Abstract
An aromatic amino acid decarboxylase DNA fragment was generated from opium poppy (Papaver somniferum L.) genomic DNA by the PCR using primers designed from conserved amino acid sequences of other aromatic amino acid decarboxylase genes. Using this fragment as a probe, a genomic clone was isolated that encodes a new member of the opium poppy tyrosine/3,4-dihydroxyphenylalanine decarboxylase gene family (TyDC5). The predicted TyDC5 amino acid sequence shares extensive identity with other opium poppy tyrosine/3,4-dihydroxyphenylalanine decarboxylases (84%), and when expressed in Escherichia coli, it is active against tyrosine and to a lesser extent against 3,4-dihydroxyphenylalanine. Ribonuclease protection assays indicate that TyDC5 is expressed primarily in the roots of mature poppy plants. A peak of TyDC5 expression was also observed during germination, coincident with the emergence of the radicle from the seed coat. Parallel results were obtained in transgenic tobacco using a TyDC5 promoter fragment (-2060) translationally fused to the beta-glucuronidase reporter gene (GUS). IN TyDC5::GUS tobacco, GUS activity transiently appeared in all parts of the seedling during germination, but was limited to the roots in older plants. These results indicate that TyDC5 expression is transcriptionally regulated and suggest that the TyDC5 enzyme may play an important role in providing precursors for alkaloid synthesis in the roots and germinating seedlings of opium poppy.
Full Text
The Full Text of this article is available as a PDF (1,009.4 KB).
Selected References
These references are in PubMed. This may not be the complete list of references from this article.
- 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]
- Asghar S. S., Siddiqi M. Phenolase of papaver somniferum. I. Isolation of the enzyme and its substrate specificity. Enzymologia. 1970;39(5):289–306. [PubMed] [Google Scholar]
- 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]
- Bradford M. M. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem. 1976 May 7;72:248–254. doi: 10.1006/abio.1976.9999. [DOI] [PubMed] [Google Scholar]
- De Luca V., Fernandez J. A., Campbell D., Kurz W. G. Developmental Regulation of Enzymes of Indole Alkaloid Biosynthesis in Catharanthus roseus. Plant Physiol. 1988 Feb;86(2):447–450. doi: 10.1104/pp.86.2.447. [DOI] [PMC free article] [PubMed] [Google Scholar]
- De Luca V., Marineau C., Brisson N. Molecular cloning and analysis of cDNA encoding a plant tryptophan decarboxylase: comparison with animal dopa decarboxylases. Proc Natl Acad Sci U S A. 1989 Apr;86(8):2582–2586. doi: 10.1073/pnas.86.8.2582. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Facchini P. J., De Luca V. Differential and tissue-specific expression of a gene family for tyrosine/dopa decarboxylase in opium poppy. J Biol Chem. 1994 Oct 28;269(43):26684–26690. [PubMed] [Google Scholar]
- Facchini P. J., De Luca V. Expression in Escherichia coli and partial characterization of two tyrosine/dopa decarboxylases from opium poppy. Phytochemistry. 1995 Mar;38(5):1119–1126. doi: 10.1016/0031-9422(94)00814-a. [DOI] [PubMed] [Google Scholar]
- Hochuli E., Döbeli H., Schacher A. New metal chelate adsorbent selective for proteins and peptides containing neighbouring histidine residues. J Chromatogr. 1987 Dec 18;411:177–184. doi: 10.1016/s0021-9673(00)93969-4. [DOI] [PubMed] [Google Scholar]
- 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]
- Kawalleck P., Keller H., Hahlbrock K., Scheel D., Somssich I. E. A pathogen-responsive gene of parsley encodes tyrosine decarboxylase. J Biol Chem. 1993 Jan 25;268(3):2189–2194. [PubMed] [Google Scholar]
- Koltunow A. M., Truettner J., Cox K. H., Wallroth M., Goldberg R. B. Different Temporal and Spatial Gene Expression Patterns Occur during Anther Development. Plant Cell. 1990 Dec;2(12):1201–1224. doi: 10.1105/tpc.2.12.1201. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Nessler C. L., Kurz W. G., Pelcher L. E. Isolation and analysis of the major latex protein genes of opium poppy. Plant Mol Biol. 1990 Dec;15(6):951–953. doi: 10.1007/BF00039436. [DOI] [PubMed] [Google Scholar]
- Roberts M. F., McCarthy D., Kutchan T. M., Coscia C. J. Localization of enzymes and alkaloidal metabolites in Papaver latex. Arch Biochem Biophys. 1983 Apr 15;222(2):599–609. doi: 10.1016/0003-9861(83)90558-1. [DOI] [PubMed] [Google Scholar]
- Sanger F., Nicklen S., Coulson A. R. DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5463–5467. doi: 10.1073/pnas.74.12.5463. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Trezzini G. F., Horrichs A., Somssich I. E. Isolation of putative defense-related genes from Arabidopsis thaliana and expression in fungal elicitor-treated cells. Plant Mol Biol. 1993 Jan;21(2):385–389. doi: 10.1007/BF00019954. [DOI] [PubMed] [Google Scholar]