Effect of Insulin on Cell Cycle Progression in MCF-7 Breast Cancer Cells

We recently demonstrated that in MCF-7 breast cancer cells, insulin promoted the phosphorylation and activation of geranylgeranyltransferase I (GGTI-I), increased the amounts of geranylgeranylated Rho-A and potentiated the transactivating activity of lysophosphatidic acid (LPA) (Chappell, J., Golovchenko, I., Wall, K., Stjernholm, R., Leitner, J., Goalstone, M., and Draznin, B. (2000)J. Biol. Chem. 275, 31792–31797). In the present study, we explored the mechanism of this potentiating effect of insulin on LPA. Insulin (10 nm) potentiated the ability of LPA to stimulate cell cycle progression and DNA synthesis in MCF-7 cells. The potentiating effect of insulin appears to involve increases in the expression of cyclin E and decreases in the expression of the cyclin-dependent kinase inhibitor p27Kip1. All potentiating effects of insulin were inhibited in the presence of an inhibitor of GGTase I, GGTI-286 (3 μm) or by an expression of a dominant negative mutant of Rho-A. In contrast to its potentiating action, a direct mitogenic effect of insulin in MCF-7 cells involves activation of phosphatidylinositol 3-kinase and increased expression of cyclin D1. We conclude that the ability of insulin to increase the cellular amounts of geranylgeranylated Rho-A results in potentiation of the LPA effect on cyclin E expression and degradation of p27Kip1 and cell cycle progression in MCF-7 breast cancer cells. We recently demonstrated that in MCF-7 breast cancer cells, insulin promoted the phosphorylation and activation of geranylgeranyltransferase I (GGTI-I), increased the amounts of geranylgeranylated Rho-A and potentiated the transactivating activity of lysophosphatidic acid (LPA) (Chappell, J., Golovchenko, I., Wall, K., Stjernholm, R., Leitner, J., Goalstone, M., and Draznin, B. (2000)J. Biol. Chem. 275, 31792–31797). In the present study, we explored the mechanism of this potentiating effect of insulin on LPA. Insulin (10 nm) potentiated the ability of LPA to stimulate cell cycle progression and DNA synthesis in MCF-7 cells. The potentiating effect of insulin appears to involve increases in the expression of cyclin E and decreases in the expression of the cyclin-dependent kinase inhibitor p27Kip1. All potentiating effects of insulin were inhibited in the presence of an inhibitor of GGTase I, GGTI-286 (3 μm) or by an expression of a dominant negative mutant of Rho-A. In contrast to its potentiating action, a direct mitogenic effect of insulin in MCF-7 cells involves activation of phosphatidylinositol 3-kinase and increased expression of cyclin D1. We conclude that the ability of insulin to increase the cellular amounts of geranylgeranylated Rho-A results in potentiation of the LPA effect on cyclin E expression and degradation of p27Kip1 and cell cycle progression in MCF-7 breast cancer cells. geranylgeranyltransferase lysophosphatidic acid retinoblastoma phosphatidylinositol 3-kinase mitogen-activated protein kinase/extracellular signal-regulated kinase kinase mitogen-activated protein extracellular signal-regulated kinase cyclin-dependent kinase Both hyperinsulinemia and cancer are extremely prevalent pathophysiological conditions associated with major morbidity and mortality. The implication of insulin in the pathogenesis of cancer has been in the English medical literature since the early 1970's (1Kessler I.I. J. Chronic Dis. 1971; 23: 579-600Abstract Full Text PDF PubMed Scopus (66) Google Scholar). 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Chem. 2000; 275: 31792-31797Abstract Full Text Full Text PDF PubMed Scopus (34) Google Scholar). in the GGTase I activity in of the amounts of geranylgeranylated Rho-A in the cells and potentiation of the effects acid In the present we the mechanism of the potentiating effect of insulin on cell cycle progression and DNA synthesis in MCF-7 breast cancer cells. culture and were Insulin E and and geranylgeranyltransferase were and protein and were acid and were MCF-7 breast cancer cells were a L. of and a dominant negative mutant of Rho-A a J. J. and MCF-7 cells were to in and insulin The cells were and and with insulin (10 nm) with and or were an with LPA were the in and with the of A. J. Biol. PubMed Scopus Google Scholar). cycle a of the DNA to the were cell cycle MCF-7 cells were to in as The cells were and and with insulin (10 nm) with and were an with LPA μm) in the presence of of in and by cell and by MCF-7 cells were to as The cells were and and with insulin (10 nm) with and were an with LPA were with and a and and protein were to E and cyclin protein were the were by by and by MCF-7 cells were to as The cells were and and with insulin (10 nm) with and were an with LPA were with and as and and protein were to p27Kip1 and protein were the were by by and by the of the of the and were by or with recently J. M. D. J. Biol. Chem. 1998; 273: Full Text Full Text PDF PubMed Scopus Google Scholar) indicated that hyperinsulinemia the effects of LPA by the of geranylgeranylated the were to the mechanism of the potentiating of insulin on of MCF-7 cells. In the we the direct effects of insulin on MCF-7 breast cancer cell by activated cell cycle Insulin (10 nm) cell by in The effect of insulin with insulin by of an inhibitor of phosphatidylinositol 3-kinase the mitogenic effect of the inhibitor μm) effect on these that the direct mitogenic of insulin in MCF-7 cells is the 3-kinase of insulin's B. J. H. F. B. J. J. Biol. Chem. 1997; Full Text Full Text PDF PubMed Scopus Google Scholar, J. M. D. J. Biol. Chem. 1998; 273: Full Text Full Text PDF PubMed Scopus Google Scholar). the effect of insulin on GGTase I and of Rho-A is by the kinase and is of 3-kinase M.L. Leitner J.W. J.M. Draznin B. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus (37) Google Scholar), these cells as an to the mechanism of the potentiating of insulin of its direct mitogenic action, which is We the potentiating effect of insulin on the mitogenic of LPA. Insulin (10 nm) the increase in LPA μm) MCF-7 cell LPA to MCF-7 cells, which were with insulin to potentiating effect of insulin by the of the geranylgeranyltransferase inhibitor GGTI-286 (3 GGTI-286 effect on insulin of the effect of insulin the ability of insulin to LPA to with LPA that the potentiating effect of insulin is on of to and the potentiating effect of DNA is of DNA synthesis and cell we the ability of insulin to the effect of LPA on in MCF-7 cells. Insulin increased DNA LPA increased by LPA to cells, which were with DNA to insulin the effect of LPA by the of of of Rho-A the potentiating effect of of GGTI-286 to the the potentiating effect of the of as with the cells with insulin and LPA. in cells with insulin in with GGTI-286 that in with insulin the role of Rho-A in the potentiating effect of cells were with a dominant negative mutant of Rho-A to the insulin and LPA of a dominant negative Rho-A the ability of insulin to the effect of LPA on effect on insulin and of we direct cell of MCF-7 cells to insulin and LPA We a effect of insulin and LPA on cell that in the presence of a GGTase I inhibitor or a dominant negative mutant of Rho-A. these that the potentiating effect of insulin is to the ability of insulin to GGTase I, its direct effect is of its on an of a dominant negative Rho-A the proteins, we the effect of insulin on the phosphorylation of and in and MCF-7 cells. of a dominant negative Rho-A effect on the ability of insulin to stimulate the phosphorylation of and normal of in these cells. insulin increased the progression of MCF-7 cells cell cycle and potentiated the mitogenic effect of we the effect of insulin on cell cycle by its effect on cyclin and cyclin E The of these to the in the of the cell cycle that is potentiated by Insulin (10 nm) cyclin LPA μm) stimulate cyclin However, insulin the effect of LPA on cyclin D1. In cells with insulin and to cyclin that in the cells with LPA In contrast to the with cyclin insulin (10 nm) effect on cyclin E LPA μm) cyclin E However, of MCF-7 cells with insulin increased the cyclin E to than as with potentiating effect of insulin with the of GGTI-286 or by a of a dominant negative mutant of Rho-A that the potentiating of insulin is by its effect on GGTase I. the of cyclin E is by its with the cyclin-dependent kinase inhibitor in D. A. Mol. Biol. 2000; Google and C. J. 1999; PubMed Scopus Google Scholar) and Rho-A p27Kip1 degradation its of cyclin activity W. C. J. J. Biol. Chem. 1999; 274: Full Text Full Text PDF PubMed Scopus Google Scholar), we insulin's effect on in MCF-7 cells. insulin (10 nm) or LPA μm) a and effect on p27Kip1 However, LPA μm) to MCF-7 cells with insulin p27Kip1 by The presence of GGTI-286 or of a dominant negative Rho-A a in p27Kip1 that by insulin in with LPA of the protein the phosphorylation of and in the protein by cyclin in J. J. J. 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Draznin B. 1998; PubMed Google Scholar). we that insulin cell cycle progression and in MCF-7 breast cancer cells and that the mechanism of the cell cycle progression is the of cyclin E and of the inhibitor p27Kip1. the presence of GGTI-286 geranylgeranyltransferase or the expression of a dominant negative Rho-A the potentiating effect of insulin on of cell cycle we that insulin LPA its effect on Rho-A prenylation and of increased amounts of geranylgeranylated Rho-A. We have that the effects of GGTI-286 on insulin's ability to effects of LPA are and J. Golovchenko I. Wall K. Stjernholm R. Leitner J. Goalstone M. Draznin B. J. Biol. 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Epidemiol. 1990; PubMed Scopus Google Scholar). aspect of insulin may explain women with insulin receptor-positive breast cancer have been to have a worse prognosis than women with insulin receptor-negative tumors (21Belfiore A. Frittitta L. Costantino A. Frasca F. Pandini G. Sciacca L. Goldfine I.D. Vigneri R. Ann. N. Y. Acad. Sci. 1996; 784: 173-188Crossref PubMed Scopus (58) Google Scholar), and fasting insulin levels may predict survival and overall survival of women with breast cancer who are receiving adjuvant (22Goodwin P.J. Ennis M. Pritchard K.I. Trudeau M.E. Koo J. Madaranas Y. Hood N. Hartwick W. Hoffman B. American Society of Clinical Oncology Online. 2000; (asco.org)Google Scholar). We and the of Cancer The is by of