Skip to main content
SpringerLink
Log in

Sequences surrounding the transcription initiation site of the Arabidopsis enoyl-acyl carrier protein reductase gene control seed expression in transgenic tobacco

  • Published:
Plant Molecular Biology Aims and scope Submit manuscript

Abstract

The NADH-specific enoyl-acyl carrier protein (ACP) reductase, which catalyses the last reducing step during the fatty acid biosynthesis cycle, is encoded in Arabidopsis thaliana encoded by a single housekeeping gene (ENR-A) which is differentially expressed during plant development. To identify elements involved in its tissue-specific transcriptional control, a fragment comprising the 1470 bp region directly upstream of the ATG start codon of the ENR-A gene was fused to the uidA (GUS) reporter gene and analysed in transgenic Nicotiana tabacum plants. GUS activity found during development of the transgenic plants was similar to endogenous ENR protein levels found in both tobacco and Arabidopsis plants, except for developing flowers. In floral tissue the promoter fragment showed very little activity in contrast to the relatively high level of endogenous ENR expression. Successive deletions from the 5′ and 3′ regions of the promoter fragment revealed the presence of at least three elements which control GUS expression in different stages of development in the transgenic tobacco plants. First, expression in young developing leaves required both the presence of sequences between −329 to −201 relative to the transcription start and part of the untranslated leader comprising the first intron. Second, root-specific GUS expression was still observed after deletion of the 5′-upstream sequences up to 19 bp of the transcription initiation site. Further, the additional removal of the intron from the untranslated leader increased root-specific expression by ca. 4- to 5-fold. Third, high expression in seeds was still observed with the minimal upstream promoter segment of 19 bp. This seed expression level was found to be independent of the presence or absence of the intron in the untranslated leader. Finally, 3′ deletion of the leader sequence up to 17 bp of the transcription start greatly impaired GUS activity during all stages of plant development, suggesting that the deleted sequence of the leader either functions as an enhancer for transcription initiation or stabilizes the mRNA.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Similar content being viewed by others

References

  1. Ashton IA, Abulnaja KO, Pallett KE, Cole DJ, Harwood JL: The mechanism of inhibition of fatty acid synthase by the herbicide diflufenican. Phytochemistry 35: 587–590 (1994).

    Google Scholar 

  2. Baerson SR, Lamppa GK: Developmental regulation of an acyl carrier protein gene promoter in vegetative and reproductive tissues. Plant Mol Biol 22: 255–267 (1993).

    PubMed  Google Scholar 

  3. Baerson SR, Vanderheiden MG, Lamppa GK: Identification of domains in an Arabidopsis acyl carrier protein gene promoter required for maximal organ-specific expression. Plant Mol Biol 26: 1947–1959 (1994).

    PubMed  Google Scholar 

  4. Bao XM, Shorrosh BS, Ohlrogge JB: Isolation and characterization of an Arabidopsis biotin carboxylase gene and its promoter. Plant Mol Biol 35: 539–550 (1997).

    PubMed  Google Scholar 

  5. Bergler H, Fuchsbichler S, Hogenauer G, Turnowsky F: The enoyl-[acyl-carrier-protein] reductase (FabI) of Escherichia coli, which catalyzes a key regulatory step in fatty acid biosynthesis, accepts NADH and NADPH as cofactors and is inhibited by palmitoyl-CoA. Eur J Biochem 242: 689–694 (1996).

    PubMed  Google Scholar 

  6. Bolle C, Herrmann RG, Oelmuller R: Intron sequences are involved in the plastid-and light-dependent expression of the spinach PsaD gene. Plant J 10: 919–924 (1996).

    PubMed  Google Scholar 

  7. Caspar T, Quail PH: Promoter and leader regions involved in the expression of the Arabidopsis ferredoxin-A gene. Plant J 3: 161–174 (1993).

    Article  PubMed  Google Scholar 

  8. Conway LJ, Poethig RS: Mutations of Arabidopsis thaliana that transform leaves into cotyledons. Proc Natl Acad Sci USA 94: 10209–10214 (1997).

    PubMed  Google Scholar 

  9. de Block M, de Brouwer D, Tenning P: Transformation of Brassica napus and Brassica oleracea using Agrobacterium tumefaciens and the expression of the bar and neo genes in transgenic plants. Plant Physiol 91: 694–701 (1989).

    Google Scholar 

  10. de Boer G-J, Kater MM, Fawcett T, Slabas AR, Nijkamp HJJ, Stuitje AR: NADH-dependent enoyl-acyl carrier protein reductase: characterization of a housekeeping gene involved in storage lipid synthesis in seed of Arabidopsis and other plant species. Plant Physiol Biochem 36: 473–486 (1998).

    Google Scholar 

  11. de Silva J, Robinson SJ, Safford R: The isolation and functional characterisation of a B. napus acyl carrier protein 50-flanking region involved in the regulation of seed storage lipid synthesis. Plant Mol Biol 18: 1163–1172 (1992).

    PubMed  Google Scholar 

  12. Elborough KM, Winz R, Deka RK, Markham JE, White AJ, Rawsthorne S, Slabas AR: Biotin carboxyl carrier protein and carboxyltransferase subunits of the multi-subunit form of acetyl-CoA carboxylase from Brassica napus: cloning and analysis of expression during oilseed rape embryogenesis. Biochem J 315: 103–112 (1996).

    PubMed  Google Scholar 

  13. Fawcett T, Simon WJ, Swinhoe R, Shanklin J, Nishida I, Christie WW, Slabas AR: Expression of mRNA and steadystate levels of protein isoforms of enoyl-ACP reductase from Brassica napus. Plant Mol Biol 26: 155–163 (1994).

    PubMed  Google Scholar 

  14. Harwood JL: Recent advances in the biosynthesis of plant fatty acids. Biochim Biophys Acta 1301: 7–56 (1996).

    PubMed  Google Scholar 

  15. Hood EE, Gelvin SB, Melchers LS, Hoekema A: New Agrobacterium helper plasmids for gene transfer to plants. Transgenic Res 2: 208–218 (1993).

    Google Scholar 

  16. Horsch RB, Fry JE, Hoffmann NL, Eichholz D, Rogers SG, Fraley RT: A simple and general method for transferring genes into plants. Science 227: 1229–1231 (1985).

    Google Scholar 

  17. Jefferson RA, Kavanagh TA, Bevan MW: GUS fusions: _-glucuronidase as a sensitive and versatile gene fusion marker in higher plants. EMBO J 6: 3901–3907 (1987).

    PubMed  Google Scholar 

  18. Kater MM, Koningstein GM, Nijkamp HJJ, Stuitje AR: cDNA cloning and expression of Brassica napus enoyl-acyl carrier protein reductase in Escherichia coli. Plant Mol Biol 17: 895–909 (1991).

    PubMed  Google Scholar 

  19. Kater MM, Koningstein GM, Nijkamp HJJ, Stuitje AR: The use of a hybrid genetic system to study the functional relationship between prokaryotic and plant multi-enzyme fatty acid synthetase. Plant Mol Biol 25: 771–790 (1994).

    PubMed  Google Scholar 

  20. Ogas J, Cheng JC, Sung ZR, Somerville C: Cellular differentiation regulated by gibberellin in the Arabidopsis thaliana pickle mutant. Science 277: 91–94 (1997).

    PubMed  Google Scholar 

  21. Piffanelli P, Ross JHE, Murphy DJ: Intra-and extracellular lipid composition and associated gene expression patterns during pollen development in Brassica napus. Plant J 11: 549–562 (1997).

    PubMed  Google Scholar 

  22. Post-Beittenmiller D: Biochemistry and molecular biology of wax production in plants. Annu Rev Plant Physiol 47: 405–430 (1996).

    Google Scholar 

  23. Rose RE, DeJesus CE, Moylan SL, Ridge NP, Scherer DE, Knauf VC: The nucleotide sequence of a cDNA clone encoding acyl carrier protein (ACP) from Brassica campestris seeds. Nucl Acids Res 15: 7197 (1987).

    PubMed  Google Scholar 

  24. Scherer DE, Knauf VC: Isolation of complementary DNA clone for the acyl carrier protein-I of spinach. Plant Mol Biol 9: 127–134 (1987).

    Google Scholar 

  25. Shimakata T, Stumpf PK: The prokaryotic nature of the fatty acid synthetase of developing Carthamus tinctorius L. (saf-flower) seeds. Arch Biochem Biophys 217: 144–154 (1982).

    PubMed  Google Scholar 

  26. Shimakata T, Stumpf PK: Purification and characterisations of _-ketoacyl-[acyl-carrier-protein] reductase, _-hydroxyacyl-[acyl-carrier-protein] dehydrase, and enoyl-[acyl-carrierprotein] reductase from Spinacia oleracea leaves. Arch Biochem Biophys 218: 77–91 (1982).

    PubMed  Google Scholar 

  27. Sieburth LE, Meyerowitz EM: Molecular dissection of the AGAMOUS control region shows that cis elements for spatial regulation are located intragenically. Plant Cell 9: 355–365 (1997).

    PubMed  Google Scholar 

  28. Slabas AR, Harding J, Hellyer A, Sidebottom C, Gwynne H, Kessel R, Tombs MP: Enzymology of plant fatty acid biosynthesis. In: Siegenthaler PA, Eichenberger W (eds) Structure, Function and Metabolism of Plant Lipids, pp. 3–10. Elsevier, New York (1984).

    Google Scholar 

  29. Slocombe SP, Piffanelli P, Fairbairn D, Bowra S, Hatzopoulos P, Tsiantis M, Murphy DJ: Temporal and tissue-specific regulation of a Brassica napus stearoyl-acyl carrier protein desaturase gene. Plant Physiol 104: 1167–1176 (1994).

    PubMed  Google Scholar 

  30. Stark WM, Boocock MR, Sherratt DJ: Catalysis by sitespecific recombinases. Trends Genet 8: 432–439 (1992).

    PubMed  Google Scholar 

  31. Stumpf PK: Plants, fatty acids, compartments. Trends Biochem Sci 6: 173–176 (1981).

    Google Scholar 

  32. Turnowsky F, Fuchs K, Jeschek C, Hogenauer G: envM genes of Salmonella typhimurium and Escherichia coli. J Bact 171: 6555–6565 (1989).

    PubMed  Google Scholar 

  33. Verwoert IIGS, van der Linden K, Nijkamp HJJ, Stuitje AR: Developmental specific expression and organelle targeting of the E. coli fabD gene, encoding malonyl coenzyme A-acyl carrier protein transacylase in transgenic rape and tobacco seeds. Plant Mol Biol 26: 189–202 (1994).

    PubMed  Google Scholar 

  34. Weeks G, Wakil SJ: Studies on the mechanism of fatty acid synthesis. J Biol Chem 243: 1180–1189 (1968).

    PubMed  Google Scholar 

  35. Wen-Jun BG, Forde S: Efficient transformation of Agrobacterium spp. by high voltage electroporation. Nucl Acids Res 17: 8385 (1989).

    PubMed  Google Scholar 

Download references

Author information

Author notes

  1. Gert-Jan de Boer

    Present address: Department of Plant Biology, Carnegie Institution of Washington, 260 Panama Street, Stanford, CA, 94305, USA

Authors and Affiliations

  1. Department of Genetics, Institute for Molecular Biological Sciences, Vrije Universiteit, BioCentrum Amsterdam, De Boelelaan 1087, 1081 HV, Amsterdam, Netherlands

    Christa Testerink, Gerlof Pielage, H. John J. Nijkamp & Antoine R. Stuitje

Authors
  1. Gert-Jan de Boer
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Christa Testerink
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Gerlof Pielage
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. H. John J. Nijkamp
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Antoine R. Stuitje
    View author publications

    You can also search for this author in PubMed Google Scholar

Rights and permissions

Reprints and permissions

About this article

Cite this article

de Boer, GJ., Testerink, C., Pielage, G. et al. Sequences surrounding the transcription initiation site of the Arabidopsis enoyl-acyl carrier protein reductase gene control seed expression in transgenic tobacco. Plant Mol Biol 39, 1197–1207 (1999). https://doi.org/10.1023/A:1006129924683

Download citation

  • Issue Date:

  • DOI: https://doi.org/10.1023/A:1006129924683

Search

Navigation