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: To analyze more specifically the effect of IFN-α on the entry process, we first tested the capacity of HeLa cells to internalize Shigella in a gentamicin protection assay carried out after a short incubation time. Cells were treated with IFN-α for 20 h, then challenged with Shigella for 30 min before adding gentamicin (Materials and Methods). As shown in Fig. 1 a, IFN-α treatment at a concentration of 500 U/ml led to a 70% inhibition of Shigella entry compared with untreated cells. A slightly reduced effect was seen with 50 U/ml IFN-α (Fig. 1 a, 50). Under comparable conditions, Salmonella entry into cells showed little inhibition, with no significant differences between the 50 and 500 U/ml treatments (Fig. 1 b). These results indicate that IFN-α specifically impairs the ability of cells to internalize Shigella.