Murine leukemia virus pseudotypes of La Crosse and Hantaan Bunyaviruses: a system for analysis of cell tropism

Ma, Meihui; ., D.B. Kersten; Kamrud, Kurt I.; Wool-Lewis, Rouven J.; Schmaljohn, Connie S.; González‐Scarano, Francisco

doi:10.1016/s0168-1702(99)00070-2

Cited by 26 publications

(23 citation statements)

References 32 publications

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“…VSV⌬G*-HTN and -SEO were generated efficiently at titers in the range of 10 5 to 10 6 IU/ml, which are higher titers than those of native HTNV and SEOV (10 4 to 10 5 focus-forming units/ml). Another pseudotype system with murine leukemia virus has been previously reported to produce a hantavirus pseudotype; however, the titers in that system were much lower, i.e., in the range of 10 3 IU/ml (13). It was thought that these results reflect low expression levels of Hantaan envelope glycoproteins or inefficient virus assembly.…”

Section: Discussionmentioning

confidence: 96%

Use of Vesicular Stomatitis Virus Pseudotypes Bearing Hantaan or Seoul Virus Envelope Proteins in a Rapid and Safe Neutralization Test

Ogino

Ebihara

Lee

et al. 2003

Clin Vaccine Immunol

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A vesicular stomatitis virus (VSV) pseudotype bearing hantavirus envelope glycoproteins was produced and used in a neutralization test as a substitute for native hantavirus. The recombinant VSV, in which the enveloped protein gene (G) was replaced by the green fluorescent protein gene and complemented with G protein expressed in trans (VSV⌬G*G), was kindly provided by M. A. Whitt. 293T cells were transfected with plasmids for the expression of envelope glycoproteins (G1 and G2) of HTNV or SEOV and were then infected with VSV⌬G*G. Pseudotype VSV with the Hantaan (VSV⌬G*-HTN) or Seoul (VSV⌬G*-SEO) envelope glycoproteins were harvested from the culture supernatant. The number of infectious units (IU) of the pseudotype VSVs ranged from 10 5 to 10 6 /ml. The infectivity of VSV⌬G*-HTN and VSV⌬G*-SEO was neutralized with monoclonal antibodies, immune rabbit sera, and sera from patients with hemorrhagic fever with renal syndrome, and the neutralizing titers were similar to those obtained with native hantaviruses. These results show that VSV⌬G*-HTN and -SEO can be used as a rapid, specific, and safe neutralization test for detecting hantavirus-neutralizing antibodies as an effective substitute for the use of native hantaviruses. Furthermore, the IU of VSV⌬G*-HTN and -SEO did not decrease by more than 10-fold when stored at 4°C for up to 30 days.

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Section: Discussionmentioning

confidence: 96%

Use of Vesicular Stomatitis Virus Pseudotypes Bearing Hantaan or Seoul Virus Envelope Proteins in a Rapid and Safe Neutralization Test

Ogino

Ebihara

Lee

et al. 2003

Clin Vaccine Immunol

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“…The effects of mutations within LACV Gc 1066-1087 were assessed using conformational and non-conformational antibodies specific to Gc, a luciferase based cell-to-cell fusion assay, and pseudotype transduction assays (Ma et al, 1999) (Plassmeyer et al, 2005; Plassmeyer et al, 2007). Several mutations within this hydrophobic domain affected glycoprotein expression to some extent, but all mutations either shifted the pH threshold of fusion below that of the wild type protein, reduced fusion efficiency, or abrogated cell-to-cell fusion and pseudotype entry altogether.…”

Section: Introductionmentioning

confidence: 99%

La Crosse virus (LACV) Gc fusion peptide mutants have impaired growth and fusion phenotypes, but remain neurotoxic

et al. 2010

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La Crosse virus is a leading cause of pediatric encephalitis in the Midwestern United States and an emerging pathogen in the American South. The LACV glycoprotein Gc plays a critical role in entry as the virus attachment protein. A 22 amino acid hydrophobic region within Gc (1066-1087) was recently identified as the LACV fusion peptide. To further define the role of Gc (1066-1087) in virus entry, fusion, and neuropathogenesis, a panel of recombinant LACV (rLACV) fusion peptide mutant viruses was generated. Replication of mutant rLACVs was significantly reduced. In addition, the fusion peptide mutants demonstrated decreased fusion phenotypes relative to LACV-WT. Interestingly, these viruses maintained their ability to cause neuronal loss in culture, suggesting that the fusion peptide of LACV Gc is a determinant of properties associated with neuroinvasion (growth to high titer in muscle cells and a robust fusion phenotype), but not necessarily of neurovirulence.

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“…GP0 is cleaved in the Golgi apparatus by furin-like proteases to generate two subunits, GP1 and GP2, which are linked by a disulfide bond (17, 24, 25). The native form of GP is a homotrimer of GP1/GP2 heterodimer on virions (32).…”

mentioning

confidence: 99%

The role of the charged residues of the GP2 helical regions in Ebola entry

et al. 2009

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The glycoprotein (GP) of Ebola is the sole structural protein that forms the spikes on the viral envelope. The GP contains two subunits, GP1 and GP2, linked by a disulfide bond, which are responsible for receptor binding and membrane fusion, respectively. In this study, the full length of GP gene of Ebola Zaire species, 2028 base pairs in length, was synthesized using 38 overlapping oligonucleotides by multiple rounds of polymerase chain reaction (PCR). The synthesized GP gene was shown to be efficiently expressed in mammalian cells. Furthermore, an efficient HIV-based pseudotyping system was developed using the synthetic GP gene, providing a safe approach to dissecting the entry mechanism of Ebola viruses. Using this pseudotyping system and mutational analysis, the role of the charged residues in the GP2 helical regions was examined. It was found that substitutions of the most charged residues in the regions did not adversely affect GP expression, processing, or viral incorporation, however, most of the mutations greatly impaired the ability of GP to mediate efficient viral infection. These results demonstrate that these charged residues of GP2 play an important role in GP-mediated Ebola entry into its host cells. We propose that these charged residues are involved in forming the intermediate conformation(s) of GP in membrane fusion and Ebola entry.

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Murine leukemia virus pseudotypes of La Crosse and Hantaan Bunyaviruses: a system for analysis of cell tropism

Cited by 26 publications

References 32 publications

Use of Vesicular Stomatitis Virus Pseudotypes Bearing Hantaan or Seoul Virus Envelope Proteins in a Rapid and Safe Neutralization Test

Use of Vesicular Stomatitis Virus Pseudotypes Bearing Hantaan or Seoul Virus Envelope Proteins in a Rapid and Safe Neutralization Test

La Crosse virus (LACV) Gc fusion peptide mutants have impaired growth and fusion phenotypes, but remain neurotoxic

The role of the charged residues of the GP2 helical regions in Ebola entry

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