Two nonadjacent regions in enteroaggregative Escherichia coli flagellin are required for activation of toll-like receptor 5.
Flagellin is the major structural protein of the flagella of Gram-negative bacteria. Recent work has demonstrated that flagellin is a potent trigger of innate immune responses in a number of eukaryotic cells and organisms, including both mammals and plants. In several different human epithelial cell lines, this innate immune response involves toll-like receptor 5 (TLR5). The mechanisms by which flagellin activates TLR5 and the importance of this interaction in other model systems of flagellin-induced inflammation remain unknown. In this work, random and site-directed mutagenesis of the inflammatory flagellin from enteroaggregative Escherichia coli identified two regions in the conserved D1 domain that are required for interleukin-8 release and TLR5 activation. In contrast, large regions of the variable domain could be excised without reducing the inflammatory activity. In addition, regions of the protein analogous to epitopes that trigger innate immune responses in plants are not involved in Caco-2 flagellin responses. These results highlight the complexity of the interaction between bacterial flagellin and its eukaryotic recognition partners and provide the basis for further studies to characterize the innate immune response to flagellin.
Division of Geographic and International Medicine, University of Virginia Health Sciences Center, Charlottesville, VA 22908, USA.
Meghan A
MA
Donnelly
Theodore S
TS
Steiner
D000595:Amino Acid Sequence; D001483:Base Sequence; D018938:Caco-2 Cells; D017931:DNA Primers; D029721:Drosophila Proteins; D004926:Escherichia coli / Q000378:metabolism*; D005408:Flagellin / Q000737:chemistry / Q000378:metabolism*; D006801:Humans; D016209:Interleukin-8 / Q000378:metabolism; D008562:Membrane Glycoproteins / Q000378:metabolism*; D008969:Molecular Sequence Data; D011956:Receptors, Cell Surface / Q000378:metabolism*; D051198:Toll-Like Receptor 5; D051193:Toll-Like Receptors
D017931:DNA Primers; D029721:Drosophila Proteins; D016209:Interleukin-8; D008562:Membrane Glycoproteins; D011956:Receptors, Cell Surface; C495346:TLR5 protein, human; D051198:Toll-Like Receptor 5; D051193:Toll-Like Receptors; D005408:Flagellin