Molecular Determinants of Stimulus-Specificity in Macrophage Reprogramming

The clinical outcome of infectious diseases is largely dictated by the response of the immune system to the pathogen. Immune responses are context-specific and significantly affected by factors such as tissue microenvironment, age, chronic diseases, cytokines, or previous infections. Contextual variables alter immune function by reprogramming cells of the innate immune system such as macrophages, altering their signaling networks and epigenetic states. Importantly, this reprogramming is stimulus-specific, and both the scope and underlying mechanisms of this specificity are areas of great interest. In Chapter Two, we investigate the differential effects of Type I and II interferon (IFN) cytokines on human macrophage reprogramming by employing a sequential conditioning-stimulation approach. Whereas prior studies have examined direct effects of IFNs, we found that IFNs produced indirect effects that could only be appreciated upon subsequent stimulation with a pathogen-associated molecule and transcriptomic analysis across multiple time points. We identified 713 genes that were unaffected by IFN alone, yet after IFN conditioning had an altered gene expression response to a subsequent stimulus. Surprisingly, we also found that the IFNs were not uniformly pro- or anti-inflammatory as previously described. Instead, the effects of Type I and II IFN were gene-specific and stimulus-specific. IFN conditioning affected both signaling networks and the epigenetic state, providing mechanistic explanations for our findings.In Chapter Three we further explore the ability of stimuli to alter the epigenome of macrophages. We found that although many stimuli activate the transcription factor (TF) NFκB, only some were capable of altering the enhancer landscape through the formation of de novo enhancers. We showed that the capacity of NFκB to produce de novo enhancers was correlated with the temporal dynamics of NFκB activity, which are stimulus-specific. In particular, we found that whether NFκB is oscillatory or non-oscillatory was the primary determinant of its capacity to reprogram the epigenome. Thus, we propose a novel mechanism based on temporal dynamics to explain why TFs like NFκB reprogram macrophage epigenomes in a stimulus-specific manner. Future work will investigate the functional and disease consequences of the de novo enhancers that arise specifically from non-oscillatory NFκB-inducing stimuli.