Receptor-mediated binding of Pasteurella multocida dermonecrotic toxin to canine osteosarcoma and monkey kidney (vero) cells.
BACKGROUND
Binding and internalization of Pasteurella multocida dermonecrotic toxin (PMDT) by toxin-sensitive canine osteosarcoma and monkey kidney (vero) cells was examined ultrastructurally.


METHODS
Purified PMDT was conjugated to 20 nm colloidal gold particles in order to observe binding and internalization in the two cell lines at the ultrastructural level. The effects of various compounds on PMDT-vero binding were investigated to help elucidate the nature of putative vero cell receptors.


RESULTS
Colloidal gold-labeled PMDT was located at cell surfaces within 1 minute of its addition and rapidly transported to coated and noncoated invaginations of the plasma membrane. After extended incubation, gold particles were observed in endocytic vesicles, but not in any other intracellular structures. The magnitude of gold-PMDT cell association correlated with the cytotoxic sensitivity of the two cell lines. Early, but not late, addition of the lysosomotropic agent methylamine protected vero cells from the cytotoxic effects of PMDT without affecting binding. Biochemical and ultrastructural inhibition studies suggested the requirement for a ganglioside-type vero cell receptor.


CONCLUSIONS
This is the first report describing binding and internalization of PMDT in host cells. Biochemical and ultrastructural results suggest that PMDT interacts with a ganglioside-type receptor on vero cells and is transported to the cytosol in endocytic vesicles which do not appear to fuse with lysosomes.
National Animal Disease Center, U.S. Department of Agriculture, Ames, Iowa.
R K
RK
Pettit
M R
MR
Ackermann
R B
RB
Rimler
D000818:Animals; D001426:Bacterial Proteins; D001427:Bacterial Toxins / Q000037:antagonists & inhibitors / Q000378:metabolism*; D004285:Dogs; D006046:Gold; D008744:Methylamines / Q000494:pharmacology; D008854:Microscopy, Electron; D012516:Osteosarcoma / Q000378:metabolism* / Q000473:pathology / Q000648:ultrastructure; D016979:Pasteurella multocida; D011956:Receptors, Cell Surface / Q000378:metabolism*; D014407:Tumor Cells, Cultured; D014709:Vero Cells / Q000378:metabolism* / Q000648:ultrastructure
D001426:Bacterial Proteins; D001427:Bacterial Toxins; D008744:Methylamines; C062258:Pasteurella multocida toxin; D011956:Receptors, Cell Surface; D006046:Gold; C027451:methylamine