UBC Theses and Dissertations
Intracellular signaling networks regulate host-cell responses to coxsackievirus B3 infection Sadeghi Garmaroudi, Farshid
The host-cell response to viruses such as coxsackievirus B3 (CVB3) is determined by the signal-transduction networks modified during infection. To unravel the function of signal-transduction networks, these networks can be perturbed biochemically or genetically. But at the systems-level, it is unclear how such perturbations are best implemented to extract molecular mechanisms underlying network function. Further, the modified signal-tranuction network is a cause of multiple interdependent pathophenotypes, which makes it difficult to link virus-induced signals and responses. Either pairwise perturbations of host-cells infected by CVB3 or graded doses of CVB3 will reveal causal and unappreciated network mechanisms in CVB3-induced signaling and pathogenesis. I used CVB3 infection of cardiomyocytes as a representative virus-host system. In a first study, I combined pairwise perturbations with multiparameter phosphorylation measurements to reveal causal mechanisms within the signaling-network response of cardiomyocytes to infection. In a second study, I adopted a quantitative multiparameter approach to interrelate CVB3-induced host-cell events. I built a model that quantitatively predicts cardiomyocyte responses from time-dependent measurements of phosphorylation events. Cluster analysis of the dataset showed that paired-inhibitor data was required for accurate data-driven predictions of kinase-substrate links in the host network. With pairwise data, I also derived a high-confidence phospho-protein network based on partial correlations, which identified phospho-IkBa as a “hub”. This network helped connect phospho-IkBa with an autocrine feedback circuit in host cells involving the proinflammatory cytokines, TNF-α and IL-1. Autocrine blockade substantially inhibited CVB3 progeny release and improved host-cell viability, implicating TNF-α and IL-1 as cell-autonomous components of CVB3-induced myocardial damage. As well, analysis of multiple-dosing data revealed a tight coupling between the ERKs (ERK1/2 and ERK5) and p38 MAPK pathways with respect to CVB3-stimulated cytotoxicity. I showed that ERKs redundantly control a caspase-9-dependent apoptotic program, whereas p38 is required for CVB3-driven necrosis. I conclude that pairwise perturbations, when combined with network-level intracellular measurements, enrich for mechanisms that would be overlooked by single perturbants. Moreover, virus-host interactions may seem especially complex because multiple overlapping pathophenotypes are being executed concurrently. By monitoring these pathophenotypes experimentally and deconvolving them mathematically, the control of each could be revealed as much more simple.
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