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. 1991 Dec;10(12):3787–3793. doi: 10.1002/j.1460-2075.1991.tb04948.x

The CCAAT/enhancer binding protein and its role in adipocyte differentiation: evidence for direct involvement in terminal adipocyte development.

L Samuelsson 1, K Strömberg 1, K Vikman 1, G Bjursell 1, S Enerbäck 1
PMCID: PMC453115  PMID: 1935900

Abstract

During the course of differentiation of preadipocytes into adipocytes, several differentiation-linked genes are activated synchronously with morphological changes. To follow this process we have used 3T3-F442A cells, known to undergo adipocyte conversion with high frequency. Accumulation of lipid droplets in the cytoplasm constitutes an easily visualized sign of the terminally differentiated phenotype. In this report we demonstrate that expression of the CCAAT/enhancer binding protein (C/EBP) is an important factor in determining the ability to accumulate lipid droplets in terminally differentiated adipocytes. In one experiment we can suppress C/EBP expression through administration of hydrocortisone to differentiating 3T3-F442A cells, which is accompanied by an inability of the cells to accumulate lipid. In another experiment a C/EBP antisense expression vector has been stably introduced into 3T3-F442A cells and as compared with control cells, a 62% decrease of C/EBP mRNA (p less than 0.01) is demonstrated. This decrease of C/EBP mRNA is accompanied by a change in cellular morphology characterized by a reduced ability to form lipid droplets. We can also demonstrate a correlation between the degree of reduction of C/EBP mRNA and the amount of lipid present in the cells. These findings strongly support the view that C/EBP is a necessary component of terminal adipocyte differentiation.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Amri E. Z., Dani C., Doglio A., Grimaldi P., Ailhaud G. Coupling of growth arrest and expression of early markers during adipose conversion of preadipocyte cell lines. Biochem Biophys Res Commun. 1986 Jun 13;137(2):903–910. doi: 10.1016/0006-291x(86)91165-4. [DOI] [PubMed] [Google Scholar]
  2. Birkenmeier E. H., Gwynn B., Howard S., Jerry J., Gordon J. I., Landschulz W. H., McKnight S. L. Tissue-specific expression, developmental regulation, and genetic mapping of the gene encoding CCAAT/enhancer binding protein. Genes Dev. 1989 Aug;3(8):1146–1156. doi: 10.1101/gad.3.8.1146. [DOI] [PubMed] [Google Scholar]
  3. Chen S., Teicher L. C., Kazim D., Pollack R. E., Wise L. S. Commitment of mouse fibroblasts to adipocyte differentiation by DNA transfection. Science. 1989 May 5;244(4904):582–585. doi: 10.1126/science.2470149. [DOI] [PubMed] [Google Scholar]
  4. Chirgwin J. M., Przybyla A. E., MacDonald R. J., Rutter W. J. Isolation of biologically active ribonucleic acid from sources enriched in ribonuclease. Biochemistry. 1979 Nov 27;18(24):5294–5299. doi: 10.1021/bi00591a005. [DOI] [PubMed] [Google Scholar]
  5. Christy R. J., Yang V. W., Ntambi J. M., Geiman D. E., Landschulz W. H., Friedman A. D., Nakabeppu Y., Kelly T. J., Lane M. D. Differentiation-induced gene expression in 3T3-L1 preadipocytes: CCAAT/enhancer binding protein interacts with and activates the promoters of two adipocyte-specific genes. Genes Dev. 1989 Sep;3(9):1323–1335. doi: 10.1101/gad.3.9.1323. [DOI] [PubMed] [Google Scholar]
  6. Cleveland D. W., Lopata M. A., MacDonald R. J., Cowan N. J., Rutter W. J., Kirschner M. W. Number and evolutionary conservation of alpha- and beta-tubulin and cytoplasmic beta- and gamma-actin genes using specific cloned cDNA probes. Cell. 1980 May;20(1):95–105. doi: 10.1016/0092-8674(80)90238-x. [DOI] [PubMed] [Google Scholar]
  7. Davis R. L., Weintraub H., Lassar A. B. Expression of a single transfected cDNA converts fibroblasts to myoblasts. Cell. 1987 Dec 24;51(6):987–1000. doi: 10.1016/0092-8674(87)90585-x. [DOI] [PubMed] [Google Scholar]
  8. Dobson D. E., Groves D. L., Spiegelman B. M. Nucleotide sequence and hormonal regulation of mouse glycerophosphate dehydrogenase mRNA during adipocyte and muscle cell differentiation. J Biol Chem. 1987 Feb 5;262(4):1804–1809. [PubMed] [Google Scholar]
  9. Durnam D. M., Palmiter R. D. A practical approach for quantitating specific mRNAs by solution hybridization. Anal Biochem. 1983 Jun;131(2):385–393. doi: 10.1016/0003-2697(83)90188-4. [DOI] [PubMed] [Google Scholar]
  10. Enerbäck S., Semb H., Bengtsson-Olivecrona G., Carlsson P., Hermansson M. L., Olivecrona T., Bjursell G. Molecular cloning and sequence analysis of cDNA encoding lipoprotein lipase of guinea pig. Gene. 1987;58(1):1–12. doi: 10.1016/0378-1119(87)90023-0. [DOI] [PubMed] [Google Scholar]
  11. Feinberg A. P., Vogelstein B. A technique for radiolabeling DNA restriction endonuclease fragments to high specific activity. Anal Biochem. 1983 Jul 1;132(1):6–13. doi: 10.1016/0003-2697(83)90418-9. [DOI] [PubMed] [Google Scholar]
  12. Green H., Kehinde O. Spontaneous heritable changes leading to increased adipose conversion in 3T3 cells. Cell. 1976 Jan;7(1):105–113. doi: 10.1016/0092-8674(76)90260-9. [DOI] [PubMed] [Google Scholar]
  13. Green P. J., Pines O., Inouye M. The role of antisense RNA in gene regulation. Annu Rev Biochem. 1986;55:569–597. doi: 10.1146/annurev.bi.55.070186.003033. [DOI] [PubMed] [Google Scholar]
  14. Herrera R., Ro H. S., Robinson G. S., Xanthopoulos K. G., Spiegelman B. M. A direct role for C/EBP and the AP-I-binding site in gene expression linked to adipocyte differentiation. Mol Cell Biol. 1989 Dec;9(12):5331–5339. doi: 10.1128/mcb.9.12.5331. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Kaestner K. H., Christy R. J., Lane M. D. Mouse insulin-responsive glucose transporter gene: characterization of the gene and trans-activation by the CCAAT/enhancer binding protein. Proc Natl Acad Sci U S A. 1990 Jan;87(1):251–255. doi: 10.1073/pnas.87.1.251. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Kim S. K., Wold B. J. Stable reduction of thymidine kinase activity in cells expressing high levels of anti-sense RNA. Cell. 1985 Aug;42(1):129–138. doi: 10.1016/s0092-8674(85)80108-2. [DOI] [PubMed] [Google Scholar]
  17. Kozak L. P., Birkenmeier E. H. Mouse sn-glycerol-3-phosphate dehydrogenase: molecular cloning and genetic mapping of a cDNA sequence. Proc Natl Acad Sci U S A. 1983 May;80(10):3020–3024. doi: 10.1073/pnas.80.10.3020. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Kuri-Harcuch W., Green H. Adipose conversion of 3T3 cells depends on a serum factor. Proc Natl Acad Sci U S A. 1978 Dec;75(12):6107–6109. doi: 10.1073/pnas.75.12.6107. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Kuri-Harcuch W., Wise L. S., Green H. Interruption of the adipose conversion of 3T3 cells by biotin deficiency: differentiation without triglyceride accumulation. Cell. 1978 May;14(1):53–59. doi: 10.1016/0092-8674(78)90300-8. [DOI] [PubMed] [Google Scholar]
  20. Landschulz W. H., Johnson P. F., Adashi E. Y., Graves B. J., McKnight S. L. Isolation of a recombinant copy of the gene encoding C/EBP. Genes Dev. 1988 Jul;2(7):786–800. doi: 10.1101/gad.2.7.786. [DOI] [PubMed] [Google Scholar]
  21. Lehrach H., Diamond D., Wozney J. M., Boedtker H. RNA molecular weight determinations by gel electrophoresis under denaturing conditions, a critical reexamination. Biochemistry. 1977 Oct 18;16(21):4743–4751. doi: 10.1021/bi00640a033. [DOI] [PubMed] [Google Scholar]
  22. McKnight S. L., Lane M. D., Gluecksohn-Waelsch S. Is CCAAT/enhancer-binding protein a central regulator of energy metabolism? Genes Dev. 1989 Dec;3(12B):2021–2024. doi: 10.1101/gad.3.12b.2021. [DOI] [PubMed] [Google Scholar]
  23. Moore L., Pastan I. Effect of cell density on energy-dependent calcium uptake by Balb/c 3T3 membranes is independent of protein synthesis and attachment to substratum. J Cell Physiol. 1979 Oct;101(1):109–116. doi: 10.1002/jcp.1041010113. [DOI] [PubMed] [Google Scholar]
  24. Pairault J., Green H. A study of the adipose conversion of suspended 3T3 cells by using glycerophosphate dehydrogenase as differentiation marker. Proc Natl Acad Sci U S A. 1979 Oct;76(10):5138–5142. doi: 10.1073/pnas.76.10.5138. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Pairault J., Lasnier F. Control of the adipogenic differentiation of 3T3-F442A cells by retinoic acid, dexamethasone, and insulin: a topographic analysis. J Cell Physiol. 1987 Aug;132(2):279–286. doi: 10.1002/jcp.1041320212. [DOI] [PubMed] [Google Scholar]
  26. Paulssen R. H., Paulssen E. J., Aleström P., Gordeladze J. O., Gautvik K. M. Specific antisense RNA inhibition of growth hormone production in differentiated rat pituitary tumour cells. Biochem Biophys Res Commun. 1990 Aug 31;171(1):293–300. doi: 10.1016/0006-291x(90)91391-5. [DOI] [PubMed] [Google Scholar]
  27. Southern P. J., Berg P. Transformation of mammalian cells to antibiotic resistance with a bacterial gene under control of the SV40 early region promoter. J Mol Appl Genet. 1982;1(4):327–341. [PubMed] [Google Scholar]
  28. Spiegelman B. M., Farmer S. R. Decreases in tubulin and actin gene expression prior to morphological differentiation of 3T3 adipocytes. Cell. 1982 May;29(1):53–60. doi: 10.1016/0092-8674(82)90089-7. [DOI] [PubMed] [Google Scholar]
  29. Spiegelman B. M., Frank M., Green H. Molecular cloning of mRNA from 3T3 adipocytes. Regulation of mRNA content for glycerophosphate dehydrogenase and other differentiation-dependent proteins during adipocyte development. J Biol Chem. 1983 Aug 25;258(16):10083–10089. [PubMed] [Google Scholar]
  30. Umek R. M., Friedman A. D., McKnight S. L. CCAAT-enhancer binding protein: a component of a differentiation switch. Science. 1991 Jan 18;251(4991):288–292. doi: 10.1126/science.1987644. [DOI] [PubMed] [Google Scholar]
  31. Wright T. R. The genetics of embryogenesis in Drosophila. Adv Genet. 1970;15:261–395. doi: 10.1016/s0065-2660(08)60075-9. [DOI] [PubMed] [Google Scholar]

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