Pasteurella multocida toxin is a potent inducer of anchorage-independent cell growth.
The growth of many normal cells requires contact with an adhesive substratum, a requirement that is frequently abrogated in the transformed phenotype. We have explored pathways that can lead to the anchorage-independent growth of cultured Rat-1 fibroblasts. Pasteurella multocida toxin (PMT), a 146-kDa mitogenic protein, caused a striking increase in the formation of colonies (greater than 200 microns) from single cells in soft agar. The magnitude of the effect of PMT was greater than that achieved by epidermal growth factor or platelet-derived growth factor. The toxin was extremely potent, with half-maximal and maximal effects observed at 1 and 10 pM PMT, respectively. This concentration dependence of the action of the toxin is similar to that for the stimulation of DNA synthesis in adherent cultures of the cells. Stimulation of colony formation could be achieved by a transient exposure of the cells to PMT and it was blocked by methylamine, indicating that the toxin enters the cells to act. Colony formation was stimulated equally by native and recombinant PMT, but a truncated version (33.5 kDa) of the recombinant toxin was ineffective. PMT antiserum blocked colony formation in response to PMT. In the Rat-1 cells, PMT stimulated the phospholipase C-mediated hydrolysis of inositolphospholipids, as indicated by the stimulation of inositol phosphate release, Ca2+ mobilization, and phosphorylation of a protein kinase C substrate. The results indicate that the deregulation of signal-transduction pathways as elicited by an intracellularly acting bacterial toxin can induce a malignant phenotype.
Higgins
T E
TE
Imperial Cancer Research Fund, Lincoln's Inn Fields, London, United Kingdom.
Murphy
A C
AC
Staddon
J M
JM
Lax
A J
AJ
Rozengurt
E
E
D000818:Animals; D001426:Bacterial Proteins; D001427:Bacterial Toxins / Q000494:pharmacology*; D002118:Calcium / Q000378:metabolism*; D002448:Cell Adhesion / Q000187:drug effects; D002455:Cell Division / Q000187:drug effects*; D002460:Cell Line; D002999:Clone Cells; D004261:DNA Replication / Q000187:drug effects*; D004815:Epidermal Growth Factor / Q000187:drug effects / Q000378:metabolism; D007295:Inositol Phosphates / Q000378:metabolism*; D007700:Kinetics; D010325:Pasteurella; D015240:Phorbol 12,13-Dibutyrate / Q000494:pharmacology; D010766:Phosphorylation; D051381:Rats; D011994:Recombinant Proteins / Q000494:pharmacology; D013936:Thymidine / Q000378:metabolism; D014316:Tritium
D001426:Bacterial Proteins; D001427:Bacterial Toxins; D007295:Inositol Phosphates; C062258:Pasteurella multocida toxin; D011994:Recombinant Proteins; D014316:Tritium; D015240:Phorbol 12,13-Dibutyrate; D004815:Epidermal Growth Factor; D002118:Calcium; D013936:Thymidine