Cell-specific signal transduction of parathyroid hormone (PTH)-related protein through stably expressed recombinant PTH/PTHrP receptors in vascular smooth muscle cells.
PTH-related protein activates a G protein-coupled PTH/PTHrP receptor in many cell types and produces diverse biological actions. To study the signal transduction events associated with biological activity of the PTH/PTHrP receptor in vascular smooth muscle, a principal PTHrP-responsive tissue, rat aortic smooth muscle cells (A10) were stably transfected with a plasmid encoding a PTH/PTHrP receptor and tested for ligand binding, PTHrP-(1-34)-induced cAMP levels, inositol phosphate production, and cytosolic calcium transients. Of nineteen G418-resistant lines recovered, all exhibited high affinity binding [approximately dissociation constant (Kd) > 10(-10)) of iodinated [Tyr36]hPTHrP(1-36)NH2 and ligand-induced cAMP accumulation (2- to 100-fold), which was directly proportional to PTH/PTHrP receptor number (range 4 x 10(3) to 7 x 10(7) sites/cell]. PTHrP had no effect on intracellular calcium or inositol phosphate formation in any cell line regardless of receptor number despite the presence of detectable G alpha q). Transient overexpression of individual G alpha q proteins (G alpha q, G alpha 11 or G alpha 14) into PTH/PTHrP receptor-expressing A10 cells conferred the ability of PTHrP to increase intracellular calcium and inositol phosphate formation. Ligand activation of the recombinant PTH/PTHrP receptor elicited appropriate downstream biological effects in A10 cells including inhibition of DNA synthesis and osteopontin messenger RNA (mRNA) expression. Thus, a single PTH/PTHrP receptor, though capable of coupling to different G proteins, signals exclusively through a cAMP-dependent pathway in vascular smooth muscle.
Maeda
S
S
Department of Medicine, University of Cincinnati College of Medicine, Ohio 45267, USA.
Wu
S
S
Jüppner
H
H
Green
J
J
Aragay
A M
AM
Fagin
J A
JA
Clemens
T L
TL
D000818:Animals; D002118:Calcium / Q000378:metabolism; D002455:Cell Division; D002460:Cell Line; D000242:Cyclic AMP / Q000378:metabolism; D007295:Inositol Phosphates / Q000096:biosynthesis; D007425:Intracellular Membranes / Q000378:metabolism; D009131:Muscle, Smooth, Vascular / Q000166:cytology / Q000502:physiology*; D053495:Osteopontin; D044162:Parathyroid Hormone-Related Protein; D011506:Proteins / Q000502:physiology*; D012333:RNA, Messenger / Q000378:metabolism; D011869:Radioligand Assay; D051381:Rats; D044168:Receptor, Parathyroid Hormone, Type 1; D018016:Receptors, Parathyroid Hormone / Q000502:physiology*; D011994:Recombinant Proteins; D012795:Sialoglycoproteins / Q000235:genetics; D015398:Signal Transduction; D014162:Transfection
D007295:Inositol Phosphates; D044162:Parathyroid Hormone-Related Protein; D011506:Proteins; D012333:RNA, Messenger; D044168:Receptor, Parathyroid Hormone, Type 1; D018016:Receptors, Parathyroid Hormone; D011994:Recombinant Proteins; D012795:Sialoglycoproteins; C506897:Spp1 protein, rat; D053495:Osteopontin; D000242:Cyclic AMP; D002118:Calcium