IL-12 and IL-23 are heterodimeric cytokines produced by macrophages and dendritic cells. They share a common p40 subunit which covalently links to distinct glycosylated chains, p35 and p19 respectively. Both cytokines are highly inducible by bacteria and are known to play important roles in mediating chronic intestinal inflammation. In chronic inflammation, IL-23 likely plays the major pathogenic role as demonstrated by the finding that IL-10-/- mice crossed with IL-23 p19-/- mice do not develop colitis, in contrast to IL-10-/- mice crossed with IL-12 p35-/- mice which develop colitis. Furthermore, single nucleotide genetic polymorphisms in the receptor for IL-23 confer susceptibility to and protection against Crohn's disease. However, the molecular regulation of IL-23 p19 remains largely unknown. Inflammatory bowel disease (IBD) is thought to result from an inappropriately directed inflammatory response to the enteric microbiota in a genetically susceptible host. Our laboratory and others have shown that IFN-y synergizes with microbial toll-like receptor (TLR) ligands (LPS, SbLP, CpG etc) and augments IL-12 p40 gene expression. We have recently identified a novel interferon stimulated response element (ISRE) in an area of conserved nucleotide sequences (CMS) across multiple species on the IL-23 p19 promoter. Our preliminary data show that IFN-y inhibits TLR ligand-induced IL-23 p19 gene expression through this ISRE via induction of interferon regulatory factor-1 (IRF-1). This proposal will determine, in detail, the regulation of IL-23 p19 gene expression, with a focus on elucidating critical interactions between the newly identified ISRE with IRFs and NF-kB, and determining, paradoxically, how IFN-y inhibits IL-23 expression. We will also elucidate mechanisms involved in the mucosal regulation of IL-23 p19 by the enteric microbiota utilizing gnotobiotic and conventionalized wild type and IL-10-/- mice. This proposal will provide a comprehensive analysis of combinatorial interactions leading to the regulated expression of IL-23 p19 in macrophages, and will utilize a combination of cell based and in vivo approaches to determine how the enteric microbiota regulate intestinal IL-23 p19 in wild type and IBD-prone mice. Current therapies for IBD have non-specifically suppressed the immune response leading to significant side effects. Therefore, significant unmet medical needs exist in these debilitating human diseases. We will use a variety of modern technologies and mouse models of IBD raised germ-free and colonized with normal intestinal bacteria to determine how intestinal bacteria turn IL-23 on and off in the gut. This information could help identify specific targets that may guide the development of new treatments in IBD that can specifically turn off IL-23 in the intestine.
