Structure and assembly of a paramyxovirus matrix protein

Battisti, Anthony J.; Meng, Geng; Winkler, Dennis C.; McGinnes, Lori W.; Plevka, Pavel; Steven, Alasdair C.; Morrison, Trudy G.; Rossmann, Michael G.

doi:10.1073/pnas.1210275109

Cited by 126 publications

(173 citation statements)

References 32 publications

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“…It is known that the contacts and, thus, the relative angles between dimeric matrix proteins determine the curvature of the matrix protein layer that forms underneath the cell membrane during budding (23). Our two crystal structures show that the dimerization interface itself also shows plasticity.…”

Section: Discussionmentioning

confidence: 72%

“…RSV belongs to the order Mononegavirales. Crystal structures have been solved for a number of M proteins from the Mononegavirales (21)(22)(23)(24), all of which form dimers. Although the monomeric unit of RSV M protein is structurally closely related to that of other paramyxovirus M proteins that exist as dimers, M was crystallized as a monomer (24,25), which sparked a debate in the field whether RSV M may use a different mechanism for its oligomerization.…”

mentioning

confidence: 99%

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Dimerization of Matrix Protein Is Required for Budding of Respiratory Syncytial Virus

Förster

Maertens

Farrell

et al. 2015

J Virol

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Respiratory syncytial virus (RSV) infects epithelial cells of the respiratory tract and is a major cause of bronchiolitis and pneumonia in children and the elderly. The virus assembles and buds through the plasma membrane, forming elongated membrane filaments, but details of how this happens remain obscure. Oligomerization of the matrix protein (M) is a key step in the process of assembly and infectious virus production. In addition, it was suggested to affect the conformation of the fusion protein, the major current target for RSV antivirals, in the mature virus. The structure and assembly of M are thus key parameters in the RSV antiviral development strategy. The structure of RSV M was previously published as a monomer. Other paramyxovirus M proteins have been shown to dimerize, and biochemical data suggest that RSV M also dimerizes. Here, using size exclusion chromatography-multiangle laser light scattering, we show that the protein is dimeric in solution. We also crystallized M in two crystal forms and show that it assembles into equivalent dimers in both lattices. Dimerization interface mutations destabilize the M dimer in vitro. To assess the biological relevance of dimerization, we used confocal imaging to show that dimerization interface mutants of M fail to assemble into viral filaments on the plasma membrane. Additionally, budding and release of virus-like particles are prevented in M mutants that fail to form filaments. Importantly, we show that M is biologically active as a dimer and that the switch from M dimers to higher-order oligomers triggers viral filament assembly and virus production. IMPORTANCEHuman respiratory syncytial virus (RSV) is the most frequent cause of infantile bronchiolitis and pneumonia. The enormous burden of RSV makes it a major unmet target for a vaccine and antiviral drug therapy. Oligomerization of the matrix protein is a key step in the process of assembly and production of infectious virus, but the molecular mechanism of RSV assembly is still poorly understood. Here we show that the RSV matrix protein forms dimers in solution and in crystals; the dimer is essential for formation of higher-order oligomers. Destabilizing the dimer interface resulted in the loss of RSV filament formation and a lack of budding of virus-like particles. Importantly, our findings can potentially lead to new structure-based RSV inhibitors targeting the assembly process.H uman respiratory syncytial virus (RSV), an enveloped, nonsegmented negative-strand RNA paramyxovirus, is the major worldwide cause of bronchiolitis and pneumonia in infants and of morbidity and mortality in the elderly. Despite the large global impact of RSV infection, no specific antivirals or licensed vaccine is yet available (1, 2). Designing new antivirals requires detailed molecular and structural understanding of key steps in RSV replication.RSV encodes 11 proteins, with the 3 glycoproteins, the fusion protein (F), glycoprotein (G), and small hydrophobic protein (SH), being present in the viral envelope. The virion ...

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Section: Discussionmentioning

confidence: 72%

mentioning

confidence: 99%

Dimerization of Matrix Protein Is Required for Budding of Respiratory Syncytial Virus

Förster

Maertens

Farrell

et al. 2015

J Virol

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“…ilarities with NDV and hMPV M, speculated to form dimers as the basic biological unit for subsequent oligomerization (22,23). We decided to test the ability of WT and mutant M recombinant proteins to form dimers/higher-order oligomers.…”

Section: Resultsmentioning

confidence: 99%

“…7A) based on reference 23 indicates no real sequence conservation between RSV M (41) and most Mononegavirales M proteins (23,42,43). However, structural alignment of the area around the RSV M Thr205-Ser220 loop (Fig.…”

Section: Discussionmentioning

confidence: 99%

“…Based on structural alignment with the recently published structure of human metapneumovirus (hMPV) (22) and Newcastle disease virus (NDV) M protein (23), we hypothesized that RSV M forms structurally similar dimeric and oligomeric structures. Despite the sequence divergence of the NDV, hMPV, and RSV M proteins, their tertiary structure appears to be well conserved, and thus the mechanisms of M oligomerization may be similar (23).…”

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confidence: 99%

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The Thr205 Phosphorylation Site within Respiratory Syncytial Virus Matrix (M) Protein Modulates M Oligomerization and Virus Production

Bajorek

Caly

Tran

et al. 2014

J Virol

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Human respiratory syncytial virus (RSV) is the most common cause of bronchiolitis and pneumonia in infants and the elderly worldwide; however, there is no licensed RSV vaccine or effective drug treatment available. The RSV matrix (M) protein plays key roles in virus assembly and budding, but the protein interactions that govern budding of infectious virus are not known. In this study, we focus on M protein and identify a key phosphorylation site (Thr205) in M that is critical for RSV infectious virus production. Recombinant virus with a nonphosphorylatable alanine (Ala) residue at the site was markedly attenuated, whereas virus with a phosphomimetic aspartate (Asp) resulted in a nonviable virus which could only be recovered with an additional mutation in M (serine to asparagine at position 220), strongly implying that Thr205 is critical for viral infectivity. Experiments in vitro showed that mutation of Thr205 does not affect M stability or the ability to form dimers but implicate an effect on higherorder oligomer assembly. In transfected and infected cells, Asp substitution of Thr205 appeared to impair M oligomerization; typical filamentous structures still formed at the plasma membrane, but M assembly during the ensuing elongation process seemed to be impaired, resulting in shorter and more branched filaments as observed using electron microscopy (EM). Our data thus imply for the first time that M oligomerization, regulated by a negative charge at Thr205, may be critical to production of infectious RSV. IMPORTANCEWe show here for the first time that RSV M's role in virus assembly/release is strongly dependent on threonine 205 (Thr205), a consensus site for CK2, which appears to play a key regulatory role in modulating M oligomerization and association with virus filaments. Our analysis indicates that T205 mutations do not impair M dimerization or viruslike filament formation per se but rather the ability of M to assemble in ordered fashion on the viral filaments themselves. This appears to impact in turn upon the infectivity of released virus rather than on virus production or release itself. Thus, M oligomerization would appear to be a target of interest for the development of anti-RSV agents; further, the recombinant T205-substituted mutant viruses described here would appear to be the first RSV mutants affected in viral maturation to our knowledge and hence of considerable interest for vaccine approaches in the future.

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Nipah virus matrix protein: expert hacker of cellular machines

Watkinson

Lee

2016

FEBS Letters

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Nipah virus (NiV, Henipavirus) is a highly lethal emergent zoonotic paramyxovirus responsible for repeated human outbreaks of encephalitis in South East Asia. There are no approved vaccines or treatments, thus improved understanding of NiV biology is imperative. NiV matrix protein recruits a plethora of cellular machinery to scaffold and coordinate virion budding. Intriguingly, matrix also hijacks cellular trafficking and ubiquitination pathways to facilitate transient nuclear localization. While the biological significance of matrix nuclear localization for an otherwise cytoplasmic virus remains enigmatic, the molecular details have begun to be characterized, and are conserved among matrix proteins from divergent paramyxoviruses. Matrix protein appropriation of cellular machinery will be discussed in terms of its early nuclear targeting and later role in virion assembly.

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Structure and assembly of a paramyxovirus matrix protein

Cited by 126 publications

References 32 publications

Dimerization of Matrix Protein Is Required for Budding of Respiratory Syncytial Virus

Dimerization of Matrix Protein Is Required for Budding of Respiratory Syncytial Virus

The Thr205 Phosphorylation Site within Respiratory Syncytial Virus Matrix (M) Protein Modulates M Oligomerization and Virus Production

Nipah virus matrix protein: expert hacker of cellular machines

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