> Pathogénie microbienne moléculaire - INSERM U389
• Summary
• Saga Shigella
• Objectives
• Genetics
• Inflammation
• Immunology
• Vaccines
2 - Molecules et signals involved in Shigella entry into epithelial cells and cell-to cell spread.
Permanent scientist: Guy Tran Van Nhieu Directeur de Recherche, INSERM).
Post-doctoral scientists: Nalini Ramarao, Caroline Clair.
Graduate student : Laurence Bougnères.
Technician : Joëlle Mounier.
Shigella is a paradigm of entry into non-phagocytic cells via a “triggered” signalling process inducing the formation of a macropinocytic pocket that requires the formation of a “signaling” platform in the eukaryotic cell membrane, following the insertion of the IpaB-IpaC pore. The role of CD44, as a surface receptor interacting with IpaB, has been demonstrated ( ). CD44 is expressed on the basolateral membrane of the epithelium, consistent with the early demonstration that Shigella enters essentially through the baso-lateral pole of polarised epithelial cells ( ).
Also consistent with this observation is the demonstration by Frank Lafont and Gisou VanDerGoot, in collaboration with PMM, that this signaling platform requires the clustering of rafts, extraction of membrane cholesterol blocking both the binding and entry of Shigella ( ). Subsequent to this early stage, the actin cytoskeletal rearrangements triggered by the entering microorganism have been described in detail ( ). showing that both small GTPases of the Rho family ( , ) and the protooncogene p60c-src through tyrosine phosphorylation of cortactin ( , , ), are key elements of the signalling pathways leading to actin nucleation/polymerization. IpaC, once inserted into the epithelial cell membrane, is the triggering molecule that activates the Cdc42/Rac-1 and the c-Src pathways ( , Mounier et al., in preparation).
The elongation process of cellular extensions is furthered by the translocation of IpgD, a phospahtidyl-inositol phosphatase that decreases the surface tension between the cortical cytoskeleton and the membrane ( , ). Following this step of massive cytoskeletal extension, Shigella injects IpaA into the cell. This protein binds the N-terminal head-domain of vinculin with high affinity, thus causing the unfolding of this actin binding protein, and leading to the shortening and bundling of actin filaments in an adherence-plaque like structure essential to finalize the entry process ( , ).
Following the initial demonstration of its actin-based intracellular motility, Shigella has also become a paradigm to study how pathogens move intracellularly and spread from cell to cell. We have shown that Shigella could move like organelles along stress cables and called it the Organelle-Like Movement ( ). Moreover, in association with the group of Marie-France Carlier (CNRS, Gif), the molecular mechanisms by which IcsA induces actin nucleation via N-WASP and the Arp2/3 complex ( ). and causes rapid bacterial movement in the cytoplasm has been deciphered. This movement results in cell to cell spread of the bacteria ( ). The passage from one cell to the other involves engagement of the bacterium with components of the cell junction such as cadherins and catenins ( ). This leads to active, myosin II-dependant phagocytosis of the protrusion ( ). final escape into the cytoplasm of the adjacent cell requires secretion of Ipa proteins ( , ). thus constituting a highly coordinated process. In addition, expression of connexins by epithelial cells is required for efficient cell-to –cell spread to occur. Formation of gap junctions and hemi-connexons is essential for the transmission of signals from the infected cell to make adjacent cells of the epithelial lining more permissive to bacterial invasion. ATP is a major mediator of this process (Tran Van Nhieu et al., submitted).