Parathyroid hormone stimulates translocation of protein kinase C isozymes in UMR-106 osteoblastic osteosarcoma cells.
Studies with antagonists have provided evidence that protein kinase C (PKC) is involved in several of the actions of parathyroid hormone (PTH) on bone. PTH increases total PKC activity in bone and bone cells. The current studies investigated whether PTH can activate specific PKC isozymes, as demonstrated by translocation of these isozymes from cytosolic to membrane fractions. The isozymes selected for study, alpha, betaI, delta, epsilon, and zeta, were shown previously by us to be present in normal osteoblasts and several osteosarcoma-derived osteoblastic cells. UMR-106 cells, a widely used osteoblastic cell line, were selected for the current study. PKC isozymes in whole cell lysates and cell fractions were visualized by western blotting; isozyme distribution was also visualized by immunofluorescence. The total amounts of the isozymes and their relative distribution between membrane and cytosolic fractions in untreated cells were stable over a range of passages (5-20 from initial plating). In untreated cells, the concentrations of PKC alpha, betaI, and zeta were higher in the cytosol, and PKC delta and epsilon were higher in the membrane fraction. Treatment with 1 or 10 nmol/L PTH for 1 or 5 min stimulated translocation of PKC alpha and betaI, with variable effects on the other isozymes. Treatment with phorbol-12,13-dibutyrate (PDBu), 1 micromol/L for 5 min, elicited similar effects to those of PTH on PKC alpha and betaI. Treatment with PDBu for 48 h resulted in a downregulation of PKC alpha, whereas a 48 h treatment with PTH did not cause downregulation. The results indicate that PTH can affect specific PKC isozymes, providing a mechanism for differential regulation of cellular actions through this pathway.
Dossing
D A
DA
Department of Molecular Pharmacology, Northwestern University Medical School, Chicago, IL 60611, USA.
Radeff
J M
JM
Sanders
J
J
Lee
S K
SK
Hsieh
M R
MR
Stern
P H
PH
D000818:Animals; D001859:Bone Neoplasms; D002273:Carcinogens / Q000494:pharmacology; D002462:Cell Membrane / Q000201:enzymology; D003600:Cytosol / Q000201:enzymology; D007527:Isoenzymes / Q000378:metabolism*; D010006:Osteoblasts / Q000166:cytology / Q000187:drug effects / Q000201:enzymology*; D012516:Osteosarcoma; D010281:Parathyroid Hormone / Q000494:pharmacology; D015240:Phorbol 12,13-Dibutyrate / Q000494:pharmacology; D011493:Protein Kinase C / Q000378:metabolism*; D064546:Protein Kinase C beta; D051571:Protein Kinase C-alpha; D051745:Protein Kinase C-delta; D051744:Protein Kinase C-epsilon; D051381:Rats; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology; D014407:Tumor Cells, Cultured; D000099221:Protein Kinase C zeta
D002273:Carcinogens; D007527:Isoenzymes; D010281:Parathyroid Hormone; D015240:Phorbol 12,13-Dibutyrate; D011493:Protein Kinase C; D064546:Protein Kinase C beta; D051571:Protein Kinase C-alpha; D051745:Protein Kinase C-delta; D051744:Protein Kinase C-epsilon; D000099221:Protein Kinase C zeta; C497260:Prkcd protein, rat; C497257:Prkce protein, rat