Molecular characterisation and computational modelling of macrophage heterogeneity of major immediate early gene expression during a murine cytomegalovirus infection
Hassim, Muhamad Fairus Bin Noor
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Human cytomegalovirus (HCMV) is a major cause of morbidity and mortality amongst immuno-compromised individuals and is the leading cause of congenital diseases amongst newborn infants. Mouse CMV (MCMV) infection of inbred mice has been extensively used as a model for HCMV pathogenesis and host-virus interaction. Macrophages are a key target cell type in the pathogenesis of human and mouse CMV infections. Macrophages are semi-permissive to CMV infection, however, the nature of this restrictive mechanism of infection is open for investigation. In this thesis, I hypothesized that macrophage permissivity is determined by the dynamic interplay of the innate response during the immediate-early (IE) period of infection. To test this hypothesis, I first developed and validated a flow cytometry based assay. In MCMV infected macrophages, I found heterogeneous expression from the major IE promoter (MIEP) leading to the development of a refractory subpopulation for IE expression. I further developed a computational modelling approach to help elucidate the dynamics of infection during this period. Modelling work revealed that the occurrence of refractory subpopulation could be caused by either 1) pre-existence of heterogeneous permissivity of macrophages prior to infection or 2) through an emergent process. Experimental testing of the models shows that the heterogeneous IE expression of homogeneously infected macrophages is caused by an emergent process. MCMV infection using type I interferon receptor and signal transducers and activator of transcription 1 (Stat1) knockout macrophages reveals that the emergence of refractory subpopulation is predominantly mediated by type I interferon through Stat1. Comparative molecular analysis between progressively infected and refractory subpopulations reveals that MCMV MIEP activation in the refractory subpopulation is stochastically inhibited by high expression of type I interferon induced antiviral components.