Binding and degradation of NH2-terminal parathyroid hormone by opossum kidney cells.
The binding and cellular processing of NH2-terminal parathyroid (PTH) hormone by confluent monolayers of opossum kidney (OK) cells was characterized using radiolabeled PTH peptide analogues. Time- and temperature-dependent specific binding of 125I-labeled (Nle-8,18, Tyr-34)-NH2-bovine(b)PTH-(1-34) was accompanied by the appearance of degraded radiolabel in the cell medium. Degrading activity was observed to be a specific consequence of binding by PTH receptors. Degrading activity was inhibited by monensin, chloroquine, and NH4+ but not by chymotrypsin inhibitors. Acid washing demonstrated that greater than 80% of total cell-associated specific binding at equilibrium was located in a rapidly internalized (acid-resistant) pool. Monensin pretreatment led to increased acid-resistant binding, presumably through inhibition of turnover of internalized receptor ligand and indicated that the degradation of radiolabel was probably associated with processing of the receptor-ligand complex. Release of intact radiolabel from the acid-resistant pool indicated that some of the internalized peptide was recycled out of the cell in an undegraded form (retroendocytosis). Acid-resistant binding and degradation of 125I-(Nle-8,18, Tyr-34)-NH2-bPTH-(3-34) was minimal, indicating that this ligand was not internalized. It is concluded that the binding and internalization of PTH-(1-34) fragment by confluent OK cells is a specific receptor-mediated process. Cellular processing of PTH-(1-34) conforms to established models of internalization by receptor-mediated endocytosis.
Brown
R C
RC
Department of Medical Biochemistry, University of Wales College of Medicine, Cardiff, United Kingdom.
Silver
A C
AC
Woodhead
J S
JS
D000818:Animals; D001667:Binding, Competitive; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D005576:Colforsin / Q000494:pharmacology; D004705:Endocytosis; D007457:Iodine Radioisotopes; D007668:Kidney; D007700:Kinetics; D009893:Opossums; D010281:Parathyroid Hormone / Q000378:metabolism*; D010446:Peptide Fragments / Q000378:metabolism*; D011956:Receptors, Cell Surface / Q000187:drug effects / Q000378:metabolism*; D018016:Receptors, Parathyroid Hormone; D013816:Thermodynamics
D007457:Iodine Radioisotopes; D010281:Parathyroid Hormone; D010446:Peptide Fragments; D011956:Receptors, Cell Surface; D018016:Receptors, Parathyroid Hormone; D005576:Colforsin; C032505:parathyroid hormone (1-34)amide, 8,18-Nle-34-Tyr-, bovine