Epigenetic regulation of epidermal proinflammatory responses

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ABSTRACT It has been argued that keratinocytes at the interface of the body with the outside world have developed specialized mechanisms to fulfill the need to nimbly respond to an ever-changing array of challenges with rapid and reversible gene activation. Our past studies support the hypothesis that signaling information pertaining to skin environmental insults is rapidly translated at repressed enhancers of stress response genes into chromatin changes leading to their temporary induction. We have shown that the Mi-2β/NuRD complex sets a chromatin environment that restricts transcription at genes that will be induced in keratinocytes by stress signals, holding them silent but poised for rapid activation. Our studies highlight a dynamic switch between chromatin remodelers that restrict gene expression and early response factors that support transcriptional activation. The mechanisms that support this temporal switch and their contribution to response memory are investigated. In Aim 1, we study how Mi-2β/NuRD is tethered to stress response loci in homeostasis, removed during the stress response, and returned during resolution. We test whether Mi-2β/NuRD is targeted to these loci by stable association with TFs, using both unbiased approaches and a focused approach on the AP1 complex. We characterize post-transcriptional modifications (PTMs) of Mi-2β/NuRD or functionally associated TFs during the early stress response, and test whether these have functional consequences on and off chromatin. We ask how changes in the composition of the AP1 complex composition at target loci contribute to Mi-2β/NuRD complex displacement and return, and whether enduring changes in TF occupancy at the loci or PTMs contribute to epigenetic memory of the stress response. We examine how specificity is achieved by examining Mi-2β target loci that are regulated by different TFs that do not respond to stress signaling. In Aim 2, we investigate how histone modifications and higher order chromatin conformations contribute to induction, repression, and reactivation of stress response genes in keratinocytes. We test for changes in local histone modifications and TF recruitment at stress response genes and whether these contribute to the memory of the response, defined as accelerated kinetics and increased magnitude of the response upon rechallenge. We examine long-distance interactions between stress response enhancers, affiliated promoters, and CTCF architectural sites, whether these are already present prior to stress, if they actively form in response to it, and whether changes are maintained at some loci to alter activation thresholds as a mechanism of epigenetic memory. Targeted manipulation of Mi-2β/NuRD and AP1 complex occupancy at specific loci is employed to test their role in directing chromatin contacts and histone modifications at rest and during the response. We also examine the heterogeneity of chromatin accessibility among basal cell precursors, its heritability within clones, and the contribution of changing clonal composition of the basal layer to the evolution of the epigenetic memory of prior inflammatory insult.