Lymphocytes contained within distinct regions of the gut epithelium represent the first line of immunologic defense to gut-mucosal pathogens. Studies aimed at understanding the nature of gut-mucosal immune responses and the development of methods for effective gut-mucosal immunization are important for practical, as well as theoretical reasons since effective gut-mucosal immunity is likely important for the control and resolution of infections which commence at mucosal surfaces. We have used gut-mucosal infection with reovirus, an enteric virus which enters gut mucosal lymphoid follicles (Peyer's patches-PP) from the gut lumen, to explore basic questions of gut mucosal immunology. We have found that intraduodenal (i.d.) application of infectious reovirus perturbs both B- and T- cell populations in both an antigen specific and non-specific manner. The i.d. application of reovirus results in the appearance of virus-specific precursor CTL in PP as well as in the intestinal epithelium. I.D. immunization with reovirus has also been found to result in the transient appearance of a population of germinal center B cells, and a transient rise in the proportion of CD8+ T cells in PP which express a novel cell surface antigen termed GCT. While the functional significance of tide GCT+ B cell population is as yet undefined, we have demonstrated that reovirus-specific effector and pCTL occur within the GCT+ subpopulation of CD8+ lymphocytes. We will continue our studies of gut-mucosal immunity using reovirus as a mode. We propose to define reovirus-specific T cell epitopes via the use of a novel RNA virus (Sindbis virus) vector system. Expression of individual reovirus gene segments will be necessary as a first step to identify which reovirus genes are involved in T cell responses since we have found that common determinants shared by reovirus polypeptides from several reovirus serotypes are most likely the stimuli for the majority of T cell responses to reovirus. The identification of individual T cell epitopes will allow us to analyze their recirculatory patterns and functional significance in murine model systems. We will also continue our studies of the function and significance of the GCT antigen. The following six specific aims will guide, our continued studies of gut mucosal immunity using reovirus as a model gut mucosal immunogen. l) Definition of specific CTL epitopes of reovirus in the context of gut- mucosal and non-mucosal lymphoid tissues. 2) Definition of specific TH epitopes of reovirus in the context of gut- mucosal and non-mucosal lymphoid tissues. 3) Molecular cloning of the GCT antigen. 4) Analysis of the reovirus-specific GCT expressing CD8+ population. 5) Analysis of th redirculatory patterns, in vivo, of reovirus-specific T lymphocytes. 6) Analysis of epitope-specific T cell populations in murine models for infection/protection studies.
