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Measurement of γHV68 Infection in Mice
Published on: November 22, 2011
Murine gammaherpesvirus 68 lacking gp150 shows defective virion release but establishes normal latency in vivo
Brigitte D de Lima1, Janet S May, Philip G Stevenson
1Division of Virology, Department of Pathology, University of Cambridge, Tennis Court Road, Cambridge CB2 1QP, United Kingdom.
Abstract:
All gammaherpesviruses encode a virion glycoprotein positionally homologous to Epstein-Barr virus gp350. These glycoproteins are thought to be involved in cell binding, but little is known of the roles they might play in the whole viral replication cycle. We have analyzed the contribution of murine gammaherpesvirus 68 (MHV-68) gp150 to viral propagation in vitro and host colonization in vivo. MHV-68 lacking gp150 was viable and showed normal binding to fibroblasts and normal single-cycle lytic replication. Its capacity to infect glycosaminoglycan (GAG)-deficient CHO-K1 cells and NS0 and RAW264.7 cells, which express only low levels of GAGs, was paradoxically increased. However, gp150-deficient MHV-68 spread poorly through fibroblast monolayers, with reduced cell-free infectivity, consistent with a deficit in virus release. Electron microscopy showed gp150-deficient virions clustered on infected-cell plasma membranes. MHV-68-infected cells showed reduced surface GAG expression, suggesting that gp150 prevented virions from rebinding to infected cells after release by making MHV-68 infection GAG dependent. Surprisingly, gp150-deficient viruses showed only a transient lag in lytic replication in vivo and established normal levels of latency. Cell-to-cell virus spread and the proliferation of latently infected cells, for which gp150 was dispensable, therefore appeared to be the major route of virus propagation in an infected host.
Insights
Murine gammaherpesvirus 68 glycoprotein 150 (gp150) is crucial for efficient virus release and spread in vitro by preventing virion re-binding to infected cells. In vivo, cell-to-cell spread and latent cell proliferation are the primary propagation routes, making gp150 dispensable.
Area of Science:
- Virology
- Immunology
- Cell Biology
Background:
- Gammaherpesviruses share a conserved virion glycoprotein (homologous to Epstein-Barr virus gp350) implicated in cell binding.
- The precise role of these glycoproteins in the viral replication cycle remains largely uncharacterized.
Purpose of the Study:
- To investigate the function of murine gammaherpesvirus 68 (MHV-68) gp150 in viral propagation and host colonization.
- To elucidate the mechanisms by which gp150 influences viral spread and infectivity.
Main Methods:
- Generation and analysis of gp150-deficient MHV-68.
- In vitro studies including cell binding assays, single-cycle lytic replication, infection of glycosaminoglycan (GAG)-deficient cells, and fibroblast monolayer spread assays.
- In vivo studies assessing viral replication and latency establishment.
- Electron microscopy to visualize virion distribution.
Main Results:
- MHV-68 lacking gp150 exhibited normal fibroblast binding and single-cycle lytic replication but showed impaired spread in fibroblast monolayers due to reduced virus release.
- gp150-deficient MHV-68 displayed paradoxically increased infectivity in GAG-deficient or low-GAG-expressing cells.
- Electron microscopy revealed virion clustering on infected cell membranes, suggesting gp150 prevents virion re-binding to GAGs on infected cells.
- In vivo, gp150-deficient viruses showed only a transient replication lag and established normal latency, indicating cell-to-cell spread and latent cell proliferation are key in vivo propagation routes.
Conclusions:
- gp150 is essential for efficient MHV-68 release and cell-free spread in vitro by mediating GAG-dependent infection.
- The primary in vivo propagation mechanisms for MHV-68 involve cell-to-cell spread and latent reservoir expansion, rendering gp150 dispensable for overall host colonization.
- MHV-68 gp150 plays a critical role in regulating viral dissemination by controlling virion release and preventing premature re-binding to host cells.
