The Gram-negative coccobacillus Kingella kingae is increasingly recognized as an important invasive pediatric pathogen that causes mostly bacteremia and skeletal system infections. K. kingae secretes an RtxA toxin that belongs to a broad family of the RTX (Repeats in ToXin) cytotoxins produced by bacterial pathogens. Recently, we demonstrated that membrane cholesterol facilitates interaction of RtxA with target cells, but other cell surface structures potentially involved in toxin binding to cells remain unknown. We show that deglycosylation of cell surface structures by glycosidase treatment, or inhibition of protein N- and O-glycosylation by chemical inhibitors substantially reduces RtxA binding to target cells. Consequently, the deglycosylated cells were more resistant to cytotoxic activity of RtxA. Moreover, experiments on cells expressing or lacking cell surface integrins of the β2 family revealed that, unlike some other cytotoxins of the RTX family, K. kingae RtxA does not bind target cells via the β2 integrins. Our results, hence, show that RtxA binds cell surface oligosaccharides present on all mammalian cells but not the leukocyte-restricted β2 integrins. This explains the previously observed interaction of the toxin with a broad range of cell types of various mammalian species and reveals that RtxA belongs to the group of broadly cytolytic RTX hemolysins.

Department of Basic and Translational Sciences School of Dental Medicine University of Pennsylvania 240 S 40th St Philadelphia PA 19104 USA

Department of Biochemistry Faculty of Science Charles University Prague Hlavova 8 128 43 Prague 2 Czech Republic

Institute of Microbiology of the Czech Academy of Sciences Videnska 1083 142 20 Prague 4 Czech Republic

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A conserved tryptophan in the acylated segment of RTX toxins controls their β2 integrin-independent cell penetration

Filamentous Hemagglutinin of Bordetella pertussis Does Not Interact with the β2 Integrin CD11b/CD18

Kingella kingae RtxA Cytotoxin in the Context of Other RTX Toxins