Role of CypA and Hsp90 in membrane translocation mediated by anthrax protective antigen

Lydia Dmochewitz, Maren Lillich, Eva Kaiser, Laura D. Jennings, Alexander E. Lang, Johannes Buchner, Gunter Fischer, Klaus Aktories, R. John Collier, Holger Barth

Research output: Contribution to journalArticlepeer-review

54 Scopus citations

Abstract

Bacillus anthracis lethal toxin consists of the protective antigen (PA) and the metalloprotease lethal factor (LF). During cellular uptake PA forms pores in membranes of endosomes, and unfolded LF translocates through the pores into the cytosol. We have investigated whether host cell chaperones facilitate translocation of LF and the fusion protein LFNDTA. LFN mediates uptake of LFNDTA into the cytosol, where DTA, the catalytic domain of diphtheria toxin, ADP-ribosylates elongation factor-2, allowing for detection of small amounts of translocated LFNDTA. Cyclosporin A, which inhibits peptidyl-prolyl cis/trans isomerase activity of cyclophilins, and radicicol, which inhibits Hsp90 activity, prevented uptake of LFNDTA into the cytosol of CHO-K1 cells and protected cells from intoxication by LFNDTA/PA. Both inhibitors, as well as an antibody against cyclophilin A blocked the release of active LFNDTA from endosomal vesicles into the cytosol in vitro. In contrast, the inhibitors did not inhibit cellular uptake of LF. In vitro, cyclophilin A and Hsp90 bound to LFNDTA and DTA but not to LF, implying that DTA determines this interaction. In conclusion, cyclophilin A and Hsp90 facilitate translocation of LFNDTA, but not of LF, across endosomal membranes, and thus they function selectively in promoting translocation of certain proteins, but not of others.

Original languageEnglish
Pages (from-to)359-373
Number of pages15
JournalCellular Microbiology
Volume13
Issue number3
DOIs
StatePublished - Mar 2011

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