Host-Pathogen Interactions Effecting Entry

Elizabeth M. Adler

Science Signaling, AAAS, Washington, DC 20005, USA

Many extracellular bacteria secrete toxins that bind membrane proteins on the target cell, which allows the toxins to be endocytosed, thereby allowing catalytically active moieties to gain access to the cytoplasm. Anthrax toxin binds to two different receptors [the type I membrane proteins TEM8 (tumor endothelial marker 8) and CMG2 (capillary morphogenesis gene 2)] through its protective antigen (PA) subunit, enabling clathrin-dependent endocytosis of its two catalytically active subunits [lethal factor (LF) and edema factor (EF)] as part of a hetero-oligomeric complex. Noting that endocytosis of the toxin-bound anthrax receptors was rapid compared to that of toxin-free receptors, Abrami et al. investigated the possibility that toxin binding triggers signaling events that facilitate its own endocytosis. Western analysis of HeLa cell extracts revealed that PA elicited tyrosine phosphorylation; a combination of immunoprecipitation, SDS/PAGE, and Western analysis revealed that phosphorylation of clathrin heavy chain preceded the appearance of PA in endosomes (assessed by existence of an SDS-resistant PA heptamer). The tyrosine phosphatase inhibitor genistein blocked PA endocytosis and evidence of LF intracellular activity [mitogen-activated protein kinase kinase 1 (MEK1) cleavage] without blocking binding or oligimerization of PA or LF binding (which depends on PA oligimerization). PA elicited CMG2 tyrosine phosphorylation but not that of a mutant form in which four intracellular tyrosine residues were replaced with alanines (CMG2^Y). PA endocytosis and MEK1 cleavage were delayed in anthrax receptor–deficient CHO cells transfected with CMG2^Y compared with those transfected with wild-type CMG2, and CMG2 ubiquitination, which is required for its endocytosis, was attenuated. PA elicited the phosphorylation of Src-like kinases (SLKs). Moreover, analyses of SLK knockdown or of mouse embryonic fibroblasts lacking the SLKs Src, Fyn, and Yes implicated Src, Fyn, or both in PA-dependent CMG2 phosphorylation and MEK1 cleavage and toxin endocytosis. The authors thus conclude that anthrax toxin stimulates signaling through SLKs to facilitate its entry into the target cell.

L. Abrami, B. Kunz, F. Gisou van der Goot, Anthrax toxin triggers the activation of src-like kinases to mediate its own uptake. Proc. Natl. Acad. Sci. USA 107, 1420–1424 (2010). [Abstract] [Full Text]

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