Regulation and Targeted Modulation of Dendritic Cell Function by Bacterial Oligosaccharides

PROBLEM. Bacterial pathogens have evolved unique strategies to subvert the host immune system. Dendritic cells (DC) are specialized antigen presenting cells that have a central position in the regulation of the immune response. Pathogens direct DC function for a large proportion by targeting Toll-like receptors (TLR) and, via their oligosaccharides, C-type lectin receptors (CLR). The mechanisms via which DC integrate the multitude of TLR and CLR signals into the phenotype that drives the immune response and the importance of the local DC micromilieu for the response, are still a mystery. This lack of knowledge seriously hampers understanding of pathogen-associated immunopathology and the targeted design of novel immunomodulatory reagents for use in vaccines and immunotherapy. AIM. The aim of this project is to unravel how bacterial targeting of distinct combinations of Toll-like receptors and C-type lectin receptors directs DC function in different local microenvironments and to obtain proof-of-principle that bacterial oligosaccharides can be applied to orchestrate TLR/CLR driven DC function to our benefit. EXPERIMENTAL STRATEGY. The effects of bacterial targeting of distinct TLR and CLR subsets on DC function are investigated via two complementary strategies: (i) Dissection of the effect of variation in bacterial glycan repertoire in a background of fixed TLR stimuli, and (ii) Dissection of the effect of variation in bacterial TLR stimuli in an invariable oligosaccharide context. To discern how different lectin receptors modulate TLR driven DC functions, recombinant Campylobacter jejuni that only differ in glycan repertoire will be used. Campylobacter can express a variety of different oligosaccharides and has been associated with the development of auto-immune disorders. The impact of variation in TLR signal on CLR-mediated DC functions will be addressed with recombinant Neisseria meningitidis strains that express lipopolysaccharide (LPS) with a well-documented unique variable potency towards TLR4. Gene-arrays, cytokine profiling, expression of co-stimulatory molecules, antigen presentation, and T cell activation will serve as parameters of DC function. Importantly, experiments will be performed with monocytes-derived immature DC in the absence and presence of mucosal epithelial cells to appreciate the significance of the complexity of the DC micromilieu for DC function. Once the desired stimuli and their effects on DC function have been defined, targeted modulation of the bacterial glycan repertoire will be established to assess the power of bacterial oligosaccharides to modulate the immune response to our benefit. EXPECTED RESULTS. Broad expertise and excellent starting conditions (all recombinant strains have already been constructed and the needed immunological techniques are fully operational) give confidence that this project will unravel (i) how bacterial pathogens can modulate DC function by influencing the interplay between TLR and CLR, (ii) the significance of environmental conditions (i.e. surrounding epithelial cells) in the modulation of DC, and (iii) the potential use of bacterial oligosaccharides to direct DC function. RELEVANCE. The proposed dissection of the interplay between CLR and TLR receptors provides essential knowledge for future targeted modulation of the immune response and application of bioactive oligosaccharides in vaccine development and immunotherapy.