Mechanisms Regulating Selective Gene Activation During the Innate Immune Response

The cells of the innate immune system are responsible for the first line of defense against foreign dangers. Recognition of pathogens results in the transcriptional upregulation of a stimulus-specific inflammatory gene program to counteract infection and initiate adaptive immune responses. An appropriate response is necessary to resolve infection, but excessive inflammation can damage host tissues and lead to inflammatory diseases. Therefore, it is critical to understand how inflammatory responses are selectively achieved in response to diverse stimuli. The dissertation describes two studies that attempt to better understand the regulatory mechanisms underlying selective gene activation in response to inflammatory stimuli. The first study explores how signaling pathways, transcription factors, and chromatin act in concert to shape the inflammatory gene program. Using genome-wide techniques to interrogate chromatin-associated RNA, insight was gained into the lipid A-induced transcriptional cascade in macrophages. A quantitative analysis of transcription factor binding combined with kinetic and expression data derived from loss-of-function mutant mouse strains have allowed us to identify co-regulated genes, particularly those regulated by NF-κB, interferon response factor-3, and serum response factor. Furthermore, subsets of co-regulated secondary response genes were found to play distinct roles in immunity, underscoring the diverse mechanisms underlying selective gene activation. This has revealed insight into the unique regulatory logic for each inflammatory gene, and serves as a framework for understanding selective gene activation in various physiological settings. The second study utilizes the findings from the first study to investigate the mechanisms of LPS tolerance. A global interrogation of the effects of LPS tolerance in macrophages revealed a broad downregulation of gene expression in the tolerant state. In addition, a large subset of inducible genes exhibited prolonged transcription even after the tolerizing dose of LPS was removed but before the second LPS treatment, which could be partially explained by the presence of other cytokines mediating their activation in the tolerant state. Furthermore, previously described negative regulators of LPS signal transduction were expressed at higher levels in tolerant macrophages, including those inhibiting signals proximal to the TLR4 receptor. Together, the framework for understanding the regulatory logic of selective gene activation that can be utilized to unravel the mechanisms underlying diverse inflammatory settings