Intracellular signalling and cytoskeletal rearrangement involved in Yersinia pestis plasminogen activator (Pla) mediated HeLa cell invasion.
Yersinia pestis, the etiologic agent of plague is a highly invasive organism being able to invade non-phagocytic epithelial cells. Its plasminogen activator (Pla), encoded by the pPCP1 plasmid plays a pivotal role in internalisation of bacteria by HeLa cells. The aim of this study was to analyse the intracellular signalling processes and cytoskeletal rearrangement events associated with invasion. Wortmannin caused a 50% decrease of invasiveness at 50nM concentration pointing to the involvement of phosphatidyl-inosinol-4 kinase (PtINs4). Pre-treatment with staurosporin, a potent inhibitor of protein kinases (PKs) and with genistein, a specific tyrosine kinase inhibitor decreased the number of internalised bacteria about seven-fold and two-fold, respectively, indicating the involvement of PKs including tyrosine kinases in Pla-mediated internalisation. Cytochalasin D, an actin polymerisation inhibitor, C3 exoenzyme of Clostridium botulinum, a specific inhibitor of small GTPase Rho, and NDGA, a 5-lipoxygenase inhibitor also involved in Rho activation strongly reduced the number of internalised bacteria revealing the role of cytoskeletal events in the invasion process. All the tested inhibitors changed the invasion but not the adhesion pattern of the Pla producing recombinant strain. Actin rearrangement could also be visualised also with rhodamin-phalloidin staining.
Benedek
Orsolya
O
Department of Medical Microbiology and Immunology, Faculty of Medicine, University of Pécs, Szigeti út 12., H-7624 Pécs, Hungary.
Nagy
Gábor
G
Emody
Levente
L
D036002:ADP Ribose Transferases / Q000633:toxicity; D008841:Actin Cytoskeleton / Q000378:metabolism / Q000648:ultrastructure; D000199:Actins / Q000378:metabolism; D000730:Androstadienes / Q000494:pharmacology; D001422:Bacterial Adhesion; D001426:Bacterial Proteins / Q000378:metabolism*; D001905:Botulinum Toxins / Q000633:toxicity; D015169:Colony Count, Microbial; D015638:Cytochalasin D / Q000494:pharmacology; D003593:Cytoplasm / Q000382:microbiology; D003599:Cytoskeleton / Q000378:metabolism* / Q000648:ultrastructure; D004791:Enzyme Inhibitors / Q000494:pharmacology; D019833:Genistein / Q000494:pharmacology; D006367:HeLa Cells; D006801:Humans; D016859:Lipoxygenase Inhibitors; D000081082:Phosphoinositide-3 Kinase Inhibitors; D010960:Plasminogen Activators / Q000378:metabolism*; D047428:Protein Kinase Inhibitors; D011505:Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors; D015398:Signal Transduction; D019311:Staurosporine / Q000494:pharmacology; D000077191:Wortmannin; D015010:Yersinia pestis / Q000201:enzymology / Q000472:pathogenicity*; D020741:rho GTP-Binding Proteins / Q000037:antagonists & inhibitors
D000199:Actins; D000730:Androstadienes; D001426:Bacterial Proteins; D004791:Enzyme Inhibitors; D016859:Lipoxygenase Inhibitors; D000081082:Phosphoinositide-3 Kinase Inhibitors; D047428:Protein Kinase Inhibitors; D015638:Cytochalasin D; D019833:Genistein; D036002:ADP Ribose Transferases; C063007:exoenzyme C3, Clostridium botulinum; D011505:Protein-Tyrosine Kinases; C078140:Pla protease, Yersinia pestis; D010960:Plasminogen Activators; D001905:Botulinum Toxins; D020741:rho GTP-Binding Proteins; D019311:Staurosporine; D000077191:Wortmannin