title: Hierarchies of host factor dynamics at the entry site of Shigella flexneri during host cell invasion.
abstract: Shigella flexneri, the causative agent of bacillary dysentery, induces massive cytoskeletal rearrangement, resulting in its entry into nonphagocytic epithelial cells. The bacterium-engulfing membrane ruffles are formed by polymerizing actin, a process activated through injected bacterial effectors that target host small GTPases and tyrosine kinases. Once inside the host cell, S. flexneri escapes from the endocytic vacuole within minutes to move intra- and intercellularly. We quantified the fluorescence signals from fluorescently tagged host factors that are recruited to the site of pathogen entry and vacuolar escape. Quantitative time lapse fluorescence imaging revealed simultaneous recruitment of polymerizing actin, small GTPases of the Rho family, and tyrosine kinases. In contrast, we found that actin surrounding the vacuole containing bacteria dispersed first from the disassembling membranes, whereas other host factors remained colocalized with the membrane remnants. Furthermore, we found that the disassembly of the membrane remnants took place rapidly, within minutes after bacterial release into the cytoplasm. Superresolution visualization of galectin 3 through photoactivated localization microscopy characterized these remnants as small, specular, patchy structures between 30 and 300 nm in diameter. Using our experimental setup to track the time course of infection, we identified the S. flexneri effector IpgB1 as an accelerator of the infection pace, specifically targeting the entry step, but not vacuolar progression or escape. Together, our studies show that bacterial entry into host cells follows precise kinetics and that this time course can be targeted by the pathogen.
authors.affiliation: Group Dynamics of Host-Pathogen Interactions, Institut Pasteur, Paris, France.
authors.forename: Soudeh
authors.initials: S
authors.lastname: Ehsani
authors.forename: José Carlos
authors.initials: JC
authors.lastname: Santos
authors.forename: Cristina D
authors.initials: CD
authors.lastname: Rodrigues
authors.forename: Ricardo
authors.initials: R
authors.lastname: Henriques
authors.forename: Laurent
authors.initials: L
authors.lastname: Audry
authors.forename: Christophe
authors.initials: C
authors.lastname: Zimmer
authors.forename: Philippe
authors.initials: P
authors.lastname: Sansonetti
authors.forename: Guy
authors.initials: G
authors.lastname: Tran Van Nhieu
authors.forename: Jost
authors.initials: J
authors.lastname: Enninga
mesh_terms: D003593:Cytoplasm / Q000382:microbiology; D003599:Cytoskeleton / Q000378:metabolism*; D004847:Epithelial Cells / Q000382:microbiology*; D006367:HeLa Cells; D054884:Host-Pathogen Interactions; D006801:Humans; D008856:Microscopy, Fluorescence; D012763:Shigella flexneri / Q000472:pathogenicity*; D013997:Time Factors; D059008:Time-Lapse Imaging; D014617:Vacuoles / Q000382:microbiology