Abstract
In order to investigate the regulatory mechanisms involved in the expression of fos and jun family members by glucocorticoids, and the effect of ras transformation in intestinal epithelial cells, we used the rat cell line IEC-6. Dexamethasone treatment induced transiently c-jun mRNAs, in contrast to the sustained expression of c-fos, whereas its effect on junB expression resulted in a later increase. Dexamethasone-dependent stimulation of c-fos and c-jun was modulated predominantly at the level of transcription. Sustained levels of induced c-fos and c-jun proteins were observed after dexamethasone treatment. AP-1 DNA-binding capacity of c-fos, and to a smaller extent c-jun, was increased by glucocorticoids later than after serum treatment. To analyse the effect of ras on the glucocorticoid response of AP-1 components, we studied several IEC-6 cell clones transformed by the Ha-ras oncogene. In comparison to normal cells, these transformants displayed increased AP- 1 DNA-binding activity with higher levels of junB and variable levels of c-jun in the AP-1 complex. Ras transformation repressed the growth-inhibitory properties of glucocorticoids. Furthermore, ras inhibited the glucocorticoid-dependent induction of c-fos protein and mRNA, leading to changes in AP-1 composition as compared to normal cells. As assessed by transient transfection luciferase assays, glucocorticoids induced significantly a minimal promoter containing 3 copies of an AP-1 DNA-binding site as well as the murine c-fos -276 to +112 promoter in non-transformed cell lines. In contrast, glucocorticoid addition did not induce these constructs in two ras transformed cells. These results suggest that ras negatively modulates specific responses of intestinal epithelial cells to glucocorticoids.
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References
Kinzler KW, Vogelstein B: Lessons from hereditary colon cancer. Cell 87: 159–170, 1996
Van Aelst L, Barr M, Marcus S, Polverino A, Wigler M: Complex formation between ras and raf and other protein kinases. Proc Natl Acad Sci USA 90: 6213–6217, 1993
Vojtek AB, Hollenberg SM, Cooper JA: Mammalian ras interacts directly with the serine/threonine kinase Raf. Cell 74: 205–214, 1993
Angel P, Karin M: The role of Jun, Fos and the AP-1 complex in cell proliferation and transformation. Biochim Biophys Acta 1072: 129–157, 1991
Karin M: Signal transduction from the cell surface to the nucleus through the phosphorylation of transcription factors. Curr Opin Cell Biol 6: 415–424, 1994
Karin M: The regulation of AP-1 activity by mitogen-activated protein kinases. J Biol Chem 270: 16483–16486, 1995
Kovary K, Bravo R: Expression of different Jun and Fos proteins during the G0-to-G1 transition in mouse fibroblasts: in vitro and in vivo associations. Mol Cell Biol 11: 2451–2459, 1991
Henning SJ, Rubin DC, Shulnan RJ: Ontogeny of the intestinal mucosa. In: L.R. Johnson (ed). Physiology of the Gastrointestinal Tract, Raven Press, New York, 1994, pp 571–615
Doolan CM, Harvey BJ: Modulation of cytosolic protein kinase C and calcium ion activity by steroid hormones in rat distal colon. J Biol Chem 271: 8763–8767, 1996
Schüle R, Evans RM: Cross-coupling of signal transduction pathways: zinc finger meets leucine zipper. Trends Genet 7: 377–381, 1991
Cato ACB, Wade E: Molecular mechanisms of anti-inflammatory action of glucocorticoids. BioEssays 18: 371–378, 1996
Miner JN, Yamamoto KR: The basic region of AP-1 specifies glucocorticoid receptor activity at a composite response element. Genes Dev 6: 2491–2501, 1992
Blais S, Boudreau F, Thorneloe K, Asselin C: Differential expression of fos and jun family members during murine postnatal intestinal development. Biol Neonate 69: 342–349, 1996
Quaroni A, May RJ: Establishment and characterization of intestinal epithelial cell cultures. Meth Cell Biol 21B: 403–427, 1980
Podolsky DK: Regulation of intestinal epithelial proliferation: A few answers, many questions. Am J Physiol 264: G179–G186, 1993
Chomczynski P, Sacchi N: Single step method of RNA isolation by acid guanidinium thiocyanate-phenol-chloroform extraction. Anal Biochem 162: 156–159, 1987
Sambrook J, Fritsch EF, Maniatis T: Molecular Cloning: A Laboratory Manual. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, 1989
Lemischka IR, Farmer S, Racaniello VR, Sharp PA: Nucleotide sequence and evolution of a mammalian α-tubulin mesenger RNA. J Mol Biol 15: 101–120, 1981.
Van Beveren C, Van Straaten F, Curran T, Müller R, Verma IM: Analysis of FBJ-MuSV provirus and c-fos (mouse) gene reveals that viral and cellular fos gene products have different carboxy termini. Cell 32: 1241–1255, 1983
Ryder K, Nathans D: Induction of protooncogene c-jun by serum growth factors. Proc Natl Acad Sci USA 85: 8464–8467, 1988
Ryder K, Lau LF, Nathans D: A gene activated by growth factors is related to the oncogene v-jun. Proc Natl Acad Sci USA 85: 1487–1491, 1988
Ryder K, Lanahan A, Perez-Albuerne E, Nathans D: Jun-D: A third member of the Jun gene family. Proc Natl Acad Sci USA 86: 1500–1503, 1989
Souleimani A, Asselin C: Regulation of c-fos expression by sodium butyrate in the human colon carcinoma cell line Caco-2. Biochem Biophys Res Commun 193: 330–336, 1993
Boudreau F, Blais S, Asselin C: Regulation of CCAAT/enhancer binding protein isoform by serum and glucocorticoids in the rat intestinal epithelial crypt cell line IEC-6. Exp Cell Res 222: 1–9, 1996.
Stein B, Baldwin AS, Ballard DW, Greene WC, Angel P, Herrlich P: Cross-coupling of the NF-κB p65 and Fos/Jun transcription factors produces potentiated biological function. EMBO J 12: 3879–3891, 1993
Asselin C, Nepveu A, Marcu KB: Molecular requirements for transcriptional initiation of the murine c-myc gene. Oncogene 4: 549–558, 1989
Lee W, Mitchell P, Tjian R: Purified transcription factor AP-1 interacts with TPA-inducible enhancer elements. Cell 49: 741–752, 1987
Ron D, Brasier AR, McGehee RE, Habener J-F: Tumor necrosis factorinduced reversal of adipocyte phenotype of 3T3-Ll cells is preceded by a loss of nuclear CCAAT/enhancer binding protein (C/EBP). J Clin Invest 89: 223–233, 1992
Spandidos DA, Wilkie NM: Malignant transformation of early passage rodent cells by single mutated human oncogene. Nature 310: 469–475, 1984
Wharton W, Smyth MJ: In: R. Baserga (ed.). Cell Growth and Division. IRL Press, Oxford, 1993, pp. 139–153
Candeliere GA, Jurutka PW, Haussler MR, St-Arnaud R: A composite element binding the vitamin D receptor, retinoid X receptor α, and a member of the CTF/NF-1 family of transcription factors mediate the vitamin D responsiveness of the c-fos promoter. Mol Cell Biol 16: 584–592, 1996
Hollenberg AN, Monden T, Flynn TR, Boers M-E, Cohen O, Wondisford FE: The human thyrotropin-releasing hormone gene is regulated by thyroid hormone through two distinct classes of negative thyroid hormone response elements. Mol Endocrinol 9: 540–550, 1995
Curran T, Franza R: Fos and jun: The AP-1 connection. Cell 55: 395–397, 1988
Landers JP, Spelsberg TC: New concepts in steroid hormone action: Transcription factors, proto-oncogenes, and the cascade model for steroid regulation of gene expression. Crit Rev Euk Gene Express 2: 19–63, 1992
Wang WW, Howells RD: Sequence of the 5′-flanking region of the rat c-fos proto-oncogene. Gene 143: 261–264, 1994
Lin S-C, Macleod S, Hardin JW: Effects of glucocorticoids on expression of the fos protooncogene in AtT-20 cells. Endocrinology 130: 257–262, 1992
Subramian M, Colvard D, Keeting PE, Rasmussen K, Riggs L, Spelsberg TC: Glucocorticoid regulation of alkaline phosphatase, osteocalcin, and proto-oncogenes in normal human osteoblast-like cells. J Cell Biochem 50: 411–424, 1992
Grassilli E, Carcereri de Prati A, Monti D, Troiano L, Menegazzi M, Barbieri D, Francheschi C, Suzuki H: Studies of the relationship between cell proliferation and cell death. II. Early gene expression during concanavalin A-induced proliferation or dexamethasoneinduced apoptosis of rat thymocytes. Biochem Biophys Res Commun 188: 1261–1266, 1992
Karagianni N, Tsawdaroglou N: The c-fos serum response element (SRE) confers negative response to glucocorticoids. Oncogene 9: 2327–2334, 1994
Binetruy B, Smeal T, Karin M: H-ras augments c-Jun activity and stimulates phosphorylation of its activation domain. Nature 351: 122–127, 1991
Coffer P, De Jonge M, Mettouchi A, Binetruy B, Ghysdael J, Kruijer W: junB promoter regulation: Ras mediated transactivation by c-Ets-1 and c-Ets-2. Oncogene 9: 911–921, 1994
Mechta F, Lallemand D, Pfarr CM, Yaniv M: Transformation by ras modifies AP1 composition and activity. Oncogene 14: 837–847, 1997
Yu CL, Prochownik EV, Imperiale MJ, Jove R: Attenuation of serum inducibility of immediate early genes by oncoproteins in tyrosine kinase signaling pathways. Mol Cell Biol 13: 2011–2019, 1993
Chiu R, Angel P, Karin M: Jun-B differs in its biological properties from, and is a negative regulator of, c-jun. Cell 59: 979–986, 1989
Schütte J, Viallet J, Nau M, Segal S, Fedorko J, Minna J: jun-B inhibits and c-fos stimulates the transforming and trans-activating activities of c-jun. Cell 59: 987–997, 1989
McEwan IJ, Wright APH, Gustafson J-A: Mechanism of gene expression by the glucocorticoid receptor: Role of protein-protein interactions. BioEssays 19: 153–160, 1997
Lowy DR, Willumsen BM: Function and regulation of Ras. Annu Rev Biochem 62: 851–891, 1993
Rake JB, Quinones MA, Faller DV: Inhibition of platelet-derived growth factor-mediated signal transduction by transforming ras. Suppression of receptor autophosphorylation. J Biol Chem 266: 5348–5352, 1991
Chen C-Y, Forman LW, Faller DV: Calcium-dependent immediate-early gene induction in lymphocytes is negatively regulated by p21Ha-ras. Mol Cell Biol 16: 6582–6592, 1996
Hu J, Isom HC: Suppression of albumin enhancer activity by Ha-ras and AP-1 in hepatocyte cell lines. Mol Cell Biol 14: 1531–1543, 1994
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Boudreau, F., Zannoni, S., Pelletier, N. et al. Negative regulation of glucocorticoid-dependent induction of c-fos by ras in intestinal epithelial cells. Mol Cell Biochem 195, 99–111 (1999). https://doi.org/10.1023/A:1006987313013
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DOI: https://doi.org/10.1023/A:1006987313013