Characterization of receptors for alpha-melanocyte-stimulating hormone on human melanoma cells.
Receptors for alpha-melanocyte-stimulating hormone (alpha-MSH) on human malignant melanoma cell lines were investigated with a specific binding assay and characterized with structural analogues of alpha-MSH and adrenocorticotropic hormone and by photoaffinity cross-linking of the hormone-receptor complex. Specific binding of high-performance liquid chromatography-purified, monoiodinated alpha-MSH in the presence of 1 mM 1,10-phenanthroline as protease inhibitor was highest after a 2-h incubation at 37 degrees C. The nonspecific binding was less than 20% and dissociation of the ligand-receptor complex was relatively slow. Ten out of 12 human cell lines showed specific binding sites for alpha-MSH with Kp values ranging from 0.195 to 2.87 nM and the sites/cell being approximately 400 to approximately 1600. Virtually identical results were obtained in an assay where the cells remained attached to the culture dishes during the entire experiment. The study of hormone analogues with the D10 cell line showed that oxidized alpha-MSH had an approximately 40-fold lower affinity than alpha-MSH whereas [Nle4,D-Phe7]-alpha-MSH displayed a threefold and the adrenocorticotropic hormone fragments (1-17) and (1-24) a 20- and 8-fold higher affinity. Cross-linking of the alpha-MSH-receptor complex of three cell lines using monoiodinated [Nle4,D-Phe7,Trp(2-nitro-4-azidophenylsulfenyl)9]-alpha-MSH as photoaffinity label revealed a major Mr 45,000 protein band on sodium dodecyl sulfate-polyacrylamide gels, analogous to the MSH receptor of mouse B16 melanoma cells.
Siegrist
W
W
Department of Research, University Hospital, Basel, Switzerland.
Solca
F
F
Stutz
S
S
Giuffrè
L
L
Carrel
S
S
Girard
J
J
Eberle
A N
AN
D000818:Animals; D002460:Cell Line; D004285:Dogs; D005910:Glioma / Q000378:metabolism; D006801:Humans; D007700:Kinetics; D009074:Melanocyte-Stimulating Hormones / Q000378:metabolism*; D008545:Melanoma / Q000378:metabolism*; D008546:Melanoma, Experimental / Q000378:metabolism; D051379:Mice; D009447:Neuroblastoma / Q000378:metabolism; D011979:Receptors, Pituitary Hormone / Q000378:metabolism*; D014407:Tumor Cells, Cultured / Q000378:metabolism*
D011979:Receptors, Pituitary Hormone; C029446:MSH receptor; D009074:Melanocyte-Stimulating Hormones