Myeloid specific and CD4+ T cell dependent mechanisms of gammaherpesvirus infection control.

Abstract

The human gammaherpesviruses (g-HV), Epstein Barr virus (EBV) and Kaposirs sarcoma virus (KSHV), cause cancers. They establish chronic B cell infections through immune evasion and latency. EBV is ubiquitous in human populations, infecting over 95% of adults worldwide. Its public health burden is unknown and likely underestimated, but its undisputed disease associations make vaccination a worthwhile goal. Early infection is likely to provide the best targets. However, because this precedes clinical presentation, it has been a difficult objective. The species-restrictions of individual g-HVs make non-human EBV infections problematic to interpret. Yet g-HV infections long predate human speciation, so close parallels are likely between those of different mammalian hosts. Mice provide the main in vivo model of mammalian biology. Murid herpesvirus-4 (MuHV-4), a g-HV isolated from yellow-necked mice, offers a valuable opportunity to understand core, conserved features of early g-HV host colonization, and hence to develop vaccine strategies applicable to EBV and KSHV. Like EBV and KSHV, MuHV-4 infects B cells, drives their proliferation, and persists in resting memory cells. It was employed in this thesis to analyse g-HV infection control.EBV has been isolated in vivo from many more cell types than are routinely studied in vitro. Like EBV, MuHV-4 principally colonizes B cells at steady state, but it enters new hosts via epithelial infection, and myeloid cells provide the gateway to B cells. Hence, myeloid cell infection has been a primary focus of this thesis. Analysis of organ homogenates and flow cytometry has been the preferred approach to tracking B cell infection. However, for myeloid cells, their anatomical site can be functionally more revealing. I have therefore extensively analysed tissue sections. Functional analysis of myeloid specific events was achieved with LysM-cre and CDllc-cre transgenic mice. Myeloid cells are diverse, and no single molecule delineates a functionally distinct subset. LysM-cre and CDllc-cre mice are nonetheless reasonably well characterised, and approximately target the macrophage-like (LysM+) and dendritic cell (DC)-like (CDllc+) extremes of the myeloid spectrum. Cre expressing transgenic mice were crossed with floxed type 1 interferon receptor mice to determine the myeloid-specific effects of this central anti-viral innate effector. They were additionally crossed with floxed MHC class II mice to determine the role of myeloid cells in interacting with CD4+ T cells, a key anti-viral adaptive effector. In both settings, a novel role for anti-viral NK cell recruitment was identified.The main aim of understanding anti-viral immunity is to develop better vaccines. Recombinant subunit vaccines to EBV and MuHV-4 have failed to reduce long term infections. Live attenuated MuHV-4 has successfully protected. However, live attenuated EBV is made problematic by its oncogenic latency genes. My analysis of anti-viral immunity consistently pointed towards lytic infection of myeloid cells being a key target for infection control. I generated a recombinant MuHV-4 that lacks all known latency genes, thus eliciting immune cell priming through engaging lytic antigens alone. It proved to be an effective vaccine. It showed also that immunity to MuHV-4 specific genes is not required for protection. Translating this protection to lytic-only EBV may therefore be feasible, and provides an important future goal.