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 125 I‐CCK‐8 with high affinity (Kd = 1.1 nM) and is functionally coupled to intracellular signaling pathways including rapid and transient increase in Ca 2+ 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. © 2001 Wiley‐Liss, Inc.