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Lassa virus Z protein is a matrix protein and sufficient for the release of virus-like particles [corrected].

ABSTRACT: Lassa virus is an enveloped virus with glycoprotein spikes on its surface. It contains an RNA ambisense genome that encodes the glycoprotein precursor GP-C, the nucleoprotein NP, the polymerase L, and the Z protein. Here we demonstrate that the Lassa virus Z protein (i). is abundant in viral particles, (ii). is strongly membrane associated, (iii). is sufficient in the absence of all other viral proteins to release enveloped particles, and (iv). contains two late domains, PTAP and PPXY, necessary for the release of virus-like particles. Our data provide evidence that Z is the Lassa virus matrix protein that is the driving force for virus particle release.

SUBMITTER: Strecker T

PROVIDER: S-EPMC228511 | biostudies-literature | 2003 Oct

REPOSITORIES: biostudies-literature

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Lassa virus Z protein is a matrix protein and sufficient for the release of virus-like particles [corrected].

Strecker Thomas T Eichler Robert R Meulen Jan ter Jt Weissenhorn Winfried W Dieter Klenk Hans H Garten Wolfgang W Lenz Oliver O

Journal of virology 20031001 19

Lassa virus is an enveloped virus with glycoprotein spikes on its surface. It contains an RNA ambisense genome that encodes the glycoprotein precursor GP-C, the nucleoprotein NP, the polymerase L, and the Z protein. Here we demonstrate that the Lassa virus Z protein (i). is abundant in viral particles, (ii). is strongly membrane associated, (iii). is sufficient in the absence of all other viral proteins to release enveloped particles, and (iv). contains two late domains, PTAP and PPXY, necessary ...[more]

PMID: 12970458

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Project description:BackgroundLassa fever is a neglected tropical disease with significant impact on the health care system, society, and economy of Western and Central African nations where it is endemic. Treatment of acute Lassa fever infections has successfully utilized intravenous administration of ribavirin, a nucleotide analogue drug, but this is not an approved use; efficacy of oral administration has not been demonstrated. To date, several potential new vaccine platforms have been explored, but none have progressed toward clinical trials and commercialization. Therefore, the development of a robust vaccine platform that could be generated in sufficient quantities and at a low cost per dose could herald a subcontinent-wide vaccination program. This would move Lassa endemic areas toward the control and reduction of major outbreaks and endemic infections. To this end, we have employed efficient mammalian expression systems to generate a Lassa virus (LASV)-like particle (VLP)-based modular vaccine platform.ResultsA mammalian expression system that generated large quantities of LASV VLP in human cells at small scale settings was developed. These VLP contained the major immunological determinants of the virus: glycoprotein complex, nucleoprotein, and Z matrix protein, with known post-translational modifications. The viral proteins packaged into LASV VLP were characterized, including glycosylation profiles of glycoprotein subunits GP1 and GP2, and structural compartmentalization of each polypeptide. The host cell protein component of LASV VLP was also partially analyzed, namely glycoprotein incorporation, though the identity of these proteins remain unknown. All combinations of LASV Z, GPC, and NP proteins that generated VLP did not incorporate host cell ribosomes, a known component of native arenaviral particles, despite detection of small RNA species packaged into pseudoparticles. Although VLP did not contain the same host cell components as the native virion, electron microscopy analysis demonstrated that LASV VLP appeared structurally similar to native virions, with pleiomorphic distribution in size and shape. LASV VLP that displayed GPC or GPC+NP were immunogenic in mice, and generated a significant IgG response to individual viral proteins over the course of three immunizations, in the absence of adjuvants. Furthermore, sera from convalescent Lassa fever patients recognized VLP in ELISA format, thus affirming the presence of native epitopes displayed by the recombinant pseudoparticles.ConclusionsThese results established that modular LASV VLP can be generated displaying high levels of immunogenic viral proteins, and that small laboratory scale mammalian expression systems are capable of producing multi-milligram quantities of pseudoparticles. These VLP are structurally and morphologically similar to native LASV virions, but lack replicative functions, and thus can be safely generated in low biosafety level settings. LASV VLP were immunogenic in mice in the absence of adjuvants, with mature IgG responses developing within a few weeks after the first immunization. These studies highlight the relevance of a VLP platform for designing an optimal vaccine candidate against Lassa hemorrhagic fever, and warrant further investigation in lethal challenge animal models to establish their protective potential.

Project description:PurposeHeterologous virus-like particle (VLP) assembly involving influenza or the Newcastle disease virus matrix protein (M) has been extensively used to explore the efficacies of VLP vaccines against the respiratory syncytial virus (RSV). Here, we attempted to generate homologous RSV VLPs by expressing the pre-fusion (pre-F) or the glycoprotein (G) on the RSV M protein and evaluated their protective efficacy in mice.MethodsWe generated VLPs using the baculovirus expression system in Spodoptera frugiperda (Sf9) insect cells. Recombinant baculoviruses expressing the RSV pre-F, G, and M antigens were inoculated into Sf9 cells, and particles were self-assembled. Mice were immunized with either pre-F or G-expressing VLPs, and immune parameters were assessed to determine protection.ResultsOur findings show that successful VLP assembly can be achieved by utilizing recombinant baculoviruses expressing the RSV pre-F or G proteins with the native matrix protein. Mice immunized with either pre-F or the G antigen-expressing VLPs elicited robust serum-mediated virus neutralization. VLP immunization evoked Th1-biased RSV-specific antibody responses in the sera of mice. Following challenge infection with the RSV A2 strain, immunized mice experienced lesser eosinophil and IL-4 accumulation in the lungs, though a substantial increase in TNF-α secretion was observed from CD4+ T cells. Interestingly, splenic antibody-secreting cell responses were substantially enhanced against RSV F antigen, but not against the RSV G antigen following immunization and challenge infection. Immunizing mice with the VLPs significantly inhibited pulmonary histopathology development, as indicated by the diminished inflammatory immune cell influx and mucin secretion.ConclusionCombined, these vaccine-induced immune responses contributed to successfully inhibiting the RSV replication in the lungs of mice and demonstrated that RSV VLP assembly using insect cell-derived homologous RSV matrix protein is a feasible approach.

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Lassa virus Z protein is a matrix protein and sufficient for the release of virus-like particles [corrected].

Publications

Lassa virus Z protein is a matrix protein and sufficient for the release of virus-like particles [corrected].

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