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GRK 1482 Jahrbuch 2011-2014

Abstract In intestinal epithelial cells, the presence of bacteria and bacteria-de- rived peptides is proposed to cause an activation of NOD and NF-κB with PEPT1 mediating the uptake of these peptides into cells. Duox1/2 expression was shown to be activated by NF-κB and is increased by calcium [4]. Thus, we will investigate the interplay of the three brush border membrane proteins dual oxidases DUOX1/2, peptide tran- porter PEPT1 and Na+/H+ exchanger NHE3 by using C. elegans, the human colon carcinoma cell line Caco-2, as well as murine tissue samples. Introduction The epithelial cells in small and, to a larger extend, large intestine express on their luminal surface DUOX (dual oxidase) proteins, a class of NADPH- oxidases that generate H2O2 from O2. Its function is coupled to proton efflux via the sodium-proton exchanger NHE3. This provides a pro-oxidative sur- face environment that affects microbiota homeos- tasis and seems also be part of a NOD-mediated antibacterial response (see Figure 1). From a screen using the model organism C. elegans we obtained evidence that peptide transporter PEPT1 and NHE3 possess some co-regulation in the intestine [1] in the context of ROS production, and we speculate that this interplay is part of a distinct microenvironment at the gut surface that affects the microbiotasurface interaction. The re- gulation of the intracellular pH is tightly coupled to nutrient transport processes and to innate im- munity/pathogen defense mechanisms. We have demonstrated that intracellular pH homeostasis and proton flux dependent on Na+/H+ exchange by NHE3 and PEPT1 activity modulate fatty acid uptake and body fat content [2], and these processes might also be involved in DUOX functional regulation. The C. elegans Duox homologue Ce-Duox/BLI-3 was shown to be responsible for H2O2 production after infection with E. faecalis [3]. Wild type C. ele- gans were more susceptible to the infection when bli-3 mRNA expression was reduced by RNA inter- ference. By measuring the nematodal H2O2 pro- duction by the Amplex Red Assay this well charac- terized model organism will be used as an in vivo system for analyzing the physiological and molecu- lar interactions of the host with pathogenic bacteria. PhD FELLOWS Page 46 | GRK Progress Report 2011-2014 Julia Vörös (M.Sc.) Physiology of Human Nutrition PhD 8/2 Colonic DUOX2-mediated epithelial H2O2 production, its functional coupling to pH-homeostasis and its role in defence against bacteria