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Proceedings of the National Academy of Sciences of the United States of America logoLink to Proceedings of the National Academy of Sciences of the United States of America
. 1992 Aug 15;89(16):7561–7565. doi: 10.1073/pnas.89.16.7561

Hibernoma formation in transgenic mice and isolation of a brown adipocyte cell line expressing the uncoupling protein gene.

S R Ross 1, L Choy 1, R A Graves 1, N Fox 1, V Solevjeva 1, S Klaus 1, D Ricquier 1, B M Spiegelman 1
PMCID: PMC49750  PMID: 1323843

Abstract

Transgenic mice were produced containing the adipocyte-specific regulatory region from the adipocyte P2 (aP2) gene linked to the simian virus 40 transforming genes. Most of the transgenic mice developed brown fat tumors (hibernomas) in their interscapular brown adipose tissue. Hibernoma formation was noticeable in some of the mice as early as 1 day after birth and most of the mice developed very large tumors by 1 month of age. All of the tumor tissue expressed the brown fat-specific uncoupling protein (UCP) gene as well as the aP2 gene. Several of the tumors have been used to establish cultured cell lines and at least one of these lines can be induced to differentiate into brown adipocytes. The cultured adipocytes express mRNA for UCP upon stimulation with N6,O2'-dibutyryladenosine 3',5'-cyclic monophosphate, norepinephrine, isoproterenol or D7114, a beta 3 adrenergic agonist. Thus, regulation of the key thermogenic gene UCP can now be studied in an established cell line.

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Selected References

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  1. Bouillaud F., Weissenbach J., Ricquier D. Complete cDNA-derived amino acid sequence of rat brown fat uncoupling protein. J Biol Chem. 1986 Feb 5;261(4):1487–1490. [PubMed] [Google Scholar]
  2. Brinster R. L., Chen H. Y., Messing A., van Dyke T., Levine A. J., Palmiter R. D. Transgenic mice harboring SV40 T-antigen genes develop characteristic brain tumors. Cell. 1984 Jun;37(2):367–379. doi: 10.1016/0092-8674(84)90367-2. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Casteilla L., Nouguès J., Reyne Y., Ricquier D. Differentiation of ovine brown adipocyte precursor cells in a chemically defined serum-free medium. Importance of glucocorticoids and age of animals. Eur J Biochem. 1991 May 23;198(1):195–199. doi: 10.1111/j.1432-1033.1991.tb16001.x. [DOI] [PubMed] [Google Scholar]
  4. Ceci J. D., Kovatch R. M., Swing D. A., Jones J. M., Snow C. M., Rosenberg M. P., Jenkins N. A., Copeland N. G., Meisler M. H. Transgenic mice carrying a murine amylase 2.2/SV40 T antigen fusion gene develop pancreatic acinar cell and stomach carcinomas. Oncogene. 1991 Feb;6(2):323–332. [PubMed] [Google Scholar]
  5. Champigny O., Ricquier D., Blondel O., Mayers R. M., Briscoe M. G., Holloway B. R. Beta 3-adrenergic receptor stimulation restores message and expression of brown-fat mitochondrial uncoupling protein in adult dogs. Proc Natl Acad Sci U S A. 1991 Dec 1;88(23):10774–10777. doi: 10.1073/pnas.88.23.10774. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. 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]
  7. Choi Y. W., Henrard D., Lee I., Ross S. R. The mouse mammary tumor virus long terminal repeat directs expression in epithelial and lymphoid cells of different tissues in transgenic mice. J Virol. 1987 Oct;61(10):3013–3019. doi: 10.1128/jvi.61.10.3013-3019.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Choi Y. W., Lee I. C., Ross S. R. Requirement for the simian virus 40 small tumor antigen in tumorigenesis in transgenic mice. Mol Cell Biol. 1988 Aug;8(8):3382–3390. doi: 10.1128/mcb.8.8.3382. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Efrat S., Linde S., Kofod H., Spector D., Delannoy M., Grant S., Hanahan D., Baekkeskov S. Beta-cell lines derived from transgenic mice expressing a hybrid insulin gene-oncogene. Proc Natl Acad Sci U S A. 1988 Dec;85(23):9037–9041. doi: 10.1073/pnas.85.23.9037. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Fox N., Crooke R., Hwang L. H., Schibler U., Knowles B. B., Solter D. Metastatic hibernomas in transgenic mice expressing an alpha-amylase-SV40 T antigen hybrid gene. Science. 1989 Apr 28;244(4903):460–463. doi: 10.1126/science.2785714. [DOI] [PubMed] [Google Scholar]
  11. Graves R. A., Tontonoz P., Ross S. R., Spiegelman B. M. Identification of a potent adipocyte-specific enhancer: involvement of an NF-1-like factor. Genes Dev. 1991 Mar;5(3):428–437. doi: 10.1101/gad.5.3.428. [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. Hamaguchi K., Gaskins H. R., Leiter E. H. NIT-1, a pancreatic beta-cell line established from a transgenic NOD/Lt mouse. Diabetes. 1991 Jul;40(7):842–849. doi: 10.2337/diab.40.7.842. [DOI] [PubMed] [Google Scholar]
  14. Hanahan D. Dissecting multistep tumorigenesis in transgenic mice. Annu Rev Genet. 1988;22:479–519. doi: 10.1146/annurev.ge.22.120188.002403. [DOI] [PubMed] [Google Scholar]
  15. Hanahan D. Heritable formation of pancreatic beta-cell tumours in transgenic mice expressing recombinant insulin/simian virus 40 oncogenes. Nature. 1985 May 9;315(6015):115–122. doi: 10.1038/315115a0. [DOI] [PubMed] [Google Scholar]
  16. Himms-Hagen J. Brown adipose tissue thermogenesis: interdisciplinary studies. FASEB J. 1990 Aug;4(11):2890–2898. [PubMed] [Google Scholar]
  17. Holloway B. R. Reactivation of brown adipose tissue. Proc Nutr Soc. 1989 Jul;48(2):225–230. doi: 10.1079/pns19890033. [DOI] [PubMed] [Google Scholar]
  18. Hopkins N. K., Gorman R. R. Regulation of 3T3-L1 fibroblast differentiation by prostacyclin (prostaglandin I2). Biochim Biophys Acta. 1981 Feb 23;663(2):457–466. doi: 10.1016/0005-2760(81)90174-0. [DOI] [PubMed] [Google Scholar]
  19. Jat P. S., Noble M. D., Ataliotis P., Tanaka Y., Yannoutsos N., Larsen L., Kioussis D. Direct derivation of conditionally immortal cell lines from an H-2Kb-tsA58 transgenic mouse. Proc Natl Acad Sci U S A. 1991 Jun 15;88(12):5096–5100. doi: 10.1073/pnas.88.12.5096. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Klaus S., Cassard-Doulcier A. M., Ricquier D. Development of Phodopus sungorus brown preadipocytes in primary cell culture: effect of an atypical beta-adrenergic agonist, insulin, and triiodothyronine on differentiation, mitochondrial development, and expression of the uncoupling protein UCP. J Cell Biol. 1991 Dec;115(6):1783–1790. doi: 10.1083/jcb.115.6.1783. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Klaus S., Casteilla L., Bouillaud F., Ricquier D. The uncoupling protein UCP: a membraneous mitochondrial ion carrier exclusively expressed in brown adipose tissue. Int J Biochem. 1991;23(9):791–801. doi: 10.1016/0020-711x(91)90062-r. [DOI] [PubMed] [Google Scholar]
  22. Klingenberg M. Nucleotide binding to uncoupling protein. Mechanism of control by protonation. Biochemistry. 1988 Jan 26;27(2):781–791. doi: 10.1021/bi00402a044. [DOI] [PubMed] [Google Scholar]
  23. 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]
  24. Mahon K. A., Chepelinsky A. B., Khillan J. S., Overbeek P. A., Piatigorsky J., Westphal H. Oncogenesis of the lens in transgenic mice. Science. 1987 Mar 27;235(4796):1622–1628. doi: 10.1126/science.3029873. [DOI] [PubMed] [Google Scholar]
  25. Melton D. A., Krieg P. A., Rebagliati M. R., Maniatis T., Zinn K., Green M. R. Efficient in vitro synthesis of biologically active RNA and RNA hybridization probes from plasmids containing a bacteriophage SP6 promoter. Nucleic Acids Res. 1984 Sep 25;12(18):7035–7056. doi: 10.1093/nar/12.18.7035. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Muzzin P., Revelli J. P., Ricquier D., Meier M. K., Assimacopoulos-Jeannet F., Giacobino J. P. The novel thermogenic beta-adrenergic agonist Ro 16-8714 increases the interscapular brown-fat beta-receptor-adenylate cyclase and the uncoupling-protein mRNA level in obese (fa/fa) Zucker rats. Biochem J. 1989 Aug 1;261(3):721–724. doi: 10.1042/bj2610721. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Ornitz D. M., Hammer R. E., Messing A., Palmiter R. D., Brinster R. L. Pancreatic neoplasia induced by SV40 T-antigen expression in acinar cells of transgenic mice. Science. 1987 Oct 9;238(4824):188–193. doi: 10.1126/science.2821617. [DOI] [PubMed] [Google Scholar]
  28. Ornitz D. M., Palmiter R. D., Messing A., Hammer R. E., Pinkert C. A., Brinster R. L. Elastase I promoter directs expression of human growth hormone and SV40 T antigen genes to pancreatic acinar cells in transgenic mice. Cold Spring Harb Symp Quant Biol. 1985;50:399–409. doi: 10.1101/sqb.1985.050.01.050. [DOI] [PubMed] [Google Scholar]
  29. Rehnmark S., Kopecký J., Jacobsson A., Néchad M., Herron D., Nelson B. D., Obregon M. J., Nedergaard J., Cannon B. Brown adipocytes differentiated in vitro can express the gene for the uncoupling protein thermogenin: effects of hypothyroidism and norepinephrine. Exp Cell Res. 1989 May;182(1):75–83. doi: 10.1016/0014-4827(89)90280-2. [DOI] [PubMed] [Google Scholar]
  30. Ross S. R., Graves R. A., Greenstein A., Platt K. A., Shyu H. L., Mellovitz B., Spiegelman B. M. A fat-specific enhancer is the primary determinant of gene expression for adipocyte P2 in vivo. Proc Natl Acad Sci U S A. 1990 Dec;87(24):9590–9594. doi: 10.1073/pnas.87.24.9590. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Rothwell N. J., Stock M. J. A role for brown adipose tissue in diet-induced thermogenesis. Nature. 1979 Sep 6;281(5726):31–35. doi: 10.1038/281031a0. [DOI] [PubMed] [Google Scholar]
  32. Rothwell N. J., Stock M. J., Stribling D. Diet-induced thermogenesis. Pharmacol Ther. 1982;17(2):251–268. doi: 10.1016/0163-7258(82)90016-x. [DOI] [PubMed] [Google Scholar]
  33. Russell T. R., Ho R. Conversion of 3T3 fibroblasts into adipose cells: triggering of differentiation by prostaglandin F2alpha and 1-methyl-3-isobutyl xanthine. Proc Natl Acad Sci U S A. 1976 Dec;73(12):4516–4520. doi: 10.1073/pnas.73.12.4516. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. 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]
  35. Windle J. J., Weiner R. I., Mellon P. L. Cell lines of the pituitary gonadotrope lineage derived by targeted oncogenesis in transgenic mice. Mol Endocrinol. 1990 Apr;4(4):597–603. doi: 10.1210/mend-4-4-597. [DOI] [PubMed] [Google Scholar]
  36. Yanai N., Suzuki M., Obinata M. Hepatocyte cell lines established from transgenic mice harboring temperature-sensitive simian virus 40 large T-antigen gene. Exp Cell Res. 1991 Nov;197(1):50–56. doi: 10.1016/0014-4827(91)90478-d. [DOI] [PubMed] [Google Scholar]
  37. Young P., Arch J. R., Ashwell M. Brown adipose tissue in the parametrial fat pad of the mouse. FEBS Lett. 1984 Feb 13;167(1):10–14. doi: 10.1016/0014-5793(84)80822-4. [DOI] [PubMed] [Google Scholar]

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