Hsp70 and a novel axis of type I interferon-dependent antiviral immunity in the measles virus-infected brain.

J Virol. 2013 Jan;87(2):998-1009.

Kim MY, Shu Y, Carsillo T, Zhang J, Yu L, Peterson C, Longhi S, Girod S, Niewiesk S, Oglesbee M.

Department of Veterinary Biosciences, The Ohio State University, Columbus, OH, USA.

Abstract

The major inducible 70-kDa heat shock protein (hsp70) is host protective in a mouse model of measles virus (MeV) brain infection. Transgenic constitutive expression of hsp70 in neurons, the primary target of MeV infection, abrogates neurovirulence in neonatal H-2(d) congenic C57BL/6 mice. A significant level of protection is retained after depletion of T lymphocytes, implicating innate immune mechanisms. The focus of the present work was to elucidate the basis for hsp70-dependent innate immunity using this model. Transcriptome analysis of brains from transgenic (TG) and nontransgenic (NT) mice 5 days after infection identified type I interferon (IFN) signaling, macrophage activation, and antigen presentation as the main differences linked to survival. The pivotal role of type I IFN in hsp70-mediated protection was demonstrated in mice with a genetically disrupted type I IFN receptor (IFNAR(-/-)), where IFNAR(-/-) eliminated the difference in survival between TG and NT mice. Brain macrophages, not neurons, are the predominant source of type I IFN in the virus-infected brain, and in vitro studies provided a mechanistic basis by which MeV-infected neurons can induce IFN-β in uninfected microglia in an hsp70-dependent manner. MeV infection induced extracellular release of hsp70 from mouse neuronal cells that constitutively express hsp70, and extracellular hsp70 induced IFN-β transcription in mouse microglial cells through Toll-like receptors 2 and 4. Collectively, our results support a novel axis of type I IFN-dependent antiviral immunity in the virus-infected brain that is driven by hsp70.

PMID: 23135720

 

Supplement:

Immune clearance of non-cytopathic measles virus infection of neurons requires activation of brain macrophages (microglia) to cross present viral antigen and to produce type I interferon, which is primarily interferon beta (IFN-β) in the brain. These macrophage functions support stimulation of virus-specific T cells that drive viral clearance through the production of interferon gamma. A long-standing question has been the mechanism by which virus infected neurons activate brain macrophages and stimulate the production of IFN-β. Levels of IFN-β in particular are pivotal to protection against neurovirulence by a number of viruses. Results of the present work supports a model in which extracellular 70 kDa heat shock protein (hsp70) is the link between the virus infected neuron and uninfected macrophages, representing a novel axis of antiviral immunity.

The mouse model of measles virus (MeV) brain infection was used to define the biological relevance of virus-hsp70 interaction. A model summarizing our results is provided in figure 1. Mice, unlike humans, lack constitutive expression of hsp70. As such, transgenic constitutive expression of hsp70 in mice was used to determine the influence of this heat shock protein on the outcome of infection. In vitro studies in mouse neuronal cells show that MeV induces hsp70 and that hsp70 in turn enhances MeV gene expression. If hsp70 is constitutively expressed within the cell or induced by transient hyperthermia, MeV gene expression is increased. This relationship stimulates the extracellular release of hsp70 from viable infected cells, a mechanism that may involve microvesicular shedding or exosomal secretion. Extracellular hsp70 is ligand for Toll-like receptor 4, expressed primarily on brain macrophages, and may thus induce macrophage activation and antigen presentation functions through NFκB, and IFN-β production through IRF3. In vivo studies show that constitutive expression of hsp70 confers protection that is correlated to gene expression profiles indicating enhanced macrophage activation and type I interferon production. Addition studies show that hsp70-mediated protection is type I interferon-dependent.  Although not addressed in the current work, the model may also explain how brain macrophages can cross-present viral antigen derived from neurons. Hsp70 binds the MeV nucleocapsid protein, particularly the domain containing epitopes that are determinants of cell-mediated viral clearance. Hsp70-antigen complexes can be internalized by macrophages through scavenger receptors such as CD91, resulting in cross-presentation of exogenous antigen by the activated macrophage.

More recent studies have documented the more broad virological relevance of this novel axis of antiviral immunity using highly cytopathic mouse brain infections caused by vesicular stomatitis virus(1). As with MeV, constitutive hsp70 expression increases VSV gene expression which in turn stimulates extracellular release of hsp70 from both viable and apoptotic infected cells. Mice that constitutively express hsp70 are protected against VSV neurovirulence, and protection is correlated to enhanced production of IFN-β in brain. Time course analysis of brain viral burden shows that the predominant influence of hsp70-dependent innate immune responses is on viral clearance, similar to earlier work with MeV in which hsp70 was induced by transient hyperthermia. Work with VSV also included expression of hsp70 from the viral genome, showing that it is hsp70 expressed within the virus infected cell that mediates protective host responses.

Collectively, results show that elevating levels of hsp70 within the virus infected cell evokes a novel axis of innate immunity that drives viral clearance. The febrile response may thus be viewed as a host protective response to viral infection, given that fever is a potent inducer of hsp70. Morever, the protection mechanism may apply to both non-cytopathic and cytopathic viral neuronal infections. Collectively, results suggest novel therapeutic approaches to viral encephalitis that are based upon increasing intracellular levels and subsequent extracellular release of hsp70.

Innate immunity update3

1.            Kim, M. Y., Y. Ma, Y. Zhang, J. Li, Y. Shu, and M. Oglesbee. 2013. Hsp70-Dependent Antiviral Immunity against Cytopathic Neuronal Infection by Vesicular Stomatitis Virus. J Virol. 87(19);10668-10678.

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