Parathyroid hormone and glucagon compete for binding to low affinity sites on human skin fibroblasts.
We have studied the binding sites for parathyroid hormone (PTH) present on normal human fibroblasts by studying these receptors with respect to (1) affinity and specificity towards various peptide hormones including human PTH(1-34) and porcine glucagon(1-29); (2) ability to mediate stimulation of cAMP production in response to these hormones; and (3) molecular size. Binding assays using 125I-labelled human PTH(1-34) and 125I-labelled porcine glucagon(1-29), and hormone stimulations of cAMP production were performed on confluent fibroblasts grown in 24-well dishes (passage 4-10). The molecular sizes of the binding sites for PTH and glucagon were assessed after cross-linking to the corresponding 125I-labelled ligand using the heterobifunctional reagent 1,4-difluoro-2,5-dinitrobenzene (DFDNB), by sodium dodecyl sulphate gel electrophoresis and autoradiography. The results demonstrate: (1) Biologically active PTH and glucagon, but not other peptide hormones tested, are equipotent competitors for binding on human fibroblasts to sites which have a relatively low affinity for these ligands (Kd approximately 0.8-2.4 x 10(-7) M); these sites have an apparent molecular weight of 95 kDa and are not linked to stimulation of cAMP production by PTH. (2) A distinct class of receptors for PTH with an apparent molecular weight of 60 kDa and which probably are linked to stimulation of cAMP production by PTH is also expressed by these cells; glucagon cannot compete with PTH for binding to these sites, and does not interfere with the stimulation of cAMP production by PTH. (3) Glucagon does not stimulate cAMP production in human fibroblasts.(ABSTRACT TRUNCATED AT 250 WORDS)
CNRS URA 583, Hôpital des Enfants Malades, Paris, France.
A
A
el Hessni
F
F
Authier
C
C
Silve
D001665:Binding Sites; D001667:Binding, Competitive; D000242:Cyclic AMP / Q000096:biosynthesis; D005347:Fibroblasts / Q000378:metabolism*; D005934:Glucagon / Q000378:metabolism*; D006801:Humans; D007700:Kinetics; D008970:Molecular Weight; D010281:Parathyroid Hormone / Q000378:metabolism*; D010446:Peptide Fragments / Q000378:metabolism*; D011485:Protein Binding; D011956:Receptors, Cell Surface / Q000378:metabolism*; D011964:Receptors, Gastrointestinal Hormone / Q000378:metabolism*; D018027:Receptors, Glucagon; D018016:Receptors, Parathyroid Hormone; D015290:Second Messenger Systems; D012867:Skin / Q000166:cytology / Q000378:metabolism*; D019379:Teriparatide
D010281:Parathyroid Hormone; D010446:Peptide Fragments; D011956:Receptors, Cell Surface; D011964:Receptors, Gastrointestinal Hormone; D018027:Receptors, Glucagon; D018016:Receptors, Parathyroid Hormone; D019379:Teriparatide; D005934:Glucagon; D000242:Cyclic AMP