Rapamycin dissociates p70(S6K) activation from DNA synthesis stimulated by bombesin and insulin in Swiss 3T3 cells.
Phosphorylation and subsequent activation of p70 S6 kinase (S6K) are events that are highly conserved in the cellular response to mitogens. The neuropeptide bombesin, which is a potent mitogen for Swiss 3T3 cells, stimulated a time- and dose-dependent activation of p70(S6K) as determined by gel mobility shift and immune complex kinase assays. This effect was inhibited by the immunosuppressant rapamycin at 10 nM, which also completely abolished bombesin-stimulated DNA synthesis at identical concentrations. In striking contrast, the combination of bombesin and insulin in synergy stimulated maximum DNA synthesis in these cells despite persistent inhibition of p70(S6K) by rapamycin throughout G1. These results indicate that activation of p70(S6K) is not required for the transition of quiescent cells to the S phase of the cell cycle. The inhibitory effects of rapamycin on bombesin-stimulated cell cycle progression did not involve accumulation of the cyclin-dependent kinase inhibitor p27(kip1) but a striking inhibition of the expression of cyclin D1. This effect was circumvented by bombesin with insulin, suggesting that a rapamycin-insensitive pathway stimulated by this combination leads to cyclin D1 expression. Thus, these findings, dissociating the mitogenic effects of bombesin in synergy with insulin from activation of p70(S6K), support the hypothesis that this kinase is a component of one of the parallel pathways that can lead to DNA synthesis rather than an obligatory point of convergence in mitogenic signaling.
Withers
D J
DJ
Imperial Cancer Research Fund, P. O. Box 123, 44 Lincoln's Inn Fields, London, WC2A 3PX, United Kingdom.
Seufferlein
T
T
Mann
D
D
Garcia
B
B
Jones
N
N
Rozengurt
E
E
D016475:3T3 Cells; D000818:Animals; D001839:Bombesin / Q000494:pharmacology*; D018797:Cell Cycle Proteins; D019938:Cyclin D1; D050760:Cyclin-Dependent Kinase Inhibitor p27; D018844:Cyclin-Dependent Kinases / Q000037:antagonists & inhibitors; D016213:Cyclins / Q000378:metabolism; D004261:DNA Replication / Q000187:drug effects*; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000378:metabolism; D007166:Immunosuppressive Agents / Q000494:pharmacology*; D007328:Insulin / Q000494:pharmacology*; D051379:Mice; D008869:Microtubule-Associated Proteins / Q000378:metabolism; D015513:Oncogene Proteins / Q000378:metabolism; D010766:Phosphorylation; D011090:Polyenes / Q000494:pharmacology*; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D019893:Ribosomal Protein S6 Kinases; D020123:Sirolimus; D013936:Thymidine / Q000378:metabolism; D013997:Time Factors; D025521:Tumor Suppressor Proteins
C493961:Cdkn1b protein, mouse; D018797:Cell Cycle Proteins; D016213:Cyclins; D004791:Enzyme Inhibitors; D007166:Immunosuppressive Agents; D007328:Insulin; D008869:Microtubule-Associated Proteins; D015513:Oncogene Proteins; D011090:Polyenes; D025521:Tumor Suppressor Proteins; D019938:Cyclin D1; D050760:Cyclin-Dependent Kinase Inhibitor p27; D017346:Protein Serine-Threonine Kinases; D019893:Ribosomal Protein S6 Kinases; D018844:Cyclin-Dependent Kinases; D001839:Bombesin; D013936:Thymidine; D020123:Sirolimus