Cytotoxic necrotizing factor-2 of Escherichia coli alters the morphology of cultured hippocampal neurons.
Certain pathogenic strains of E. coli produce the cytotoxic necrotizing factors-1 or -2. Cytotoxic necrotizing factor-1 irreversibly activates the small GTPases of the Rho family Rho, Rac and Cdc42. Cytotoxic necrotizing factor-2 may have similar effects. Since the Rho proteins play an important role in the organization of the actin cytoskeleton and neuronal differentiation, we have investigated whether cytotoxic necrotizing factor-2 affects the morphology of cultured hippocampal neurons. The toxin indeed caused dendrite retraction and axon shortening. Within 4 h of application, cytotoxic necrotizing factor-2 induced a transient formation of short finger-like extensions. To study the role of the Rho proteins in the morphological changes caused by cytotoxic necrotizing factor-2, we transfected neurons with recombinant Rho proteins. Dominant-negative forms of Rac or Rho but not of Cdc42 prevented the formation of short extensions induced by cytotoxic necrotizing factor-2, indicating synergistic effects of Rac and Rho. In contrast, the retraction of dendrites induced by cytotoxic necrotizing factor-2 was only prevented by dominant-negative Rho. Analysis with pull-down assays showed that cytotoxic necrotizing factor-2 strongly activated Rac and Rho, whereas an effect on Cdc42 was not observed. Cytotoxic necrotizing factor-2 also diminished the total amount of Rac and Rho. The degradation of Rac was so pronounced that the increase in Rac activity was only transient. In organotypic cultures of the hippocampus, cytotoxic necrotizing factor-2 reduced the number of neurites per neuron, suggesting that neurons in the tissue context were also vulnerable. We conclude that cytotoxic necrotizing factor-2 has pronounced effects on neuronal morphology, which are due to activation of the GTPases Rho and Rac.
Boutillier
S
S
Institut für Experimentelle und Klinische Pharmakologie und Toxikologie, Albert-Ludwigs-Universität, Albertstrasse 23, Zentrum für Neurowissenschaften, 79104 Freiburg, Germany.
Rapp
J
J
Staeb
T
T
Olenik
C
C
Schmidt
G
G
Meyer
D K
DK
Leemhuis
J
J
D000577:Amides / Q000494:pharmacology; D000818:Animals; D000831:Animals, Newborn; D001427:Bacterial Toxins / Q000494:pharmacology*; D002478:Cells, Cultured; D015638:Cytochalasin D / Q000494:pharmacology; D003603:Cytotoxins / Q000494:pharmacology*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004926:Escherichia coli / Q000737:chemistry*; D029968:Escherichia coli Proteins; D006624:Hippocampus / Q000166:cytology / Q000187:drug effects*; D047908:Intracellular Signaling Peptides and Proteins; D009474:Neurons / Q000166:cytology / Q000187:drug effects* / Q000648:ultrastructure; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors; D011725:Pyridines / Q000494:pharmacology; D051381:Rats; D017208:Rats, Wistar; D011994:Recombinant Proteins / Q000494:pharmacology; D014162:Transfection; D014404:Tubulin / Q000378:metabolism / Q000648:ultrastructure; D020764:cdc42 GTP-Binding Protein / Q000378:metabolism; D020744:rac GTP-Binding Proteins / Q000378:metabolism; D020741:rho GTP-Binding Proteins / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology; D054460:rho-Associated Kinases
D000577:Amides; D001427:Bacterial Toxins; D003603:Cytotoxins; D004791:Enzyme Inhibitors; D029968:Escherichia coli Proteins; D047908:Intracellular Signaling Peptides and Proteins; D011725:Pyridines; D011994:Recombinant Proteins; D014404:Tubulin; C086995:cytotoxic necrotizing factor type 2; C060296:cytotoxic necrotizing factor type 1; C108830:Y 27632; D015638:Cytochalasin D; D017346:Protein Serine-Threonine Kinases; D054460:rho-Associated Kinases; D020764:cdc42 GTP-Binding Protein; D020744:rac GTP-Binding Proteins; D020741:rho GTP-Binding Proteins