Escherichia coli cyclomodulin Cif induces G2 arrest of the host cell cycle without activation of the DNA-damage checkpoint-signalling pathway.
The cycle inhibiting factor (Cif) belongs to a family of bacterial toxins and effector proteins, the cyclomodulins, that deregulate the host cell cycle. Upon injection into HeLa cells by the enteropathogenic Escherichia coli (EPEC) type III secretion system, Cif induces a cytopathic effect characterized by the recruitment of focal adhesion plates and the formation of stress fibres, an irreversible cell cycle arrest at the G(2)/M transition, and sustained inhibitory phosphorylation of mitosis inducer, CDK1. Here, we report that the reference typical EPEC strain B171 produces a functional Cif and that lipid-mediated delivery of purified Cif into HeLa cells induces cell cycle arrest and actin stress fibres, implying that Cif is necessary and sufficient for these effects. EPEC infection of intestinal epithelial cells (Caco-2, IEC-6) also induces cell cycle arrest and CDK1 inhibition. The effect of Cif is strikingly similar to that of cytolethal distending toxin (CDT), which inhibits the G(2)/M transition by activating the DNA-damage checkpoint pathway. However, in contrast to CDT, Cif does not cause phosphorylation of histone H2AX, which is associated with DNA double-stranded breaks. Following EPEC infection, the checkpoint effectors ATM/ATR, Chk1 and Chk2 are not activated, the levels of the CDK-activating phosphatases Cdc25B and Cdc25C are not affected, and Cdc25C is not sequestered in host cell cytoplasm. Hence, Cif activates a DNA damage-independent signalling pathway that leads to inhibition of the G(2)/M transition.
Taieb
Frédéric
F
UMR 1225, IHAP, INRA-ENVT, 23 Chemin des Capelles, BP 87614, 31000 Toulouse, France. f.taieb@envt.fr
Nougayrède
Jean-Philippe
JP
Watrin
Claude
C
Samba-Louaka
Ascel
A
Oswald
Eric
E
D000818:Animals; D001427:Bacterial Toxins / Q000378:metabolism; D018938:Caco-2 Cells; D002453:Cell Cycle; D018797:Cell Cycle Proteins / Q000378:metabolism; D000071877:Checkpoint Kinase 1; D064447:Checkpoint Kinase 2; D053903:DNA Breaks, Double-Stranded; D004249:DNA Damage; D004926:Escherichia coli / Q000378:metabolism / Q000472:pathogenicity*; D029968:Escherichia coli Proteins / Q000378:metabolism*; D016195:G2 Phase; D006367:HeLa Cells; D006801:Humans; D007413:Intestinal Mucosa / Q000382:microbiology*; D011494:Protein Kinases / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D011817:Rabbits; D051381:Rats; D015398:Signal Transduction; D020687:cdc25 Phosphatases / Q000378:metabolism
D001427:Bacterial Toxins; D018797:Cell Cycle Proteins; C481998:Cif protein, E coli; D029968:Escherichia coli Proteins; C067044:cytolethal distending toxin; D011494:Protein Kinases; D064447:Checkpoint Kinase 2; C000605867:CHEK1 protein, human; C578634:CHEK2 protein, human; D000071877:Checkpoint Kinase 1; C000605871:Chek1 protein, rat; C578632:Chek2 protein, rat; D017346:Protein Serine-Threonine Kinases; C517008:CDC25B protein, human; C517009:CDC25C protein, human; D020687:cdc25 Phosphatases