Interferon alpha inhibits a Src-mediated pathway necessary for Shigella-induced cytoskeletal rearrangements in epithelial cells.
Bottom Line: Shigella flexneri, the causative agent of bacillary dysentery, has the ability to enter nonphagocytic cells.The interferon (IFN) family of cytokines was found to inhibit Shigella invasion of cultured epithelial cells.Immunofluorescence studies showed that IFN-alpha inhibits Shigella-induced actin polymerization required for bacterial entry into cells.
Affiliation: Unité de Génétique Humaine, INSERM U276.
Shigella flexneri, the causative agent of bacillary dysentery, has the ability to enter nonphagocytic cells. The interferon (IFN) family of cytokines was found to inhibit Shigella invasion of cultured epithelial cells. We show here that IFN-alpha inhibits a Src-dependent signaling cascade triggered by Shigella that leads to the reorganization of the host cell cytoskeleton. Immunofluorescence studies showed that IFN-alpha inhibits Shigella-induced actin polymerization required for bacterial entry into cells. Phosphorylation of cortactin, a Src-substrate specifically tyrosyl-phosphorylated during Shigella entry, was inhibited by IFN-alpha. Overexpression of a dominant interfering form of pp60c-src led to inhibition of Shigella-induced cytoskeletal rearrangements and decreased cortactin phosphorylation indicating a role for Src in Shigella entry. Also, Shigella uptake in cells that expressed constitutively active Src was unaffected by IFN-alpha treatment. We conclude that Src kinase activity is necessary for Shigella invasion of epithelial cells and that IFN-alpha inhibits this Src-dependent signaling pathway.
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Mentions: As shown above, IFN-α treatment induces a decrease in Shigella-induced cortactin phosphorylation suggesting that IFN-α inhibits Shigella entry by affecting Src activation. To further define the relationship between IFN-α inhibition of Shigella entry and Src, we investigated the effects of IFN-α on cells that express a constitutively active form of Src (Cartwright et al., 1987). HeLa cells were transfected with an allele of src bearing a mutation at position 527 that substituted a tyrosine into a phenyl alanine (Y527F). Stable clones were selected (srcF) and their relative levels of Src was analyzed by Western blot. Overexpression of Src was about two- to threefold relative to the amounts in the parental cells (Fig. 8 b, Src). When challenged with Shigella, srcF cells shared characteristics similar to srcK+ cells. The kinetics of foci formation (Fig. 7 g, diamonds), the efficiency of Shigella entry and the phosphorylation pattern (not shown) after Shigella invasion were indistinguishable in srcF cells and in srcK+ cells.