombinant HPIV3 (rHPIV3) containing the ML305Amutation (rHPIV3-ML305A) could be successfully recovered. In the present study, we found that the titer of rHPIV3-ML305Awas at least 10-fold lower than the titer of rHPIV3. Using VLP incorporation and coimmunoprecipitation assays, we found that VLPs expressing the M protein (M-VLPs) can efficiently incorporate N and P via an N-M or P-M interaction and ML305A-VLPs had an ability to incorporate P via a P-M interaction similar to that of M-VLPs but were unable to incorporate N and no longer interacted with N. Furthermore, we found that the incorporation of P into ML305A-VLPs but not M-VLPs was inhibited in the presence of N. In addition, we provide evidence that the C-terminal region of P is involved in its interaction with both N and M and N binding to the C-terminal region of P inhibits the incorporation of P into ML305A-VLPs. Our findings provide new molecular details to support the idea that the N-M interaction and not the P-M interaction is critical for packaging N and P into infectious viral particles.IMPORTANCEHuman parainfluenza virus type 3 (HPIV3) is a nonsegmented, negative-sense, single-stranded RNA virus that belongs to theParamyxoviridaefamily and can cause lower respiratory tract infections in infants and young children as well as elderly or immunocompromised individuals. However, no effective vaccine has been developed or licensed. We used virus-like particle (VLP) incorporation and coimmunoprecipitation assays to determine how the M protein assembles internal viral proteins. We demonstrate that both nucleoprotein (N) and phosphoprotein (P) can incorporate into M-VLPs and N inhibits the M-P interaction via the binding of N to the C terminus of P. We also provide additional evidence that the N-M interaction but not the P-M interaction is critical for the regulation of HPIV3 assembly. Our studies provide a more complete characterization of HPIV3 virion assembly and substantiation that N interaction with M regulates internal viral organization.

scholarly journals Indigenous Wildlife Rabies in Taiwan: Ferret Badgers, a Long Term Terrestrial Reservoir

2017 ◽  
Vol 2017 ◽  
pp. 1-6 ◽  
Author(s):  
Yu-Ching Lan ◽  
Tzai-Hung Wen ◽  
Chao-chin Chang ◽  
Hsin-Fu Liu ◽  
Pei-Fen Lee ◽  
...  
Keyword(s):  
Rabies Virus ◽  
Temporal Dynamics ◽  
Animal Health ◽  
Glycoprotein G ◽  
Coalescent Approach ◽  
Coalescent Tree ◽  
Southern Taiwan ◽  
World Organization ◽  
Nucleoprotein N

The emerging disease of rabies was confirmed in Taiwan ferret badgers (FBs) and reported to the World Organization for Animal Health (OIE) on July 17, 2013. The spread of wildlife rabies can be related to neighborhood countries in Asia. The phylogenetic analysis was conducted by maximum likelihood (ML) methods and the Bayesian coalescent approach based on the glycoprotein (G) and nucleoprotein (N) genes. The phylogeographic and spatial temporal dynamics of viral transmission were determined by using SPREAD, QGIS. Therefore, the origin and the change with time of the viruses can be identified. Results showed the rabies virus of FB strains in Taiwan is a unique clade among other strains in Asia. According to the phylogeographic coalescent tree, three major genotypes of the FB rabies virus have circulated in three different geographical areas in Taiwan. Two genotypes have distributed into central and southern Taiwan between two ecological river barriers. The third genotype has been limited in southeastern Taiwan by the natural mountain barrier. The diversity of FB rabies viruses indicates that the biological profile of FBs could vary in different geographical areas in Taiwan. An enhanced surveillance system needs to be established near the currently identified natural barriers for early warnings of the rabies virus outbreak in Taiwan.

scholarly journals The Glycoprotein and the Matrix Protein of Rabies Virus Affect Pathogenicity by Regulating Viral Replication and Facilitating Cell-to-Cell Spread

2007 ◽  
Vol 82 (5) ◽  
pp. 2330-2338 ◽  
Author(s):  
Rojjanaporn Pulmanausahakul ◽  
Jianwei Li ◽  
Matthias J. Schnell ◽  
Bernhard Dietzschold
Keyword(s):  
Viral Replication ◽  
Rabies Virus ◽  
Matrix Protein ◽  
M Gene ◽  
Predominant Role ◽  
Highly Pathogenic ◽  
Recombinant Viruses ◽  
Glycoprotein G ◽  
The Matrix ◽  
Cell Spread

ABSTRACT While the glycoprotein (G) of rabies virus (RV) is known to play a predominant role in the pathogenesis of rabies, the function of the RV matrix protein (M) in RV pathogenicity is not completely clear. To further investigate the roles of these proteins in viral pathogenicity, we constructed chimeric recombinant viruses by exchanging the G and M genes of the attenuated SN strain with those of the highly pathogenic SB strain. Infection of mice with these chimeric viruses revealed a significant increase in the pathogenicity of the SN strain bearing the RV G from the pathogenic SB strain. Moreover, the pathogenicity was further increased when both G and M from SB were introduced into SN. Interestingly, the replacement of the G or M gene or both in SN by the corresponding genes of SB was associated with a significant decrease in the rate of viral replication and viral RNA synthesis. In addition, a chimeric SN virus bearing both the M and G genes from SB exhibited more efficient cell-to-cell spread than a chimeric SN virus in which only the G gene was replaced. Together, these data indicate that both G and M play an important role in RV pathogenesis by regulating virus replication and facilitating cell-to-cell spread.

scholarly journals Pathogenicity of Different Rabies Virus Variants Inversely Correlates with Apoptosis and Rabies Virus Glycoprotein Expression in Infected Primary Neuron Cultures

1999 ◽  
Vol 73 (1) ◽  
pp. 510-518 ◽  
Author(s):  
Kinjiro Morimoto ◽  
D. Craig Hooper ◽  
Sergei Spitsin ◽  
Hilary Koprowski ◽  
Bernhard Dietzschold
Keyword(s):  
Protein Expression ◽  
G Protein ◽  
Rabies Virus ◽  
Antigenic Site ◽  
Expression Levels ◽  
N Protein ◽  
Adult Mice ◽  
Glycoprotein G ◽  
Neuronal Processes ◽  
Nucleoprotein N

ABSTRACT The mouse-adapted rabies virus strain CVS-24 has stable variants, CVS-B2c and CVS-N2c, which differ greatly in their pathogenicity for normal adult mice and in their ability to infect nonneuronal cells. The glycoprotein (G protein), which has previously been implicated in rabies virus pathogenicity, shows substantial structural differences between these variants. Although prior studies have identified antigenic site III of the G protein as the major pathogenicity determinant, CVS-B2c and CVS-N2c do not vary at this site. The possibility that pathogenicity is inversely related to G protein expression levels is suggested by the finding that CVS-B2c, the less pathogenic variant, expresses at least fourfold-higher levels of G protein than CVS-N2c in infected neurons. Although there is some difference between CVS-B2c- and CVS-N2c-infected neurons in G protein mRNA expression levels, the differential expression of G protein appears to be largely determined by posttranslational mechanisms that affect G protein stability. Pulse-chase experiments indicated that the G protein of CVS-B2c is degraded more slowly than that of CVS-N2c. The accumulation of G protein correlated with the induction of programmed cell death in CVS-B2c-infected neurons. The extent of apoptosis was considerably lower in CVS-N2c-infected neurons, where G protein expression was minimal. While nucleoprotein (N protein) expression levels were similar in neurons infected with either variant, the transport of N protein into neuronal processes was strongly inhibited in CVS-B2c-infected cells. Thus, downregulation of G protein expression in neuronal cells evidently contributes to rabies virus pathogenesis by preventing apoptosis and the apparently associated failure of the axonal transport of N protein.

Rabies virus matrix protein targets host actin cytoskeleton: A Protein-Protein interaction analysis

2020 ◽  
Author(s):  
Fatemeh Zandi ◽  
Vahid Khalaj ◽  
Fatemeh Goshadrou ◽  
Anna Meyfour ◽  
Alireza Gholami ◽  
...  
Keyword(s):  
Rabies Virus ◽  
Matrix Protein ◽  
Molecular Mechanisms ◽  
Interaction Analysis ◽  
Super Resolution ◽  
M Protein ◽  
Docking Simulations ◽  
Cytoskeleton Organization ◽  
Rat Brainstem ◽  
Rabies Pathogenesis

Abstract Multifunctional matrix protein (M) of rabies virus (RABV) plays essential roles in the pathogenesis of rabies infection. Identification of M protein interacting partners in target hosts could help to elucidate the biological pathways and molecular mechanisms involved in the pathogenesis of this virus. In this study, two-dimensional Far-western blotting (2D-Far-WB) technique was applied to find possible matrix protein partners in the rat brainstem. Recombinant RABV M was expressed in Pichia pastoris (P. pastoris) and was partially purified. Subsequently, 2D-Far-WB determined six rat brainstem proteins interacted with recombinant M protein which were identified by mass spectrometry. Functional annotation by gene ontology analysis determined these proteins were involved in the regulation of synaptic transmission processes, metabolic process, and cell morphogenesis-cytoskeleton organization. The interaction of viral M protein with selected host proteins in mouse Neuro-2a cells infected with RABV was verified by super-resolution confocal microscopy. Molecular docking simulations also demonstrated the formation of RABV M complexes. However, further confirmation with co-immunoprecipitation (Co-IP), was only successful for M-actin cytoplasmic1 interaction. Totally, our study revealed actin cytoplasmic1 as a binding partner of M protein, which might have important role(s) in rabies pathogenesis.

scholarly journals A single amino acid mutation, R42A, in the Newcastle disease virus matrix protein abrogates its nuclear localization and attenuates viral replication and pathogenicity

2014 ◽  
Vol 95 (5) ◽  
pp. 1067-1073 ◽  
Author(s):  
Zhiqiang Duan ◽  
Juan Li ◽  
Jie Zhu ◽  
Jian Chen ◽  
Haixu Xu ◽  
...  
Keyword(s):  
Viral Replication ◽  
Newcastle Disease Virus ◽  
Disease Virus ◽  
Nuclear Localization ◽  
Newcastle Disease ◽  
Matrix Protein ◽  
M Protein ◽  
Single Amino Acid ◽  
Basic Residue

The Newcastle disease virus (NDV) matrix (M) protein is a highly basic and nucleocytoplasmic shuttling viral protein. Previous study has demonstrated that the N-terminal 100 aa of NDV M protein are somewhat acidic overall, but the remainder of the polypeptide is strongly basic. In this study, we investigated the role of the N-terminal basic residues in the subcellular localization of M protein and in the replication and pathogenicity of NDV. We found that mutation of the basic residue arginine (R) to alanine (A) at position 42 disrupted M’s nuclear localization. Moreover, a recombinant virus with R42A mutation in the M protein reduced viral replication in DF-1 cells and attenuated the virulence and pathogenicity of the virus in chickens. This is the first report to show that a basic residue mutation in the NDV M protein abrogates its nuclear localization and attenuates viral replication and pathogenicity.

scholarly journals Generation of Monoclonal Antibodies against Variable Epitopes of the M Protein of Rabies Virus

Viruses ◽  
2019 ◽  
Vol 11 (4) ◽  
pp. 375
Author(s):  
Liu ◽  
Zhao ◽  
He ◽  
Wang ◽  
Su ◽  
...  
Keyword(s):  
Monoclonal Antibodies ◽  
Rabies Virus ◽  
Western Blotting ◽  
Indirect Immunofluorescence ◽  
Matrix Protein ◽  
Amino Acid Sequences ◽  
Diagnostic Methods ◽  
M Protein ◽  
Particle Assembly ◽  
Novel Target

Rabies virus (RABV), the causative agent of rabies, is highly neurovirulent for warm-blooded animals with a mortality rate of up to 100%. The RABV matrix protein (M) is required for virus particle assembly and budding. However, little is known about antigenic differences in the M protein. In this study, five monoclonal antibodies (mAbs), designated 3B9, 4A1, 2B11, 2C1, and 4B11, against the RABV M protein were generated using a recombinant M protein. All five mAbs reacted with the CVS-11 strain but showed no reactivity against the HEP-Flury strain in indirect immunofluorescence and western blotting. The epitope targeted by these mAbs was further identified by peptide scanning using GST-fused peptides. The 25PPYDDD30 peptide was defined as the minimal linear epitope. Alignment of amino acid sequences and phylogenetic analysis of different RABV strains indicated that the variable epitope 25PPDGDD30 is only present in the HEP-Flury and variant Flury strains of clade III, while the other strains resembling ERA and SRVA9 within the clade had another variable epitope, 25PLDDDD30. A Y27D mutation within the epitope was found among the rest of the RABV strains distributed in different clades. However, a single D28G mutation eliminated the reactivity of these five mAbs. In addition, the mAbs were able to recognize wildtype RABV strain in indirect immunofluorescence and western blotting and detect RABV-infected brain tissue using immunohistochemistry. The newly established mAbs and identified epitope may facilitate future investigations in the structure and function of the M protein and the development of diagnostic methods for the detection of different RABV strains worldwide. Most importantly, the epitope recognized by the mAbs against M protein might serve as a novel target for the development of a vaccine targeting RABV virulent strains.

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