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. 1997 Dec 1;328(Pt 2):453–461. doi: 10.1042/bj3280453

Characterization of the promoter of human adipocyte hormone-sensitive lipase.

J Grober 1, H Laurell 1, R Blaise 1, B Fabry 1, S Schaak 1, C Holm 1, D Langin 1
PMCID: PMC1218941  PMID: 9371701

Abstract

Hormone-sensitive lipase (HSL) catalyses the rate-limiting step of adipose tissue lipolysis. The human HSL gene is composed of nine exons encoding the adipocyte form and a testis-specific coding exon. Northern blot analyses showed that human adipocytes express a 2.8 kb HSL mRNA, suggesting the presence of a short (20-150 bp) 5' untranslated region (5'-UTR). A single 5'-UTR of approx. 70 nt was detected in RNase H mapping experiments. Two 5'-UTRs of 70 and 170 nt respectively were obtained by rapid amplification of cDNA ends and cDNA library screenings. RNase protection experiments, with probes derived from the two products, showed that human adipocyte HSL mRNA contains only the 70 nt product. Primer extension analysis mapped the transcriptional start site 74 nt upstream of the start codon. In HT29, a human cell line expressing HSL, the presence of the short or the long 5'-UTR is mutually exclusive. The short and long 5'-UTR exons were located 1.5 and approx. 13 kb respectively upstream of the first coding exon. Various portions of the 5'-flanking region upstream of the short product exon were linked to the luciferase gene and transfected into cells that express HSL (HT29 cells and rat adipocytes) and do not express HSL (HeLa cells). High luciferase activity was found for constructs containing the sequence between nt -2400 and -86, but not for shorter constructs. An analysis of 14 kb of genomic sequence revealed the presence of five DNase I hypersensitive sites associated with active gene transcription. Three of the sites are located in the vicinity of the transcriptional start site and could be linked to the minimal promoter activity. Two of the sites are located downstream of the exon containing the start codon, suggesting the presence of intronic regulatory elements.

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Selected References

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

  1. Berlin I., Berlan M., Crespo-Laumonnier B., Landault C., Payan C., Puech A. J., Turpin G. Alterations in beta-adrenergic sensitivity and platelet alpha 2-adrenoceptors in obese women: effect of exercise and caloric restriction. Clin Sci (Lond) 1990 Jan;78(1):81–87. doi: 10.1042/cs0780081. [DOI] [PubMed] [Google Scholar]
  2. Boisclair Y. R., Brown A. L., Casola S., Rechler M. M. Three clustered Sp1 sites are required for efficient transcription of the TATA-less promoter of the gene for insulin-like growth factor-binding protein-2 from the rat. J Biol Chem. 1993 Nov 25;268(33):24892–24901. [PubMed] [Google Scholar]
  3. Bucher P. Weight matrix descriptions of four eukaryotic RNA polymerase II promoter elements derived from 502 unrelated promoter sequences. J Mol Biol. 1990 Apr 20;212(4):563–578. doi: 10.1016/0022-2836(90)90223-9. [DOI] [PubMed] [Google Scholar]
  4. Chomczynski P., Sacchi N. Single-step method of RNA isolation by acid guanidinium thiocyanate-phenol-chloroform extraction. Anal Biochem. 1987 Apr;162(1):156–159. doi: 10.1006/abio.1987.9999. [DOI] [PubMed] [Google Scholar]
  5. Das H. K., Jackson C. L., Miller D. A., Leff T., Breslow J. L. The human apolipoprotein C-II gene sequence contains a novel chromosome 19-specific minisatellite in its third intron. J Biol Chem. 1987 Apr 5;262(10):4787–4793. [PubMed] [Google Scholar]
  6. Elgin S. C. The formation and function of DNase I hypersensitive sites in the process of gene activation. J Biol Chem. 1988 Dec 25;263(36):19259–19262. [PubMed] [Google Scholar]
  7. Foufelle F., Lepetit N., Bosc D., Delzenne N., Morin J., Raymondjean M., Ferré P. DNase I hypersensitivity sites and nuclear protein binding on the fatty acid synthase gene: identification of an element with properties similar to known glucose-responsive elements. Biochem J. 1995 Jun 1;308(Pt 2):521–527. doi: 10.1042/bj3080521. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Frayn K. N., Coppack S. W., Fielding B. A., Humphreys S. M. Coordinated regulation of hormone-sensitive lipase and lipoprotein lipase in human adipose tissue in vivo: implications for the control of fat storage and fat mobilization. Adv Enzyme Regul. 1995;35:163–178. doi: 10.1016/0065-2571(94)00011-q. [DOI] [PubMed] [Google Scholar]
  9. Gautron S., Maire P., Hakim V., Kahn A. Regulation of the multiple promoters of the human aldolase A gene: response of its two ubiquitous promoters to agents promoting cell proliferation. Nucleic Acids Res. 1991 Feb 25;19(4):767–774. doi: 10.1093/nar/19.4.767. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Harada N., Utsumi T., Takagi Y. Tissue-specific expression of the human aromatase cytochrome P-450 gene by alternative use of multiple exons 1 and promoters, and switching of tissue-specific exons 1 in carcinogenesis. Proc Natl Acad Sci U S A. 1993 Dec 1;90(23):11312–11316. doi: 10.1073/pnas.90.23.11312. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Hellström L., Langin D., Reynisdottir S., Dauzats M., Arner P. Adipocyte lipolysis in normal weight subjects with obesity among first-degree relatives. Diabetologia. 1996 Aug;39(8):921–928. doi: 10.1007/BF00403911. [DOI] [PubMed] [Google Scholar]
  12. Holm C., Belfrage P., Fredrikson G. Human adipose tissue hormone-sensitive lipase: identification and comparison with other species. Biochim Biophys Acta. 1989 Nov 28;1006(2):193–197. doi: 10.1016/0005-2760(89)90195-1. [DOI] [PubMed] [Google Scholar]
  13. Holm C., Belfrage P., Fredrikson G. Immunological evidence for the presence of hormone-sensitive lipase in rat tissues other than adipose tissue. Biochem Biophys Res Commun. 1987 Oct 14;148(1):99–105. doi: 10.1016/0006-291x(87)91081-3. [DOI] [PubMed] [Google Scholar]
  14. Holm C., Kirchgessner T. G., Svenson K. L., Fredrikson G., Nilsson S., Miller C. G., Shively J. E., Heinzmann C., Sparkes R. S., Mohandas T. Hormone-sensitive lipase: sequence, expression, and chromosomal localization to 19 cent-q13.3. Science. 1988 Sep 16;241(4872):1503–1506. doi: 10.1126/science.3420405. [DOI] [PubMed] [Google Scholar]
  15. Holst L. S., Langin D., Mulder H., Laurell H., Grober J., Bergh A., Mohrenweiser H. W., Edgren G., Holm C. Molecular cloning, genomic organization, and expression of a testicular isoform of hormone-sensitive lipase. Genomics. 1996 Aug 1;35(3):441–447. doi: 10.1006/geno.1996.0383. [DOI] [PubMed] [Google Scholar]
  16. Javahery R., Khachi A., Lo K., Zenzie-Gregory B., Smale S. T. DNA sequence requirements for transcriptional initiator activity in mammalian cells. Mol Cell Biol. 1994 Jan;14(1):116–127. doi: 10.1128/mcb.14.1.116. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Langin D., Holm C., Lafontan M. Adipocyte hormone-sensitive lipase: a major regulator of lipid metabolism. Proc Nutr Soc. 1996 Mar;55(1B):93–109. doi: 10.1079/pns19960013. [DOI] [PubMed] [Google Scholar]
  18. Langin D., Laurell H., Holst L. S., Belfrage P., Holm C. Gene organization and primary structure of human hormone-sensitive lipase: possible significance of a sequence homology with a lipase of Moraxella TA144, an antarctic bacterium. Proc Natl Acad Sci U S A. 1993 Jun 1;90(11):4897–4901. doi: 10.1073/pnas.90.11.4897. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Laurell H., Grober J., Holst L. S., Holm C., Mohrenweiser H. W., Langin D. The hormone-sensitive lipase (LIPE) gene located on chromosome 19q13.1-->13.2 is not duplicated on 19p13.3. Int J Obes Relat Metab Disord. 1995 Aug;19(8):590–592. [PubMed] [Google Scholar]
  20. Mahendroo M. S., Mendelson C. R., Simpson E. R. Tissue-specific and hormonally controlled alternative promoters regulate aromatase cytochrome P450 gene expression in human adipose tissue. J Biol Chem. 1993 Sep 15;268(26):19463–19470. [PubMed] [Google Scholar]
  21. Plée-Gautier E., Grober J., Duplus E., Langin D., Forest C. Inhibition of hormone-sensitive lipase gene expression by cAMP and phorbol esters in 3T3-F442A and BFC-1 adipocytes. Biochem J. 1996 Sep 15;318(Pt 3):1057–1063. doi: 10.1042/bj3181057. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Pugh B. F., Tjian R. Mechanism of transcriptional activation by Sp1: evidence for coactivators. Cell. 1990 Jun 29;61(7):1187–1197. doi: 10.1016/0092-8674(90)90683-6. [DOI] [PubMed] [Google Scholar]
  23. Rajagopalan S., Wan D. F., Habib G. M., Sepulveda A. R., McLeod M. R., Lebovitz R. M., Lieberman M. W. Six mRNAs with different 5' ends are encoded by a single gamma-glutamyltransferase gene in mouse. Proc Natl Acad Sci U S A. 1993 Jul 1;90(13):6179–6183. doi: 10.1073/pnas.90.13.6179. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Remaury A., Laurell H., Grober J., Reynisdottir S., Dauzats M., Holm C., Langin D. Expression of hormone-sensitive lipase in the human colon adenocarcinoma cell line HT29. Biochem Biophys Res Commun. 1995 Feb 6;207(1):175–182. doi: 10.1006/bbrc.1995.1169. [DOI] [PubMed] [Google Scholar]
  25. Reynisdottir S., Eriksson M., Angelin B., Arner P. Impaired activation of adipocyte lipolysis in familial combined hyperlipidemia. J Clin Invest. 1995 May;95(5):2161–2169. doi: 10.1172/JCI117905. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Richard-Foy H., Hager G. L. Sequence-specific positioning of nucleosomes over the steroid-inducible MMTV promoter. EMBO J. 1987 Aug;6(8):2321–2328. doi: 10.1002/j.1460-2075.1987.tb02507.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Rolland V., Dugail I., Le Liepvre X., Lavau M. Evidence of increased glyceraldehyde-3-phosphate dehydrogenase and fatty acid synthetase promoter activities in transiently transfected adipocytes from genetically obese rats. J Biol Chem. 1995 Jan 20;270(3):1102–1106. doi: 10.1074/jbc.270.3.1102. [DOI] [PubMed] [Google Scholar]
  28. 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]
  29. Schwartz R. S., Jaeger L. F., Veith R. C., Lakshminarayan S. The effect of diet or exercise on plasma norepinephrine kinetics in moderately obese young men. Int J Obes. 1990 Jan;14(1):1–11. [PubMed] [Google Scholar]
  30. Sepulveda A. R., Carter B. Z., Habib G. M., Lebovitz R. M., Lieberman M. W. The mouse gamma-glutamyl transpeptidase gene is transcribed from at least five separate promoters. J Biol Chem. 1994 Apr 8;269(14):10699–10705. [PubMed] [Google Scholar]
  31. Stich V., Harant I., De Glisezinski I., Crampes F., Berlan M., Kunesova M., Hainer V., Dauzats M., Rivière D., Garrigues M. Adipose tissue lipolysis and hormone-sensitive lipase expression during very-low-calorie diet in obese female identical twins. J Clin Endocrinol Metab. 1997 Mar;82(3):739–744. doi: 10.1210/jcem.82.3.3793. [DOI] [PubMed] [Google Scholar]
  32. Tavernier G., Galitzky J., Valet P., Remaury A., Bouloumie A., Lafontan M., Langin D. Molecular mechanisms underlying regional variations of catecholamine-induced lipolysis in rat adipocytes. Am J Physiol. 1995 Jun;268(6 Pt 1):E1135–E1142. doi: 10.1152/ajpendo.1995.268.6.E1135. [DOI] [PubMed] [Google Scholar]

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