PROJECT SUMMARY/ABSTRACT The disorders collectively referred to as inflammatory bowel disease (IBD) are characterized by aberrant immune responses to the commensal microbiota. The establishment and maintenance of tolerance to the microbiota is accomplished via a complex and dynamic network of cells and molecules that collectively stave off immune reactivity. Elucidation of the mechanisms whereby these natural pathways limit immunity to normally benign commensals, especially those that occupy niches at the epithelial border, will ultimately enhance our ability to target these interactions for the prevention and/or treatment of intestinal inflammation. We hypothesize that high-affinity, thymus-dependent (T-dependent) antibody responses are essential for the regulation of mucus-associated bacterial communities. T-dependent antibody responses are especially important in the host response to pathogenic invasion of the gut. Since some helpful commensal species utilize similar mechanisms as pathogens to establish a niche in the mucus layer lining the intestinal epithelium, it stands to reason that similar immune mechanisms are deployed to control these organisms. We have found that mice colonized with a minimal mucus-associated consortium display a predominantly T-dependent IgG response. Moreover, mice with impaired production of T-dependent antibodies have significantly diminished total antibody responses to antigens derived from mucus-associated bacteria including specific flagellins derived from members of the family Lachnospiraceae. Colonization with mucus-associated bacteria, or the deficiency of T-dependent antibodies in the presence of such organisms, coincides with increased production of the immunoregulatory cytokine interleukin-10 (IL-10) by CD4 T cells in the large intestine. Accordingly, simultaneous disruption of these 2 pathways results in spontaneous colonic inflammation. In this proposal, we will utilize defined microbes and the antigens they generate to elucidate the mechanisms whereby interactions between flagellated mucus-associated bacteria and the gut immune system promote tolerance of these communities. In addition, we will explore novel roles for T-dependent anti-commensal IgG1 in the establishment and maintenance of immune homeostasis. If successful, our studies will define the mechanisms whereby TD anti-commensal antibodies promote immune normalcy at the colonic epithelial border and constantly mitigate the risk of deleterious reactivity to the diverse community of mucus-associated bacteria.