Photoaffinity labeling of parathyroid hormone receptors: comparison of receptors across species and target tissues and after desensitization to hormone.
Cells derived from human giant cell tumors of bone and fibroblasts derived from human neonatal foreskin respond to parathyroid hormone (PTH) by increasing the intracellular and extracellular levels of adenosine cyclic 3',5'-phosphate (cAMP). Using photoaffinity labeling methods, we examined these cells for the presence of a PTH receptor or a binding subunit of a receptor complex. A previously designed biologically active and photolabile radioligand analogue of PTH was reacted with these intact cells. After photolysis, the cells were extracted, and the proteins were denatured, reduced, and separated by electrophoresis on sodium dodecyl sulfate (Na-DodSO4)-polyacrylamide gels followed by autoradiography. A single membrane component, Mr 70 000, was labeled specifically in intact cells cultured from skeletal and dermal tissue. By mixing, in pairs, photolabeled proteins from (a) intact human cells derived from giant cell tumors of bone, (b) intact human fibroblasts, and (c) canine renal cortical membranes, the receptors (or their binding subunits) for PTH were compared directly and found to be identical in terms of molecular size (as determined by the migration position on NaDod-SO4-polyacrylamide gels) across species (dog and human) and target tissue (bone, skin, and kidney). Preincubation of cells cultured from giant cell tumors of bone with PTH resulted in loss of the PTH-induced cAMP response (desensitization). Preincubation with PTH was accompanied by a marked decrease in photoaffinity labeling of the PTH binding component and suggests that the loss of hormone response in cells preincubated with PTH was related to a decrease in the number or availability of PTH receptors.(ABSTRACT TRUNCATED AT 250 WORDS)
D000345:Affinity Labels / Q000494:pharmacology*; D000818:Animals; D001859:Bone Neoplasms / Q000378:metabolism*; D002277:Carcinoma / Q000378:metabolism*; D002462:Cell Membrane / Q000378:metabolism; D002478:Cells, Cultured; D000242:Cyclic AMP / Q000378:metabolism; D004285:Dogs; D006801:Humans; D007231:Infant, Newborn; D007672:Kidney Cortex / Q000378:metabolism*; D007700:Kinetics; D008297:Male; D010281:Parathyroid Hormone / Q000378:metabolism*; D011956:Receptors, Cell Surface / Q000378:metabolism*; D018016:Receptors, Parathyroid Hormone; D012867:Skin / Q000378:metabolism*; D013045:Species Specificity; D013329:Structure-Activity Relationship
D000345:Affinity Labels; D010281:Parathyroid Hormone; D011956:Receptors, Cell Surface; D018016:Receptors, Parathyroid Hormone; D000242:Cyclic AMP