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. 1991 Jul;10(7):1767–1775. doi: 10.1002/j.1460-2075.1991.tb07701.x

Exonic sequences are required for elicitor and light activation of a plant defense gene, but promoter sequences are sufficient for tissue specific expression.

C J Douglas 1, K D Hauffe 1, M E Ites-Morales 1, M Ellard 1, U Paszkowski 1, K Hahlbrock 1, J L Dangl 1
PMCID: PMC452848  PMID: 2050114

Abstract

The parsley 4CL-1 gene encodes 4-coumarate:CoA ligase, a key enzyme of general phenylpropanoid metabolism. As well as being transcriptionally activated by such stresses as pathogen infection, UV-irradiation, and wounding, expression of 4CL-1 is developmentally regulated. In this paper we present evidence that 4CL-1 cis-acting elements which control stress-induced and developmental expression are physically separated. The ability of a series of 4CL gene constructions to respond to elicitor and light in stably or transiently transformed parsley cells was tested. While inducible expression was observed from all templates in which the 4CL-1 structural gene was fused to the 4CL-1 promoter, fusions of the promoter to the GUS reporter gene were completely unresponsive. The element(s) required for responsiveness appear to be exonic, since 4CL-1 introns and 3' flanking DNA had no effect on inducibility. Furthermore, this unconventional regulatory mode operates in transgenic tobacco plants, where we show that a 4CL-1 promoter fragment specifies correct cell-specific expression when fused to GUS yet is unresponsive to elicitor and light.

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Selected References

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

  1. A simple and general method for transferring genes into plants. Science. 1985 Mar 8;227(4691):1229–1231. doi: 10.1126/science.227.4691.1229. [DOI] [PubMed] [Google Scholar]
  2. An G., Mitra A., Choi H. K., Costa M. A., An K., Thornburg R. W., Ryan C. A. Functional analysis of the 3' control region of the potato wound-inducible proteinase inhibitor II gene. Plant Cell. 1989 Jan;1(1):115–122. doi: 10.1105/tpc.1.1.115. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Ayers A. R., Ebel J., Valent B., Albersheim P. Host-Pathogen Interactions: X. Fractionation and Biological Activity of an Elicitor Isolated from the Mycelial Walls of Phytophthora megasperma var. sojae. Plant Physiol. 1976 May;57(5):760–765. doi: 10.1104/pp.57.5.760. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Basler K., Siegrist P., Hafen E. The spatial and temporal expression pattern of sevenless is exclusively controlled by gene-internal elements. EMBO J. 1989 Aug;8(8):2381–2386. doi: 10.1002/j.1460-2075.1989.tb08367.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Bevan M. Binary Agrobacterium vectors for plant transformation. Nucleic Acids Res. 1984 Nov 26;12(22):8711–8721. doi: 10.1093/nar/12.22.8711. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Bevan M., Shufflebottom D., Edwards K., Jefferson R., Schuch W. Tissue- and cell-specific activity of a phenylalanine ammonia-lyase promoter in transgenic plants. EMBO J. 1989 Jul;8(7):1899–1906. doi: 10.1002/j.1460-2075.1989.tb03592.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Block A., Dangl J. L., Hahlbrock K., Schulze-Lefert P. Functional borders, genetic fine structure, and distance requirements of cis elements mediating light responsiveness of the parsley chalcone synthase promoter. Proc Natl Acad Sci U S A. 1990 Jul;87(14):5387–5391. doi: 10.1073/pnas.87.14.5387. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Bornstein P., McKay J., Liska D. J., Apone S., Devarayalu S. Interactions between the promoter and first intron are involved in transcriptional control of alpha 1(I) collagen gene expression. Mol Cell Biol. 1988 Nov;8(11):4851–4857. doi: 10.1128/mcb.8.11.4851. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Callis J., Fromm M., Walbot V. Introns increase gene expression in cultured maize cells. Genes Dev. 1987 Dec;1(10):1183–1200. doi: 10.1101/gad.1.10.1183. [DOI] [PubMed] [Google Scholar]
  10. Chappell J., Nable R. Induction of sesquiterpenoid biosynthesis in tobacco cell suspension cultures by fungal elicitor. Plant Physiol. 1987 Oct;85(2):469–473. doi: 10.1104/pp.85.2.469. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Coulombe B., Ponton A., Daigneault L., Williams B. R., Skup D. Presence of transcription regulatory elements within an intron of the virus-inducible murine TIMP gene. Mol Cell Biol. 1988 Aug;8(8):3227–3234. doi: 10.1128/mcb.8.8.3227. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Dangl J. L., Hauffe K. D., Lipphardt S., Hahlbrock K., Scheel D. Parsley protoplasts retain differential responsiveness to u.v. light and fungal elicitor. EMBO J. 1987 Sep;6(9):2551–2556. doi: 10.1002/j.1460-2075.1987.tb02543.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Dean C., Favreau M., Bond-Nutter D., Bedbrook J., Dunsmuir P. Sequences downstream of translation start regulate quantitative expression of two petunia rbcS genes. Plant Cell. 1989 Feb;1(2):201–208. doi: 10.1105/tpc.1.2.201. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Dixon R. A., Harrison M. J. Activation, structure, and organization of genes involved in microbial defense in plants. Adv Genet. 1990;28:165–234. doi: 10.1016/s0065-2660(08)60527-1. [DOI] [PubMed] [Google Scholar]
  15. Douglas C., Hoffmann H., Schulz W., Hahlbrock K. Structure and elicitor or u.v.-light-stimulated expression of two 4-coumarate:CoA ligase genes in parsley. EMBO J. 1987 May;6(5):1189–1195. doi: 10.1002/j.1460-2075.1987.tb02353.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Dron M., Clouse S. D., Dixon R. A., Lawton M. A., Lamb C. J. Glutathione and fungal elicitor regulation of a plant defense gene promoter in electroporated protoplasts. Proc Natl Acad Sci U S A. 1988 Sep;85(18):6738–6742. doi: 10.1073/pnas.85.18.6738. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Elliott R. C., Dickey L. F., White M. J., Thompson W. F. cis-Acting Elements for Light Regulation of Pea Ferredoxin I Gene Expression Are Located within Transcribed Sequences. Plant Cell. 1989 Jul;1(7):691–698. doi: 10.1105/tpc.1.7.691. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Fritzemeier K. H., Cretin C., Kombrink E., Rohwer F., Taylor J., Scheel D., Hahlbrock K. Transient Induction of Phenylalanine Ammonia-Lyase and 4-Coumarate: CoA Ligase mRNAs in Potato Leaves Infected with Virulent or Avirulent Races of Phytophthora infestans. Plant Physiol. 1987 Sep;85(1):34–41. doi: 10.1104/pp.85.1.34. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Geiduschek E. P., Tocchini-Valentini G. P. Transcription by RNA polymerase III. Annu Rev Biochem. 1988;57:873–914. doi: 10.1146/annurev.bi.57.070188.004301. [DOI] [PubMed] [Google Scholar]
  20. Gruissem W. Chloroplast gene expression: how plants turn their plastids on. Cell. 1989 Jan 27;56(2):161–170. doi: 10.1016/0092-8674(89)90889-1. [DOI] [PubMed] [Google Scholar]
  21. Jefferson R. A., Kavanagh T. A., Bevan M. W. GUS fusions: beta-glucuronidase as a sensitive and versatile gene fusion marker in higher plants. EMBO J. 1987 Dec 20;6(13):3901–3907. doi: 10.1002/j.1460-2075.1987.tb02730.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Kaulen Hildegard, Schell Jeff, Kreuzaler Fritz. Light-induced expression of the chimeric chalcone synthase-NPTII gene in tobacco cells. EMBO J. 1986 Jan;5(1):1–8. doi: 10.1002/j.1460-2075.1986.tb04169.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Kozak M. Possible role of flanking nucleotides in recognition of the AUG initiator codon by eukaryotic ribosomes. Nucleic Acids Res. 1981 Oct 24;9(20):5233–5252. doi: 10.1093/nar/9.20.5233. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Krauskopf A., Resnekov O., Aloni Y. A cis downstream element participates in regulation of in vitro transcription initiation from the P38 promoter of minute virus of mice. J Virol. 1990 Jan;64(1):354–360. doi: 10.1128/jvi.64.1.354-360.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Kuhn D. N., Chappell J., Boudet A., Hahlbrock K. Induction of phenylalanine ammonia-lyase and 4-coumarate:CoA ligase mRNAs in cultured plant cells by UV light or fungal elicitor. Proc Natl Acad Sci U S A. 1984 Feb;81(4):1102–1106. doi: 10.1073/pnas.81.4.1102. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Lamb C. J., Lawton M. A., Dron M., Dixon R. A. Signals and transduction mechanisms for activation of plant defenses against microbial attack. Cell. 1989 Jan 27;56(2):215–224. doi: 10.1016/0092-8674(89)90894-5. [DOI] [PubMed] [Google Scholar]
  27. Lawton M. A., Lamb C. J. Transcriptional activation of plant defense genes by fungal elicitor, wounding, and infection. Mol Cell Biol. 1987 Jan;7(1):335–341. doi: 10.1128/mcb.7.1.335. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Leong K., Lee W., Berk A. J. High-level transcription from the adenovirus major late promoter requires downstream binding sites for late-phase-specific factors. J Virol. 1990 Jan;64(1):51–60. doi: 10.1128/jvi.64.1.51-60.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Liang X. W., Dron M., Cramer C. L., Dixon R. A., Lamb C. J. Differential regulation of phenylalanine ammonia-lyase genes during plant development and by environmental cues. J Biol Chem. 1989 Aug 25;264(24):14486–14492. [PubMed] [Google Scholar]
  30. Liang X. W., Dron M., Schmid J., Dixon R. A., Lamb C. J. Developmental and environmental regulation of a phenylalanine ammonia-lyase-beta-glucuronidase gene fusion in transgenic tobacco plants. Proc Natl Acad Sci U S A. 1989 Dec;86(23):9284–9288. doi: 10.1073/pnas.86.23.9284. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Lipphardt S., Brettschneider R., Kreuzaler F., Schell J., Dangl J. L. UV-inducible transient expression in parsley protoplasts identifies regulatory cis-elements of a chimeric Antirrhinum majus chalcone synthase gene. EMBO J. 1988 Dec 20;7(13):4027–4033. doi: 10.1002/j.1460-2075.1988.tb03296.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Logemann J., Schell J., Willmitzer L. Improved method for the isolation of RNA from plant tissues. Anal Biochem. 1987 May 15;163(1):16–20. doi: 10.1016/0003-2697(87)90086-8. [DOI] [PubMed] [Google Scholar]
  33. Lois R., Dietrich A., Hahlbrock K., Schulz W. A phenylalanine ammonia-lyase gene from parsley: structure, regulation and identification of elicitor and light responsive cis-acting elements. EMBO J. 1989 Jun;8(6):1641–1648. doi: 10.1002/j.1460-2075.1989.tb03554.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Lozoya E., Hoffmann H., Douglas C., Schulz W., Scheel D., Hahlbrock K. Primary structures and catalytic properties of isoenzymes encoded by the two 4-coumarate: CoA ligase genes in parsley. Eur J Biochem. 1988 Oct 1;176(3):661–667. doi: 10.1111/j.1432-1033.1988.tb14328.x. [DOI] [PubMed] [Google Scholar]
  35. Moore D. D., Marks A. R., Buckley D. I., Kapler G., Payvar F., Goodman H. M. The first intron of the human growth hormone gene contains a binding site for glucocorticoid receptor. Proc Natl Acad Sci U S A. 1985 Feb;82(3):699–702. doi: 10.1073/pnas.82.3.699. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Rapid switching of plant gene expression induced by fungal elicitor. Science. 1985 Mar 8;227(4691):1240–1243. doi: 10.1126/science.227.4691.1240. [DOI] [PubMed] [Google Scholar]
  37. Schmelzer E., Jahnen W., Hahlbrock K. In situ localization of light-induced chalcone synthase mRNA, chalcone synthase, and flavonoid end products in epidermal cells of parsley leaves. Proc Natl Acad Sci U S A. 1988 May;85(9):2989–2993. doi: 10.1073/pnas.85.9.2989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Schmelzer E., Kruger-Lebus S., Hahlbrock K. Temporal and Spatial Patterns of Gene Expression around Sites of Attempted Fungal Infection in Parsley Leaves. Plant Cell. 1989 Oct;1(10):993–1001. doi: 10.1105/tpc.1.10.993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Schmid J., Doerner P. W., Clouse S. D., Dixon R. A., Lamb C. J. Developmental and environmental regulation of a bean chalcone synthase promoter in transgenic tobacco. Plant Cell. 1990 Jul;2(7):619–631. doi: 10.1105/tpc.2.7.619. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Schulze-Lefert P., Becker-André M., Schulz W., Hahlbrock K., Dangl J. L. Functional architecture of the light-responsive chalcone synthase promoter from parsley. Plant Cell. 1989 Jul;1(7):707–714. doi: 10.1105/tpc.1.7.707. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Schulze-Lefert P., Dangl J. L., Becker-André M., Hahlbrock K., Schulz W. Inducible in vivo DNA footprints define sequences necessary for UV light activation of the parsley chalcone synthase gene. EMBO J. 1989 Mar;8(3):651–656. doi: 10.1002/j.1460-2075.1989.tb03422.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Staiger D., Kaulen H., Schell J. A Nuclear Factor Recognizing a Positive Regulatory Upstream Element of the Antirrhinum majus Chalcone Synthase Promoter. Plant Physiol. 1990 Aug;93(4):1347–1353. doi: 10.1104/pp.93.4.1347. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Tietjen K. G., Hunkler D., Matern U. Differential response of cultured parsley cells to elicitors from two non-pathogenic strains of fungi. 1. Identification of induced products as coumarin derivatives. Eur J Biochem. 1983 Mar 15;131(2):401–407. doi: 10.1111/j.1432-1033.1983.tb07277.x. [DOI] [PubMed] [Google Scholar]
  44. Töpfer R., Schell J., Steinbiss H. H. Versatile cloning vectors for transient gene expression and direct gene transfer in plant cells. Nucleic Acids Res. 1988 Sep 12;16(17):8725–8725. doi: 10.1093/nar/16.17.8725. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. Wingender R., Röhrig H., Höricke C., Wing D., Schell J. Differential regulation of soybean chalcone synthase genes in plant defence, symbiosis and upon environmental stimuli. Mol Gen Genet. 1989 Aug;218(2):315–322. doi: 10.1007/BF00331284. [DOI] [PubMed] [Google Scholar]
  46. Yang J. Q., Remmers E. F., Marcu K. B. The first exon of the c-myc proto-oncogene contains a novel positive control element. EMBO J. 1986 Dec 20;5(13):3553–3562. doi: 10.1002/j.1460-2075.1986.tb04682.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  47. van de Löcht U., Meier I., Hahlbrock K., Somssich I. E. A 125 bp promoter fragment is sufficient for strong elicitor-mediated gene activation in parsley. EMBO J. 1990 Sep;9(9):2945–2950. doi: 10.1002/j.1460-2075.1990.tb07486.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  48. van der Meer I. M., Spelt C. E., Mol J. N., Stuitje A. R. Promoter analysis of the chalcone synthase (chsA) gene of Petunia hybrida: a 67 bp promoter region directs flower-specific expression. Plant Mol Biol. 1990 Jul;15(1):95–109. doi: 10.1007/BF00017727. [DOI] [PubMed] [Google Scholar]

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