Inhibition of cytokinesis by Clostridium difficile toxin B and cytotoxic necrotizing factors--reinforcing the critical role of RhoA in cytokinesis.
Low molecular weight GTP-binding proteins of the Rho family control the organization of the actin cytoskeleton in eukaryotic cells. RhoA governs the formation of actin stress fibers and is responsible for the formation of the contractile ring in cytokinesis. Cytokinesis completion requires RhoA inactivation resulting in disassembly of the contractile ring. Cytokinesis thus requires switching of RhoA activity. This switch of RhoA activity is blocked by Rho-modifying bacterial protein toxins that either activate or inactivate RhoA by covalent modifications. Exoenzyme C3 from Clostridium limosum (C3-lim) and Clostridium difficile toxin B (TcdB) inactivate RhoA by mono-ADP-ribosylation and mono-glucosylation, respectively. Cytotoxic necrotizing factors (CNF), produced by either Yersinia pseudotuberculosis (CNFY) or uropathogenic strains of E. coli (CNF1), deamidate and thereby activate RhoA. This study provides evidence that RhoA-activating as well as RhoA-inactivating toxins cause inhibition of cytokinesis and cell division. The toxins' effects on cytokinesis were analyzed in Hela cells synchronized using the thymidine double block technique. Treatment of G2-phase cells with either the RhoA-activating CNFY or CNF1 or the RhoA-inactivating C3-lim or TcdB resulted in cytokinesis inhibition, as evidenced by the formation of a 4N population on flow cytometry, the inhibition of contractile ring formation, and the formation of bi-nucleated cells. While TcdB and CNF1 modify a broad-spectrum of Rho proteins, C3-lim and CNFY specifically target RhoA. Since C3-lim and CNFY both caused cytokinesis inhibition, our study re-inforces the critical role of RhoA (not Rac1 or Cdc42) in cytokinesis and cell division.
Huelsenbeck
Stefanie C
SC
Institute of Toxicology, Hannover Medical School, Hannover, Germany.
May
Martin
M
Schmidt
Gudula
G
Genth
Harald
H
D000199:Actins / Q000502:physiology; D001426:Bacterial Proteins / Q000494:pharmacology*; D001427:Bacterial Toxins / Q000494:pharmacology*; D002467:Cell Nucleus / Q000187:drug effects / Q000473:pathology; D048749:Cytokinesis / Q000187:drug effects*; D029968:Escherichia coli Proteins / Q000494:pharmacology*; D006367:HeLa Cells; D006801:Humans; D020742:rhoA GTP-Binding Protein / Q000378:metabolism*
D000199:Actins; D001426:Bacterial Proteins; D001427:Bacterial Toxins; D029968:Escherichia coli Proteins; C031815:toxB protein, Clostridium difficile; C060296:cytotoxic necrotizing factor type 1; D020742:rhoA GTP-Binding Protein