Intestinal M Cells: Uptake and Fate of Enteric Pathogens

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DESCRIPTION (provided by applicant): Enteric pathogens can exploit the transport activity of M cells to invade the intestinal mucosa. The mouse pathogen reovirus (Type 1 Lang) adheres selectively to M cells in adult mice and establishes infection in Peyer's patches before spreading systemically. Reovirus provides an excellent model for understanding pathogen-M cell adherence and for following the earliest stages of viral pathogenesis in Peyer's patches. Aim I is to elucidate the molecular interactions that allow reovirus to selectively adhere to M cells. The hypothesis that the reovirus outer capsid protein sigma 1 is the viral adhesin responsible for M cell binding will be tested using recombinant outer capsid proteins and reconstituted viral cores. We will use affinity methods to separate apical membrane glycoconjugates of epithelial cells, and overlay approaches to identify the epithelial membrane glycoproteins and/or glycolipids that can serve as reovirus receptors. Also, the hypothesis that integral membrane mucins can mask reovirus binding sites will be tested. Aim II is to use the reovirus model to clarify the multiple, paradoxical effects that secreted IgA may have on entry of pathogens via M cells, and on subsequent mucosal immune responses. IgA can prevent mucosal infection, yet IgA-antigen complexes can adhere to M cell apical surfaces. A unifying hypotheses to be tested is that 1) naturally-secreted IgAs against outer capsid proteins can protect by entrapping virus in mucus and preventing mucosal contact, but 2) if virus coated with IgA succeeds in contacting the FAE, it can adhere to M cells via IgA and enter the mucosa. In addition, the possible protective effect of the endogenous IgA coat on M cells will be examined using IgA knockout and normal mice. Aim III will examine the fate of reovirus and reovirus-infected cells after M cell transport. We will test two hypotheses: 1) that M cell-transported reovirus is taken up by and infects a specific subset of dendritic cells (DCs) in the subepithelial dome region, and 2) that the mucosal adjuvant cholera toxin can drive the movement of the DCS carrying virus or virus-sized particles to other sites in Peyer's patches and beyond, facilitating viral dissemination. The proposed studies will elucidate how M cells "select" pathogens for mucosal immune surveillance, how pathogens exploit the M cell transport pathway to cause disease, and how vaccines and vaccine vectors may be efficiently targeted to the mucosal immune system.