Abstract
Salmonella can invade non-phagocytic cells through its type III secretion system (T3SS-1), which induces a Trigger entry process. This study showed that Salmonella enterica, subspecies enterica serovar Enteritidis can also invade cells via the Rck outer membrane protein. Rck was necessary and sufficient to enable non-invasive E. coli and Rck-coated beads to adhere to and invade different cells. Internalization analysis of latex beads coated with different Rck peptides showed that the peptide containing amino acids 140-150 promoted adhesion, whereas amino acids between 150 and 159 modulated invasion. Expression of dominant-negative derivatives and use of specific inhibitors demonstrated the crucial role of small GTPases Rac1 and Cdc42 in activating the Arp2/3 complex to trigger formation of actin-rich accumulation, leading to Rck-dependent internalization. Finally, scanning and transmission electron microscopy with Rck-coated beads and E. coli expressing Rck revealed microvillus-like extensions that formed a Zipper-like structure, engulfing the adherent beads and bacteria. Overall, our results provide new insights into the Salmonella T3SS-independent invasion mechanisms and strongly suggest that Rck induces a Zipper-like entry mechanism. Consequently, Salmonella seems to be the first bacterium found to be able to induce both Zipper and Trigger mechanisms to invade host cells.
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Acknowledgements
This work was supported by the Région Centre and performed within the 'SAVIRE' project, granted by the Délégation Régionale à la Recherche et à la Technologie du Centre (FEDER) (No 1634-32245) and by the Région Centre (No 2008-00036085). M Rosselin holds a Doctoral fellowship granted by the Région Centre and the Institut National de la Recherche Agronomique. We thank C Kieda for HBrMEC cell donation, Koreaki Ito (Kyoto University, Kyoto, Japan) for anti-SecG antibody donation, P Cossart and J Bertoglio for plasmid donation and J De Rycke for his critical reading of the manuscript.
Emmanuelle Caron passed away on July 8, 2009. We would like to take the opportunity to thank her for her guidance and suppport over the years. We will miss the friend and the great scientist!
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(Supplementary information is linked to the online version of the paper on the Cell Research website.)
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Supplementary information, Figure S1
Rck is able to induce adhesion to and internalization of a non invasive E. coli strain by human cells. (PDF 54 kb)
Supplementary information, Figure S2
rck deletion did not significantly reduce adhesion to and internalization of S. Enteritidis into MA104 and 3T3 cell lines. (PDF 82 kb)
Supplementary information, Figure S3
GST-113-159 Rck-coated beads are internalized by MA104 cells. (PDF 138 kb)
Supplementary information, Figure S4
Rck mediates a Zipper-like membrane rearrangement. (PDF 158 kb)
Supplementary information, Figure S5
The transduced TAT-N17Rac and -N17Cdc42 fusion proteins are active. (PDF 76 kb)
Supplementary information, Figure S6
G118D amino acid substitution in Rck inhibited adhesion and cell internalization. (PDF 37 kb)
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Rosselin, M., Virlogeux-Payant, I., Roy, C. et al. Rck of Salmonella enterica, subspecies enterica serovar Enteritidis, mediates Zipper-like internalization. Cell Res 20, 647–664 (2010). https://doi.org/10.1038/cr.2010.45
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DOI: https://doi.org/10.1038/cr.2010.45
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