Receptor kinetics differ for endothelin-1 and endothelin-2 binding to Swiss 3T3 fibroblasts.
The equilibrium binding, kinetics of ligand-receptor interactions, and biological activity of endothelin-1 and -2 have been studied in Swiss 3T3 fibroblasts. Scatchard analyses of saturation binding data for ET-1 and -2, performed at 4 degrees C to prevent internalization of the occupied receptor, revealed similar affinity constants and numbers of binding sites for endothelin-1 and -2. Experiments designed to determine ligand-induced effects on 45Ca efflux demonstrated no qualitative or quantitative differences between the two endothelin isoforms. In contrast, kinetic studies resulted in different rates of dissociation for the two isoforms and different extents of dissociation. Specifically, only 40% of the bound [125I]endothelin-1 was dissociated at 4 h following the addition of excess unlabeled ligand, whereas 85-90% of the bound [125I]endothelin-2 was dissociated under the same conditions. Endothelin-1 and -2 also differed in the percent of specific cell-associated ligand bound after a 2 h incubation at 37 degrees C following an initial equilibration at 4 degrees C. The differences in dissociation rates and association or internalization rates at 37 degrees C are the first data that differentiate between the two isoforms. It is suggested that isoform-specific differences in the rate of dissociation from cell surface endothelin receptors influence the level of cell-associated endothelin and may be important in determining physiologic responses in vivo.
Devesly
P
P
Phillips
P E
PE
Johns
A
A
Rubanyi
G
G
Parker-Botelho
L H
LH
D000818:Animals; D002460:Cell Line; D004705:Endocytosis; D016232:Endothelins / Q000378:metabolism*; D005347:Fibroblasts / Q000378:metabolism; D007700:Kinetics; D051379:Mice; D011956:Receptors, Cell Surface / Q000378:metabolism*; D017466:Receptors, Endothelin; D013379:Substrate Specificity; D013696:Temperature
D016232:Endothelins; D011956:Receptors, Cell Surface; D017466:Receptors, Endothelin