IL-4 is expressed in Th2 cells and basophils. Increased expression of IL-4 is an established hallmark of allergic diseases. Recent studies have allowed the identification of nuclear factors (NF) whose expression or activation are associated with Th2-restricted expression of IL-4. We have identified two NF, CP2 and NF-kappaB, exerting diverging effects on IL-4 and IL-2 transcription. We show that CP2 and the NF-kappaB species p65 and p50 are differentially expressed in Thl and Th2 cells and regulate transcription by directly interacting with the IL-4 promoter via a complex interplay with other NF. We hypothesize that these proteins play a relevant role in activation of the IL-4 gene. We will investigate: 1) The role of CP2 in IL-4 gene activation (Aim 1). We will analyze the expression and function of CP2 in Th cells and basophils and the effect of CP2 depletion, functional impairment or forced expression on IL-4 production. 2) The significance of IL-4 regulation by NF-kappaB molecular species (Aim 2). Our findings implicate that the p65-p50 balance directly affects IL-4 expression in Th cells and basophils. We will look at the expression of p65 in these cells, and the impact of selective p65 blockade on cytokine induction in mice transgenic for a transdominant repressor. 3) The mechanism of IL-4 gene activation by CP2 and p50 (Aim 3). By in vitro and in vivo analysis of protein-DNA interactions we will study the functional interaction of CP2 and p50 with the IL-4 promoter and distal regulatory elements and their impact on IL-4 locus remodeling and accessibility. We will study the significance of their interaction with the cofactors Yin Yang 1, Bcl-3 and NF-IL6. Dissecting the mechanisms of IL-4 regulation is of crucial importance to understanding the molecular basis of allergic disease. The proposed approach, by defining the roles of CP2 and NF-kappaB in IL-4 transcription, will provide a basis for studies of the expression ana function of these factors in allergic inflammation, and identify novel, specific molecular targets for immunomodulation. Given the involvement of CP2 and NF-kappaB in transcription of a diverse host of cellular and viral genes, our results will also have broader implications in the definition of the significance and function of these proteins in the regulation of gene expression in health and disease.
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