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. 1997 Mar;113(3):755–763. doi: 10.1104/pp.113.3.755

Isolation of a cDNA and a genomic clone encoding cinnamate 4-hydroxylase from Arabidopsis and its expression manner in planta.

M Mizutani 1, D Ohta 1, R Sato 1
PMCID: PMC158193  PMID: 9085571

Abstract

We have isolated a cDNA for a cytochrome P450, cinnamate 4-hydroxylase (C4H), of Arabidopsis thaliana using a C4H cDNA from mung been as a hybridization probe. The deduced amino acid sequence is 84.7% identical to that of mung bean C4H and therefore was designated CYP73A5. The CYP73A5 protein was expressed in insect cells using the baculovirus expression system and when reconstituted with lipid and NADPH-cytochrome P450 reductase resulted in C4H activity with a specific activity of 68 nmol min-1 nmol-1 P450. Southern blot analysis revealed that CYP73A5 is a single-copy gene in Arabidopsis. C4H (CYP73A5) expression was apparently coordinated in Arabidopsis with both PAL1 and 4CL in response to light and wounding. Although the light induction of CHS followed a time course similar to that observed with C4H, no induction of CHS was detected upon wounding. On the other hand, the C4H expression patterns exhibited no significant coordination with those of PAL2 and PAL3. A C4H promoter region of 907 bp contained all of the three cis-acting elements (boxes P, A, and L) conserved among the PAL and 4CL genes so far reported as controlling expression.

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Selected References

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  1. Appert C., Logemann E., Hahlbrock K., Schmid J., Amrhein N. Structural and catalytic properties of the four phenylalanine ammonia-lyase isoenzymes from parsley (Petroselinum crispum Nym.). Eur J Biochem. 1994 Oct 1;225(1):491–499. doi: 10.1111/j.1432-1033.1994.00491.x. [DOI] [PubMed] [Google Scholar]
  2. 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]
  3. Dixon R. A., Paiva N. L. Stress-Induced Phenylpropanoid Metabolism. Plant Cell. 1995 Jul;7(7):1085–1097. doi: 10.1105/tpc.7.7.1085. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. 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]
  5. Fahrendorf T., Dixon R. A. Stress responses in alfalfa (Medicago sativa L.). XVIII: Molecular cloning and expression of the elicitor-inducible cinnamic acid 4-hydroxylase cytochrome P450. Arch Biochem Biophys. 1993 Sep;305(2):509–515. doi: 10.1006/abbi.1993.1454. [DOI] [PubMed] [Google Scholar]
  6. Feinbaum R. L., Ausubel F. M. Transcriptional regulation of the Arabidopsis thaliana chalcone synthase gene. Mol Cell Biol. 1988 May;8(5):1985–1992. doi: 10.1128/mcb.8.5.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Hamerski D., Schmitt D., Matern U. Induction of two prenyltransferases for the accumulation of coumarin phytoalexins in elicitor-treated Ammi majus cell suspension cultures. Phytochemistry. 1990;29(4):1131–1135. doi: 10.1016/0031-9422(90)85417-e. [DOI] [PubMed] [Google Scholar]
  8. Hauffe K. D., Lee S. P., Subramaniam R., Douglas C. J. Combinatorial interactions between positive and negative cis-acting elements control spatial patterns of 4CL-1 expression in transgenic tobacco. Plant J. 1993 Aug;4(2):235–253. doi: 10.1046/j.1365-313x.1993.04020235.x. [DOI] [PubMed] [Google Scholar]
  9. Johnson E. F. Mapping determinants of the substrate selectivities of P450 enzymes by site-directed mutagenesis. Trends Pharmacol Sci. 1992 Mar;13(3):122–126. doi: 10.1016/0165-6147(92)90042-5. [DOI] [PubMed] [Google Scholar]
  10. Lagrimini L. M., Burkhart W., Moyer M., Rothstein S. Molecular cloning of complementary DNA encoding the lignin-forming peroxidase from tobacco: Molecular analysis and tissue-specific expression. Proc Natl Acad Sci U S A. 1987 Nov;84(21):7542–7546. doi: 10.1073/pnas.84.21.7542. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. 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]
  12. Lee D., Ellard M., Wanner L. A., Davis K. R., Douglas C. J. The Arabidopsis thaliana 4-coumarate:CoA ligase (4CL) gene: stress and developmentally regulated expression and nucleotide sequence of its cDNA. Plant Mol Biol. 1995 Aug;28(5):871–884. doi: 10.1007/BF00042072. [DOI] [PubMed] [Google Scholar]
  13. Lindberg R. L., Negishi M. Alteration of mouse cytochrome P450coh substrate specificity by mutation of a single amino-acid residue. Nature. 1989 Jun 22;339(6226):632–634. doi: 10.1038/339632a0. [DOI] [PubMed] [Google Scholar]
  14. 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]
  15. Lois R., Hahlbrock K. Differential wound activation of members of the phenylalanine ammonia-lyase and 4-coumarate:CoA ligase gene families in various organs of parsley plants. Z Naturforsch C. 1992 Jan-Feb;47(1-2):90–94. doi: 10.1515/znc-1992-1-216. [DOI] [PubMed] [Google Scholar]
  16. Mauch-Mani B., Slusarenko A. J. Production of Salicylic Acid Precursors Is a Major Function of Phenylalanine Ammonia-Lyase in the Resistance of Arabidopsis to Peronospora parasitica. Plant Cell. 1996 Feb;8(2):203–212. doi: 10.1105/tpc.8.2.203. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Nairn C. J., Winesett L., Ferl R. J. Nucleotide sequence of an actin gene from Arabidopsis thaliana. Gene. 1988 May 30;65(2):247–257. doi: 10.1016/0378-1119(88)90461-1. [DOI] [PubMed] [Google Scholar]
  18. Nelson D. R., Strobel H. W. On the membrane topology of vertebrate cytochrome P-450 proteins. J Biol Chem. 1988 May 5;263(13):6038–6050. [PubMed] [Google Scholar]
  19. OMURA T., SATO R. THE CARBON MONOXIDE-BINDING PIGMENT OF LIVER MICROSOMES. I. EVIDENCE FOR ITS HEMOPROTEIN NATURE. J Biol Chem. 1964 Jul;239:2370–2378. [PubMed] [Google Scholar]
  20. Ohl S., Hedrick S. A., Chory J., Lamb C. J. Functional properties of a phenylalanine ammonia-lyase promoter from Arabidopsis. Plant Cell. 1990 Sep;2(9):837–848. doi: 10.1105/tpc.2.9.837. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Potts J. R., Weklych R., Conn E. E., Rowell J. The 4-hydroxylation of cinnamic acid by sorghum microsomes and the requirement for cytochrome P-450. J Biol Chem. 1974 Aug 25;249(16):5019–5026. [PubMed] [Google Scholar]
  22. Russell D. W. The metabolism of aromatic compounds in higer plants. X. Properties of the cinnamic acid 4-hydroxylase of pea seedlings and some aspects of its metabolic and developmental control. J Biol Chem. 1971 Jun 25;246(12):3870–3878. [PubMed] [Google Scholar]
  23. Teutsch H. G., Hasenfratz M. P., Lesot A., Stoltz C., Garnier J. M., Jeltsch J. M., Durst F., Werck-Reichhart D. Isolation and sequence of a cDNA encoding the Jerusalem artichoke cinnamate 4-hydroxylase, a major plant cytochrome P450 involved in the general phenylpropanoid pathway. Proc Natl Acad Sci U S A. 1993 May 1;90(9):4102–4106. doi: 10.1073/pnas.90.9.4102. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Yalpani N., Leon J., Lawton M. A., Raskin I. Pathway of Salicylic Acid Biosynthesis in Healthy and Virus-Inoculated Tobacco. Plant Physiol. 1993 Oct;103(2):315–321. doi: 10.1104/pp.103.2.315. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Yamazaki S., Sato K., Suhara K., Sakaguchi M., Mihara K., Omura T. Importance of the proline-rich region following signal-anchor sequence in the formation of correct conformation of microsomal cytochrome P-450s. J Biochem. 1993 Nov;114(5):652–657. doi: 10.1093/oxfordjournals.jbchem.a124232. [DOI] [PubMed] [Google Scholar]

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