Bombesin-like peptide receptor subtypes promote mitogenesis, which requires persistent receptor signaling

Bombesin-like peptides (BLPs) can regulate the growth of normal and transformed cells. To compare the relative activities of the three known human BLP receptor subtypes [i.e., the gastrin-releasing peptide (GRP) receptor, neuromedin B (NMB) receptor, or BLP receptor subtype 3] in growth regulation, we expressed each receptor in a receptor-deficient host, Balb/3T3 cells. None of the receptor agonists used in our study promoted DNA synthesis by quiescent parental, nontransfected Balb/3T3 cells. Using clones stably transfected with the NMB receptor however, we found that NMB stimulated the incorporation of [3H]thymidine 2.5- to 8-fold over basal levels. The greatest net stimulation of [3H]thymidine incorporation occurred when the medium contained insulin. In quiescent Balb/3T3 cells transfected with the GRP receptor, GRP promoted a 15-fold increase in DNA synthesis in the absence of insulin or other growth factors. GRP also induced the labeling of a large percentage (53%) of the cells with bromodeoxyuridine. To determine the length of time that GRP receptor signaling was required to drive quiescent cells into the S phase of the cell cycle, we blocked GRP receptor signaling by addition of a competitive GRP receptor antagonist at different times after stimulating cells with GRP. Our data demonstrate that persistent GRP receptor signaling throughout a large part of the G1 phase of the cell cycle is important in the mitogenic effects of GRP in these cells. Hitherto uncharacterized GRP receptor signaling pathways may be important in this process. BLPs also stimulated a mitogenic response by transfectants expressing the BLP receptor subtype 3 if insulin was contained in the medium. Taken together, these studies indicate that all three BLP receptor subtypes may contribute to growth regulation in vivo.