scholarly journals Decision letter: Efficient support of virus-like particle assembly by the HIV-1 packaging signal

2018 ◽  
Keyword(s):  
Packaging Signal ◽  
Particle Assembly ◽  
Virus Like Particle ◽  
Hiv 1

scholarly journals Efficient support of virus-like particle assembly by the HIV-1 packaging signal

eLife ◽  
2018 ◽  
Vol 7 ◽  
Author(s):  
Mauricio Comas-Garcia ◽  
Tomas Kroupa ◽  
Siddhartha AK Datta ◽  
Demetria P Harvin ◽  
Wei-Shau Hu ◽  
...  
Keyword(s):  
Virus Assembly ◽  
Genomic Rna ◽  
Packaging Signal ◽  
Structural Element ◽  
Particle Assembly ◽  
Genomic Rnas ◽  
Gag Protein ◽  
Virus Particles ◽  
Virus Like Particle ◽  
Hiv 1

The principal structural component of a retrovirus particle is the Gag protein. Retroviral genomic RNAs contain a ‘packaging signal’ (‘Ψ') and are packaged in virus particles with very high selectivity. However, if no genomic RNA is present, Gag assembles into particles containing cellular mRNA molecules. The mechanism by which genomic RNA is normally selected during virus assembly is not understood. We previously reported (<xref ref-type="bibr" rid="bib9">Comas-Garcia et al., 2017</xref>) that at physiological ionic strength, recombinant HIV-1 Gag binds with similar affinities to RNAs with or without Ψ, and proposed that genomic RNA is selectively packaged because binding to Ψ initiates particle assembly more efficiently than other RNAs. We now present data directly supporting this hypothesis. We also show that one or more short stretches of unpaired G residues are important elements of Ψ; Ψ may not be localized to a single structural element, but is probably distributed over >100 bases.

scholarly journals Author response: Efficient support of virus-like particle assembly by the HIV-1 packaging signal

2018 ◽  
Author(s):  
Mauricio Comas-Garcia ◽  
Tomas Kroupa ◽  
Siddhartha AK Datta ◽  
Demetria P Harvin ◽  
Wei-Shau Hu ◽  
...  
Keyword(s):  
Author Response ◽  
Packaging Signal ◽  
Particle Assembly ◽  
Virus Like Particle ◽  
Hiv 1

scholarly journals HIV-1 matrix protein repositioning in nucleocapsid region fails to confer virus-like particle assembly

Virology ◽  
2008 ◽  
Vol 378 (1) ◽  
pp. 97-104 ◽  
Author(s):  
Ching-Yuan Chang ◽  
Yu-Fen Chang ◽  
Shiu-Mei Wang ◽  
Ying-Tzu Tseng ◽  
Kuo-Jung Huang ◽  
...  
Keyword(s):  
Matrix Protein ◽  
Particle Assembly ◽  
Virus Like Particle ◽  
Hiv 1

scholarly journals HIV-1 Gag protein with or without p6 specifically dimerizes on the viral RNA packaging signal

2020 ◽  
Vol 295 (42) ◽  
pp. 14391-14401 ◽  
Author(s):  
Samantha Sarni ◽  
Banhi Biswas ◽  
Shuohui Liu ◽  
Erik D. Olson ◽  
Jonathan P. Kitzrow ◽  
...  
Keyword(s):  
Mammalian Cells ◽  
Viral Particle ◽  
Rna Binding ◽  
Packaging Signal ◽  
Independent Manner ◽  
Particle Assembly ◽  
Gag Protein ◽  
Viral Particle Assembly ◽  
Hiv 1 ◽  
Psi Rna

The HIV-1 Gag protein is responsible for genomic RNA (gRNA) packaging and immature viral particle assembly. Although the presence of gRNA in virions is required for viral infectivity, in its absence, Gag can assemble around cellular RNAs and form particles resembling gRNA-containing particles. When gRNA is expressed, it is selectively packaged despite the presence of excess host RNA, but how it is selectively packaged is not understood. Specific recognition of a gRNA packaging signal (Psi) has been proposed to stimulate the efficient nucleation of viral assembly. However, the heterogeneity of Gag–RNA interactions renders capturing this transient nucleation complex using traditional structural biology approaches challenging. Here, we used native MS to investigate RNA binding of wild-type (WT) Gag and Gag lacking the p6 domain (GagΔp6). Both proteins bind to Psi RNA primarily as dimers, but to a control RNA primarily as monomers. The dimeric complexes on Psi RNA require an intact dimer interface within Gag. GagΔp6 binds to Psi RNA with high specificity in vitro and also selectively packages gRNA in particles produced in mammalian cells. These studies provide direct support for the idea that Gag binding to Psi specifically promotes nucleation of Gag–Gag interactions at the early stages of immature viral particle assembly in a p6-independent manner.

scholarly journals HIV-1 Gag protein with or without p6 specifically dimerizes on the viral RNA packaging signal

2020 ◽  
Author(s):  
Samantha Sarni ◽  
Banhi Biswas ◽  
Shuohui Liu ◽  
Erik D. Olson ◽  
Jonathan P. Kitzrow ◽  
...  
Keyword(s):  
Mammalian Cells ◽  
Viral Particle ◽  
Rna Binding ◽  
Packaging Signal ◽  
Independent Manner ◽  
Particle Assembly ◽  
Gag Protein ◽  
Viral Particle Assembly ◽  
Hiv 1 ◽  
Psi Rna

AbstractThe HIV-1 Gag protein is responsible for genomic RNA (gRNA) packaging and immature viral particle assembly. While the presence of gRNA in virions is required for viral infectivity, in its absence, Gag can assemble around cellular RNAs and form particles resembling gRNA-containing particles. When gRNA is expressed, it is selectively packaged despite the presence of excess host RNA, but how it is selectively packaged is not understood. Specific recognition of a gRNA packaging signal (Psi) has been proposed to stimulate the efficient nucleation of viral assembly. However, the heterogeneity of Gag-RNA interactions renders capturing this transient nucleation complex using traditional structural biology approaches challenging. Here, we used native mass spectrometry to investigate RNA binding of wild-type Gag and Gag lacking the p6 domain (GagΔp6). Both proteins bind to Psi RNA primarily as dimers, but to a control RNA primarily as monomers. The dimeric complexes on Psi RNA require an intact dimer interface within Gag. GagΔp6 binds to Psi RNA with high specificity in vitro and also selectively packages gRNA in particles produced in mammalian cells. These studies provide direct support for the idea that Gag binding to Psi specifically nucleates Gag-Gag interactions at the early stages of immature viral particle assembly in a p6-independent manner.

scholarly journals Analysis of Human Immunodeficiency Virus Type 1 Gag Dimerization-Induced Assembly

2005 ◽  
Vol 79 (23) ◽  
pp. 14498-14506 ◽  
Author(s):  
Ayna Alfadhli ◽  
Tenzin Choesang Dhenub ◽  
Amelia Still ◽  
Eric Barklis
Keyword(s):  
Human Immunodeficiency Virus ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Protein Assembly ◽  
Particle Assembly ◽  
Gag Protein ◽  
Immunodeficiency Virus ◽  
Hiv 1 ◽  
Terminal Domains

ABSTRACT The nucleocapsid (NC) domains of retrovirus precursor Gag (PrGag) proteins play an essential role in virus assembly. Evidence suggests that NC binding to viral RNA promotes dimerization of PrGag capsid (CA) domains, which triggers assembly of CA N-terminal domains (NTDs) into hexamer rings that are interconnected by CA C-terminal domains. To examine the influence of dimerization on human immunodeficiency virus type 1 (HIV-1) Gag protein assembly in vitro, we analyzed the assembly properties of Gag proteins in which NC domains were replaced with cysteine residues that could be linked via chemical treatment. In accordance with the model that Gag protein pairing triggers assembly, we found that cysteine cross-linking or oxidation reagents induced the assembly of virus-like particles. However, efficient assembly also was observed to be temperature dependent or required the tethering of NTDs. Our results suggest a multistep pathway for HIV-1 Gag protein assembly. In the first step, Gag protein pairing through NC-RNA interactions or C-terminal cysteine linkage fosters dimerization. Next, a conformational change converts assembly-restricted dimers or small oligomers into assembly-competent ones. At the final stage, final particle assembly occurs, possibly through a set of larger intermediates.

Structure and Stability of Wild-type and Mutant RNA Internal Loops from the SL-1 Domain of the HIV-1 Packaging Signal

2002 ◽  
Vol 322 (3) ◽  
pp. 543-557 ◽  
Author(s):  
Jane Greatorex ◽  
José Gallego ◽  
Gabriele Varani ◽  
Andrew Lever
Keyword(s):  
Wild Type ◽  
Packaging Signal ◽  
Structure And Stability ◽  
Internal Loops ◽  
Hiv 1

scholarly journals Identification of a Structural Element in HIV-1 Gag Required for Virus Particle Assembly and Maturation

mBio ◽  
2018 ◽  
Vol 9 (5) ◽  
Author(s):  
Mariia Novikova ◽  
Lucas J. Adams ◽  
Juan Fontana ◽  
Anna T. Gres ◽  
Muthukumar Balasubramaniam ◽  
...  
Keyword(s):  
Virus Assembly ◽  
Critical Role ◽  
Replication Cycle ◽  
Structural Defects ◽  
Fold Axis ◽  
Structural Element ◽  
Particle Assembly ◽  
Compensatory Mutations ◽  
Hiv 1 ◽  
Selection Of

ABSTRACTLate in the HIV-1 replication cycle, the viral structural protein Gag is targeted to virus assembly sites at the plasma membrane of infected cells. The capsid (CA) domain of Gag plays a critical role in the formation of the hexameric Gag lattice in the immature virion, and, during particle release, CA is cleaved from the Gag precursor by the viral protease and forms the conical core of the mature virion. A highly conserved Pro-Pro-Ile-Pro (PPIP) motif (CA residues 122 to 125) [PPIP(122–125)] in a loop connecting CA helices 6 and 7 resides at a 3-fold axis formed by neighboring hexamers in the immature Gag lattice. In this study, we characterized the role of this PPIP(122–125) loop in HIV-1 assembly and maturation. While mutations P123A and P125A were relatively well tolerated, mutation of P122 and I124 significantly impaired virus release, caused Gag processing defects, and abolished infectivity. X-ray crystallography indicated that the P122A and I124A mutations induce subtle changes in the structure of the mature CA lattice which were permissive forin vitroassembly of CA tubes. Transmission electron microscopy and cryo-electron tomography demonstrated that the P122A and I124A mutations induce severe structural defects in the immature Gag lattice and abrogate conical core formation. Propagation of the P122A and I124A mutants in T-cell lines led to the selection of compensatory mutations within CA. Our findings demonstrate that the CA PPIP(122–125) loop comprises a structural element critical for the formation of the immature Gag lattice.IMPORTANCECapsid (CA) plays multiple roles in the HIV-1 replication cycle. CA-CA domain interactions are responsible for multimerization of the Gag polyprotein at virus assembly sites, and in the mature virion, CA monomers assemble into a conical core that encapsidates the viral RNA genome. Multiple CA regions that contribute to the assembly and release of HIV-1 particles have been mapped and investigated. Here, we identified and characterized a Pro-rich loop in CA that is important for the formation of the immature Gag lattice. Changes in this region disrupt viral production and abrogate the formation of infectious, mature virions. Propagation of the mutants in culture led to the selection of second-site compensatory mutations within CA. These results expand our knowledge of the assembly and maturation steps in the viral replication cycle and may be relevant for development of antiviral drugs targeting CA.

scholarly journals Oropouche virus glycoprotein topology and cellular requirements for virus-like particle assembly

2020 ◽  
Author(s):  
Natalia S. Barbosa ◽  
Luis L. P. daSilva ◽  
Colin M. Crump ◽  
Stephen C. Graham
Keyword(s):  
Virus Infection ◽  
South America ◽  
Particle Production ◽  
Febrile Illness ◽  
Etiological Agent ◽  
Particle Assembly ◽  
Virus Surface ◽  
Virus Like Particle ◽  
Surface Glycoproteins

AbstractOropouche virus (OROV; Genus: Orthobunyavirus) is the etiological agent of Oropouche fever, a debilitating febrile illness common in South America. To facilitate studies of OROV budding and assembly, we developed a system for producing OROV virus-like particles (VLPs). Using this system we show that the OROV surface glycoproteins Gn and Gc self-assemble to form VLPs independently of the non-structural protein NSm. Mature OROV Gn has two trans-membrane domains that are crucial for glycoprotein translocation to the Golgi complex and VLP production. Inhibition of Golgi function using the drugs brefeldin A and monensin inhibit VLP secretion, with monensin treatment leading to an increase in co-localisation of OROV glycoproteins with the cis-Golgi marker protein GM130. Infection studies have previously shown that the cellular Endosomal Sorting Complexes Required for Transport (ESCRT) machinery is recruited to Golgi membranes during OROV assembly and that ESCRT activity is required for virus secretion. We demonstrate that a dominant negative form of the ESCRT-associated ATPase VPS4 significantly reduces Gn secretion in our VLP assay. Proteasome inhibition using the drug MG132 also disrupts VLPs secretion, suggesting that ubiquitylation promotes ESCRT-mediated VLP release. Additionally, we observe co-localisation between OROV glycoproteins and a specific subset of fluorescently-tagged ESCRT-III components, providing the first insights into which ESCRT-III components are required for OROV assembly. Our in vitro assay for OROV VLP production has allowed us to define molecular interactions that promote OROV release and will facilitate future studies of orthobunyavirus assembly.ImportanceOropouche virus is the etiological agent of Oropouche fever, a debilitating febrile illness common in South America. The tripartite genome of this zoonotic virus is capable of reassortment and there have been multiple epidemics of Oropouche fever in South America over the last 50 years, making Oropouche virus infection a significant threat to public health. However, the molecular characteristics of this arbovirus are poorly understood. We have developed an in vitro virus-like particle production assay for Oropouche virus, allowing us to study the assembly and release of this dangerous pathogen without high-containment biosecurity. We determined the polyprotein sites that are cleaved to yield the mature Oropouche virus surface glycoproteins and characterised the cellular machinery required for glycoprotein secretion. Our study provides important insights into the molecular biology of Oropouche virus infection, in addition to presenting a robust virus-like particle production assay that should facilitate future functional and pharmacological inhibition studies.

Sign in / Sign up

Export Citation Format

Share Document