Inhibition of small G proteins by clostridium sordellii lethal toxin activates cdc2 and MAP kinase in Xenopus oocytes.
The lethal toxin (LT) from Clostridium sordellii is a glucosyltransferase that modifies and inhibits small G proteins of the Ras family, Ras and Rap, as well as Rac proteins. LT induces cdc2 kinase activation and germinal vesicle breakdown (GVBD) when microinjected into full-grown Xenopus oocytes. Toxin B from Clostridium difficile, that glucosylates and inactivates Rac proteins, does not induce cdc2 activation, indicating that proteins of the Ras family, Ras and/or Rap, negatively regulate cdc2 kinase activation in Xenopus oocyte. In oocyte extracts, LT catalyzes the incorporation of [14C]glucose into a group of proteins of 23 kDa and into one protein of 27 kDa. The 23-kDa proteins are recognized by anti-Rap1 and anti-Rap2 antibodies, whereas the 27-kDa protein is recognized by several anti-Ras antibodies and probably corresponds to K-Ras. Microinjection of LT into oocytes together with UDP-[14C]glucose results in a glucosylation pattern similar to the in vitro glucosylation, indicating that the 23- and 27-kDa proteins are in vivo substrates of LT. In vivo time-course analysis reveals that the 27-kDa protein glucosylation is completed within 2 h, well before cdc2 kinase activation, whereas the 23-kDa proteins are partially glucosylated at GVBD. This observation suggests that the 27-kDa Ras protein could be the in vivo target of LT allowing cdc2 kinase activation. Interestingly, inactivation of Ras proteins does not prevent the phosphorylation of c-Raf1 and the activation of MAP kinase that occurs normally around GVBD.
Rime
H
H
INRA/ESA-CNRS 7080, Université Pierre et Marie Curie, 4 place Jussieu, 75252 Paris Cédex 05, France.
Talbi
N
N
Popoff
M R
MR
Suziedelis
K
K
Jessus
C
C
Ozon
R
R
D000818:Animals; D001427:Bacterial Toxins / Q000633:toxicity*; D016203:CDC2 Protein Kinase / Q000378:metabolism*; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000378:metabolism*; D003013:Clostridium; D004789:Enzyme Activation / Q000187:drug effects; D005260:Female; D019204:GTP-Binding Proteins / Q000037:antagonists & inhibitors*; D009865:Oocytes / Q000187:drug effects / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects*; D014981:Xenopus
D001427:Bacterial Toxins; C072292:lethal toxin LT, Clostridium sordellii; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D016203:CDC2 Protein Kinase; D019204:GTP-Binding Proteins