The role of the C-terminal D0 domain of flagellin in activation of Toll like receptor 5

• DOI: 10.1371/journal.ppat.1006574
• Publication Date: 2017-08-21T13:37:36Z
• Authors: Vida Forstnerič; Karolina Ivičak-K...; Tjaša Plaper; Roman Jerala; Mojca Benčina
• Keywords: 0303 health sciences; QH301-705.5; Blotting, Western; Bacterial Infections; RC581-607; Immunity, Innate; Cell Line; 3. Good health; Mice; Toll-Like Receptor 5; 03 medical and health sciences; Animals; Humans; Immunoprecipitation; Immunologic diseases. Allergy; Biology (General); Research Article; Flagellin

Flagellin is a wide-spread bacterial virulence factor sensed by the membrane-bound Toll-like receptor 5 (TLR5) and by the intracellular NAIP5/NLRC4 inflammasome receptor. TLR5 recognizes a conserved region within the D1 domain of flagellin, crucial for the interaction between subunits in the flagellum and for bacterial motility. While it is known that a deletion of the D0 domain of flagellin, which lines the interior of flagella, also completely abrogates activation of TLR5, its functional role remains unknown. Using a protein fusion strategy, we propose a role for the D0 domain in the stabilization of an active dimeric signaling complex of flagellin-TLR5 at a 2:2 stoichiometric ratio. Alanine-scanning mutagenesis of flagellin revealed a previously unidentified region of flagellin, the C-terminal D0 domain, to play a crucial role in TLR5 activation. Interestingly, we show that TLR5 recognizes the same hydrophobic motif of the D0 domain of flagellin as the intracellular NAIP5/NLRC4 inflammasome receptor. Further, we show that residues within the D0 domain play a previously unrecognized role in the evasion of TLR5 recognition by Helicobacter pylori. These findings demonstrate that TLR5 is able to simultaneously sense several spatially separated sites of flagellin that are essential for its functionality, hindering bacterial evasion of immune recognition. Our findings significantly contribute to the understanding of the mechanism of TLR5 activation, which plays an important role in host defense against several pathogens, but also in several diseases, such as Crohn's disease, cystic fibrosis and rheumatoid arthritis.