CCK(B)/gastrin receptor mediates synergistic stimulation of DNA synthesis and cyclin D1, D3, and E expression in Swiss 3T3 cells.
In order to develop a model system for identifying signaling pathways and cell cycle events involved in gastrin-mediated mitogenesis, we have used high efficiency retroviral-mediated transfection of cholecystokinin (CCK)(B)/gastrin receptor into Swiss 3T3 cells. The retrovirally-transfected CCK(B)/gastrin receptor binds 125I-CCK-8 with high affinity (Kd = 1.1 nM) and is functionally coupled to intracellular signaling pathways including rapid and transient increase in Ca2+ fluxes, protein kinase C-dependent protein kinase D activation, and MEK-dependent ERK1/2 activation. In the presence of insulin, CCK-8 or gastrin induced a 66.5 +/- 8.8-fold (mean +/- SEM, n = 24 in eight independent experiments) increase in cellular DNA synthesis, reaching a level similar to that achieved by stimulation with a saturating concentration of fresh serum, and much greater than the response to each agonist added alone. CCK-8 also induced a striking increase in the expression of cyclins D1, D3, and E and hyperphosphorylation of Rb acting synergistically with insulin. Similar effects were observed when CCK(B)/gastrin receptor was activated in the presence of EGF or bombesin. Our results demonstrate that activation of CCK(B)/gastrin receptor retrovirally-transfected into Swiss 3T3 induces a potent synergistic effect on DNA synthesis, accumulation of cyclins D1, D3, and E and hyperphosphorylation of Rb in combination with insulin, EGF, or bombesin. Thus, the CCK(B)/gastrin receptor transfected into Swiss 3T3 cells provides a novel model system to elucidate mitogenic signal transduction pathways and cell cycle events activated via this receptor.
Zhukova
E
E
Department of Medicine, School of Medicine and Molecular Biology Institute, University of California, Los Angeles, CA 90095-1786, USA.
Sinnett-Smith
J
J
Wong
H
H
Chiu
T
T
Rozengurt
E
E
D016475:3T3 Cells; D000818:Animals; D002118:Calcium / Q000378:metabolism; D002453:Cell Cycle; D019938:Cyclin D1 / Q000096:biosynthesis*; D056743:Cyclin D3; D019927:Cyclin E / Q000096:biosynthesis*; D016213:Cyclins / Q000096:biosynthesis*; D004247:DNA / Q000096:biosynthesis; D004357:Drug Synergism; D005822:Genetic Vectors; D007328:Insulin / Q000494:pharmacology; D051379:Mice; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D011493:Protein Kinase C / Q000378:metabolism; D043887:Receptor, Cholecystokinin B; D011949:Receptors, Cholecystokinin / Q000235:genetics / Q000502:physiology*; D016160:Retinoblastoma Protein / Q000378:metabolism; D012190:Retroviridae / Q000235:genetics; D012844:Sincalide / Q000378:metabolism / Q000494:pharmacology; D014162:Transfection
C534697:Ccnd3 protein, mouse; D056743:Cyclin D3; D019927:Cyclin E; D016213:Cyclins; D007328:Insulin; D043887:Receptor, Cholecystokinin B; D011949:Receptors, Cholecystokinin; D016160:Retinoblastoma Protein; D019938:Cyclin D1; D004247:DNA; C089212:protein kinase D; D011493:Protein Kinase C; D020928:Mitogen-Activated Protein Kinases; D012844:Sincalide; D002118:Calcium