Abstract
3T3-L1 predipocytes differentiate into cells having the biochemical properties of adipocytes; tumor necrosis factor-α (TNF), retinoic acid (RA), and transforming growth factor-β (TGF-β), attenuate this process. Inhibition of differentiation by these agents, thought to be at the level of transcription, has been investigated by examining the accumulation of mRNA for six transcription factors and the autocrine growth factor interleukin 6 (IL-6). Upon induction of differentiation, a rapid and major accumulation of c-fos and jun-B mRNA was observed that returned to near basal levels within 4–6 h. In contrast, c-jun mRNA, although rapidly expressed following induction of differentiation, remained at relatively constant levels throughout the time course. Exposure of the cells to 5 nM TNF potentiated the accumulation of all 3 mRNAs but most significantly c-jun (12-fold), which remained elevated for at least 24 h after treatment. In control differentiating cells, krox-20 and fox-B were expressed transiently, 30 min to 2 h, while fra-1 mRNA accumulated over an extended period, 1 to 8 h. Again, TNF enhanced the accumulation of these mRNAs. Accumulation of mRNA for C/EBP, a transcription factor proposed to control expression of genes involved in the terminally differentiated state was attenuated after exposure of the cells to TNF. C/EBP expression was also inhibited in cells exposed to RA or TGF-β. IL-6 mRNA was expressed briefly (30 min to 2 h) and again transiently (at 8 h after induction of differentiation). TNF treatment markedly enhanced accumulation of IL-6 message. We propose that increased cellular content of one or more transcription factors or the suppression of C/EBP may be responsible for the attenuation of differentiation induced by exposure of the cells to TNF, RA, and TGF-β.
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References
Green H, Kehinde O: Sublines of mouse 3T3 cells that accumulate lipid. Cell 1: 113–116, 1974
Green H, Kehinde O: Spontaneous heritable changes leading to increased adipose conversion in 3T3 cells. Cell 7: 105–113, 1976
Green H, Kehinde O: Formation of normally differentiated subcutaneous fat pads by an established preadipose cell line. J Cell Physiol 101: 169–172, 1979
Rubin CS, Lai E, Rosen OM: Acquisition of increased hormone sensitivity duringin vitro adipocyte development. J Biol Chem 252: 3554–3557, 1977
Karlsson FA, Grunfeld C, Kahn CR, Roth J: Regulation of insulin receptor and insulin responsiveness in 3T3-L1 fatty fibroblasts. Endocrinology 104: 1383–1392, 1979
Reed B, Lane MD: Insulin receptor synthesis and turnover in differentiating 3T3-L1 preadipocytes. Proc Natl Acad Sci USA 77: 285–289, 1980
Mackall JC, Lane MD: Role of pyruvate carboxylase in fatty acid synthesis: alterations during preadipocyte differentiation. Biochem Biophys Res Commun 79: 720–725, 1977
Wise LS, Green H: Studies of lipoprotein lipase during the adipose conversion of 3T3 cells. Cell 13: 233–242, 1978
Spooner PM, Chernick SS, Garrison MM, Scow RO: Insulin regulation of lipoprotein lipase activity and release in 3T3-L1 adipocytes. J Biol Chem 254: 10021–10029, 1979
Spiegelman BM, Farmer SR: Decreases in tubulin and actin gene expression prior to morphological differentiation of 3T3 adipocytes. Cell 29: 53–60, 1982
Bernlohr DA, Bolanowski MA, Kelly TJ, Lane MD: Evidence for an increase in transcription of specific mRNAs during differentiation of 3T3-L1 preadipocytes. J Biol Chem 260: 5563–5567, 1985
Cook KS, Hunt CR, Spiegelman BM: Developmentally regulated mRNAs in 3T3-L1 adipocytes: analysis of transcriptional control. J Cell Biol 100 514–520, 1985
Dijan P, Phillips M, Green H: The activity of specific gene transcription in the adipose conversion of 3T3 cells. J Cell Physiol 124: 554–556, 1985
Herrera R, Ro HS, Robinson GS, Xanthopolous KO, Spiegelman BM: A direct role for C/EBP and the AP-1 binding site in gene expression linked to adipocyte differentiation. Mol Cell Biol 9: 5331–5339, 1989
Christy RJ, Yang VW, Natambi JM, Geiman DE, Landschulz WH, Friedman AD, Nakabeppu U, Kelly TJ, Lane MD: Differentiation-induced gene expression in 3T3-L1 preadipocytes: CCAAT/enhancer binding protein interacts with and activates the promoters of two adipocyte-specific genes. Genes & Development 3: 1323–1335, 1989
Distel RJ, Ro J H-S, Rasen BS, Groves DL, Spiegelman BM: Nucleoprotein complexes that regulate gene expression in adipocyte differentiation: direct participation of c-fos. Cell 49: 835–844, 1987
Yang VW, Christy RJ Cook JS, Kelly TJ, Lane MD: Mechanism of regulation of the 422(aP2) gene by cAMP during preadipocyte differentiation. Proc Natl Acad Sci USA 86: 3629–3633, 1989
Ntambi JM, Buhrow SA, Kaestner KH, Christy RJ, Sibley E, Kelly TJ, Lane MD: Differentiation-induced gene expression in 3T3-L1 preadipocytes. J Biol Chem 262: 17291–17300, 1988
Kaestner KH, McLenithan JC, Christy R, Braireiman LT Cornelius PC, Pekala PH, Lane MD: Sequence, tissue distribution, and differential expression of mRNA for a putative insulin-responsive glucose transporter in mouse 3T3-L1 adipocytes. Proc Natl Acad Sci USA 86: 3150–3154, 1989
Cook JS, Lucas JJ, Sibley E, Bolanowski MA, Christy RJ, Kelly TJ, Lane MD: Expression of the differentiation-induced gene for fatty acid-binding protein is activated by glucorticoids and cAMP. Proc Natl Acad Sci USA 85: 2949–2953, 1988
Kaestner KH, Christy RJ, Lane MD: Mouse insulin-responsive glucose transporter gene: characterization of the gene and transactivation by the CCAAT/enhancer binding protein. Proc Natl Sci USA 87: 251–255, 1990
McKnight SL, Lane MD Gluecksohn-Waelsch S: Is CCAAT/enhancer-binding protein a central regulator of energy metabolism? Genes & Development 3: 2021–2024, 1989
Old LJ: Tumor necrosis factor. Science 230: 630–632, 1985
Beutler B, Cerami A: Tumor necrosis factor and cachectin: two sides of the same biological coin. Nature 320: 584–588, 1986
Nathan CF: Secretory products of macrophages. J Clon Invest 79: 319–326, 1987
Carswell' Å, Old LJ, Kassel RL, Green S, Fiore N, Williamson B: Proc Natl Acad Sci USA 72, 3666–3670, 1975
Helson L, Green S, Carswell EÅ, Old LJ: Effect of tumour necrosis factor on cultured human melanoma cells. Nature 258: 13655–13662
Kull FC, Cuatrecasas P: Necrosin: purification and properties of a cyotoxin derived from a murine macrophage-like cell line. Proc Natl Acad Sci USA 81: 7932–7936, 1984
Sugarman BJ, Aggarwal BB Hass PE, Ïigari IS, Palladino M, Shepard HM: Recombinant human tumor necrosis factor-α: effects on proliferation of normal and transformed cellsin vitro. Science 230: 943–945, 1985
Vilcek J, Palombella V, Henrikson-De Stefano D, Swenson C, Feinman R, Hirai M, Tsujimoto M: Fibroblasts growth enhancing activity of tumor necrosis factor and its relationship to other polypeptide growth factors J Exp Med 163: 632–643, 1986
Cornelius P, Enerback S, Bjursell, Olivecrona T, Pekala PH: Regulation of lipoprotein lipase mRNA content in 3T3-L1 cells by tumour necrosis factor. Biochem J 249: 765–769, 1988
Pekala PH, Marlowe M, Heuvelman D, Connolly D: Regulation of hexose transport in aortic endothelial cells by VPF and TNF-α, but not by insulin, J Biol Chem 265: 18051–18054, 1990
Cornelius P, Lee MD, Marlowe M, Pekala PH: The growth factor-like effects of tumor necrosis factor-α. J Biol Chem 265: 20606–20516
Brenner DA, O'Hara M, Angel P, Chojkier M Karin M: Prolonged activation of jun and collagenase genes by tumour necrosis factor-α. Nature 337: 661–663, 1988
Lin J-X, Vilcek J: Tumor necrosis factor and interleukin-1 cause a rapid and transient stimulation of c-fos and c-myc mRNA levels in human fibroblasts. J Biol Chem 262: 11908–11911, 1987
Torti FÂ, Dieckmann B Beutler B, Cerami A, Ringold G: A macrophage factor inhibits adipocyte gene expression: anin vitro model of cachexia Science 229: 867–869, 1985
Torti FM, Torti SV Larrick JW, Ringold GM: Modulation of adipocyte differentiation by tumor necrosis factor and transforming growth factor beta. J Cell Biol 108: 1105–1113, 1989
Stone RL, Bernlohr DA: The role of retinoic acid in 3T3-L1 preadipocyte adipose conversion: antagonistic effects of stimulatory and inhibitory factors during differentiation. Differentiation, 45: 119–127, 1990
Curran T, Peters G, VanBeveren C Teich NM, Verma I: FBJ murine osteosarcoma virus: identification and molecular cloning of biological active proviral DNA. J Virol 44: 674–682, 1982
Cleveland D, Lopata MA, McDonald RJ, Cowan NJ Rutter WJ, Kirschner MJ: Number and evolutionary conservation of alphaand beta-tubulin and cytoplasmic beta- and gamma-actin genes using specific cloned cDNA probes Cell 20: 95–105, 1980
Lamph WW, Wamsley P, Sassone-Corsi P, Verma IM: Induction of proto-oncogene jun/AP-1 by serum and TPA. Nature 334: 629–631, 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
Van Snick J, Cayphas S, Szikora JP, Renauld JC Roost EV, Simpson RJ: cDNA cloning of murine interleukin-HP1: homology with human interleukin-6. Eur J Immunol 18: 193–197, 1988
Zerial M, Toschi L, Ryseck R-P Schuermann M, Muller R, Bravo R: The product of a novel growth factor activated gene, fos B, interacts with jun protein enhancing their DNA binding activity. EMBO J 8: 805–813, 1989
Chavrier P, Janssen-Timmen U Mattei M-G, Zerial M, Bravo R, Charnay P: Structure, chromosome location and expression of the mouse zinc finger gene krox-20: multiple gene products and coregulation with the proto-oncogene c-fos. Mol Cell Biol 9: 787–797, 1989
Chiu R, Boyle WJ, Meek J, Smeal T Hunter T, Karin M: The c-fos protein interacts with c-jun/AP-1 to stimulate transcription of AP-1 responsive genes. Cell 54: 541–552, 1988
Chiu C-P Moulds C, Coffman RL, Rennick D, Lee F: Multiple biological activities are expressed by a mouse interleukin 6 cDNA clone isolated from bone marrow stromal cells. Proc Natl Acad Sci USA 85: 8099–7103, 1988
Li L Hu J-S, Olsen EN: Different members of the jun proto-oncogene family exhibit distinct patterns of expression in response to type β transforming growth factor. J Biol Chem 264: 1556–1559, 1990
James PA, Taylor SM: Cellular differentiation, cytidine analogs and DNA methylation. Cell 20: 85–93, 1980
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Stephens, J.M., Butts, M., Stone, R. et al. Regulation of transcription factor mRNA accumulation during 3T3-L1 preadipocyte differentiation by antagonists of adipogenesis. Mol Cell Biochem 123, 63–71 (1993). https://doi.org/10.1007/BF01076476
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DOI: https://doi.org/10.1007/BF01076476