The major objective of our work is to understand the regulation of indole alkaloid biosynthesis in Camptotheca acuminata, a chinese tree that produces the anti-cancer alkaloid camptothecin (CPT). In addition to its basic scientific interest, elucidation of the regulatory mechanisms of the indole alkaloid pathway may suggest biotechnology strategies to increase the amount or alter the spectrum of indole alkaloids produced by this plant and other species. Because complex pathways often are regulated at the early stages, our approach to studying the regulation of this pathway is to examine the effects of over-expression of key enzymes on other components of indole alkaloid biosynthesis. We have over-expressed the first two enzymes of the CPT pathway, tryptophan decarboxylase (TDC) and strictosidine synthase (SS), In transgenic tobacco plants as a model system. Both proteins are highly expressed in enzymatically-active forms. We have also devised a assays for two other steps in the CPT pathway, the formation of strictosamide from strictosidine and the hydroxylation of CPT at the C-10 position. These enzymes and their genes will be isolated so their effects on alkaloid metabolism can be analyzed as well. In addition to regulation by its own metabolism, the CPT pathway may be induced by external factors, such as biotic and abiotic stress, light regimes, or developmental cues. We will analyze expression of reporter genes from the transcriptional promoters of both TDC and SS under a variety of potentially inductive conditions. Factors that regulate both genes may be general inducers of the pathway.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
5R01CA057592-02
Application #
2098312
Study Section
Bio-Organic and Natural Products Chemistry Study Section (BNP)
Project Start
1992-08-01
Project End
1995-07-31
Budget Start
1993-08-01
Budget End
1995-07-31
Support Year
2
Fiscal Year
1993
Total Cost
Indirect Cost
Name
Texas A&M University
Department
Biology
Type
Schools of Earth Sciences/Natur
DUNS #
City
College Station
State
TX
Country
United States
Zip Code
77845
Lu, Hua; Gorman, Elizabeth; McKnight, Thomas D (2005) Molecular characterization of two anthranilate synthase alpha subunit genes in Camptotheca acuminata. Planta 221:352-60
Lorence, Argelia; Nessler, Craig L (2004) Camptothecin, over four decades of surprising findings. Phytochemistry 65:2735-49
Lorence, A; Medina-Bolivar, F; Nessler, C L (2004) Camptothecin and 10-hydroxycamptothecin from Camptotheca acuminata hairy roots. Plant Cell Rep 22:437-41
Vincent, R M; Lopez-Meyer, M; McKnight, T D et al. (1997) Sustained harvest of camptothecin from the leaves of Camptotheca acuminata. J Nat Prod 60:618-9
Lopez-Meyer, M; Nessler, C L (1997) Tryptophan decarboxylase is encoded by two autonomously regulated genes in Camptotheca acuminata which are differentially expressed during development and stress. Plant J 11:1167-75
Maldonado-Mendoza, I E; Nessler, C L (1997) Molecular characterization of the AP19 gene family in Arabidopsis thaliana: components of the Golgi AP-1 clathrin assembly protein complex. Plant Mol Biol 35:865-72
Maldonado-Mendoza, I E; Vincent, R M; Nessler, C L (1997) Molecular characterization of three differentially expressed members of the Camptotheca acuminata 3-hydroxy-3-methylglutaryl CoA reductase (HMGR) gene family. Plant Mol Biol 34:781-90
Maldonado-Mendoza, I E; Lopez-Meyer, M; Galef, J R et al. (1996) Molecular analysis of a new member of the opium poppy tyrosine/3,4-dihydroxyphenylalanine decarboxylase gene family. Plant Physiol 110:43-9
Lopez-Meyer, M; Nessler, C L; McKnight, T D (1994) Sites of accumulation of the antitumor alkaloid camptothecin in Camptotheca acuminata. Planta Med 60:558-60
Nessler, C L (1994) Metabolic engineering of plant secondary products. Transgenic Res 3:109-15

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