title: Phospholipase D- and protein kinase C isoenzyme-dependent signal transduction pathways activated by the calcitonin receptor.
abstract: The calcitonin receptor expressed by the porcine LLC-PK1 renal tubule cells is a seven-transmembrane domain, G protein-coupled receptor activating adenylyl-cyclase and phospholipase C. Salmon calcitonin stimulated dose- and time-dependent release of the phospholipase D-dependent phosphatidylcholine product [3H] choline with an EC50 = 2.5 +/-0.3 x 10(-8) M, similar to that determined for phosphoinositide metabolism (EC50 = 4.5 +/-1.0 x 10(-8)M). The hormone failed to induce release of [3H]phosphocholine and [3H]glycerophosphocholine, ruling out activation of phosphatydilcholine-specific phospholipase C and phospholipase A. Calcitonin stimulated phosphatidic acid, a product of phospholipase D-dependent phosphatydilcholine hydrolysis. Activation of phospholipase D was confirmed by release of [3H]phosphatydilethanol, a specific and stable product in the presence of a primary alcohol. Activation of calcitonin receptor induced diacylglycerol formation, with a rapid peak followed by a prolonged increase, due to activation of phospholipase C and of phospholipase D. Consequently, the protein kinase-C alpha, but not the delta isoenzyme, was cytosol-to-membrane translocated by approximately 50% after 20 min exposure to calcitonin, whereas protein kinase-C zeta, which was approximately 40% membrane-linked in unstimulated cells, translocated by approximately 19%. The human calcitonin receptor expressed by BIN-67 ovary tumor cells, although displaying higher affinity for calcitonin, failed to activate phospholipase D and protein kinase-C in response to the hormone. This receptor lacks the G protein binding consensus site due to the presence of a 48-bp cassette encoding for a 16-amino acid insert in the predicted first intracellular loop. This modification is likely to prevent the calcitonin receptor from associating to phospholipase-coupled signaling.
authors.affiliation: Department of Histology and Medical Embryology, University La Sapienza, Rome, Italy.
authors.forename: F
authors.initials: F
authors.lastname: Naro
authors.forename: M
authors.initials: M
authors.lastname: Perez
authors.forename: S
authors.initials: S
authors.lastname: Migliaccio
authors.forename: D L
authors.initials: DL
authors.lastname: Galson
authors.forename: P
authors.initials: P
authors.lastname: Orcel
authors.forename: A
authors.initials: A
authors.lastname: Teti
authors.forename: S R
authors.initials: SR
authors.lastname: Goldring
mesh_terms: D000818:Animals; D002116:Calcitonin / Q000494:pharmacology; D004075:Diglycerides / Q000096:biosynthesis; D004789:Enzyme Activation / Q000502:physiology; D006801:Humans; D007527:Isoenzymes / Q000502:physiology*; D018374:LLC-PK1 Cells; D010739:Phospholipase D / Q000378:metabolism / Q000502:physiology*; D010743:Phospholipids / Q000378:metabolism; D011493:Protein Kinase C / Q000502:physiology*; D018003:Receptors, Calcitonin / Q000502:physiology*; D012474:Salmon; D015398:Signal Transduction / Q000502:physiology*; D013552:Swine; D014407:Tumor Cells, Cultured
chemical_list: D004075:Diglycerides; D007527:Isoenzymes; D010743:Phospholipids; D018003:Receptors, Calcitonin; D002116:Calcitonin; D011493:Protein Kinase C; D010739:Phospholipase D