Mononuclear phagocytes may participate in host defense against cancer through a diverse spectrum of activities which include not only direct cytotoxicity but also the recruitment and/or activation of other anti- tumor effector cell populations. Both direct and indirect anti-tumor function involves the inducible expression of many new gene products including inflammatory cytokines and other genes expressed very early after stimulation. The diversity of the tumoricidal activation process reflects the complexity of inducible macrophage inflammatory gene expression. The successful therapeutic application of Biologic Response Modifiers (BRMs), which may act in part through activation of macrophage tumoricidal function will necessitate understanding the mechanistic determinants of inflammatory cell behavior. The guiding hypothesis for the following proposal is that diverse patterns of macrophage early gene expression derive, at least in part, from the complex events which control transcriptional activation of inducible genes. Previous work supported by this grant has identified and characterized diverse expression patterns for three genes (IP-10, D3, and IL-1beta) in macrophages treated with one of several inflammatory stimuli (IFNgamma, LPS, IFNbeta). The regulation of inducible expression of these three genes provides a model in which to study the complexity evident in transcriptional control in this cell type and to estimate how much the diversity in macrophage activation may be explained at this level. To test this hypothesis and to define the mechanisms controlling inducible transcription during tumoricidal activation of macrophages we propose the following specific experimental aims: 1. To identify and define the nucleotide sequence elements which regulate transcriptional activity of a selection of inducible inflammatory gene products in response to a selection of prototypic stimuli known to promote diverse patterns of gene expression in murine macrophages. 2. To identify and define the nuclear protein(s) which recognize and bind to such sequences and to evaluate the relationship between stimulation and activity of these factors. 3. To determine if such sites are occupied in vivo in macrophages undergoing response to selected stimuli using in vivo footprinting by Ligation Mediated Polymerase Chain Reaction (LMPCR).
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