Phylogenetic analysis and functional characterisation of strictosidine synthase-like genes in Arabidopsis thaliana
Natalie A. J. Kibble A B , M. Mehdi Sohani A , Neil Shirley A B , Caitlin Byrt A B , Ute Roessner C , Antony Bacic C , Otto Schmidt A and Carolyn J. Schultz A DA School of Agriculture Food and Wine, The University of Adelaide, Glen Osmond, SA 5064, Australia.
B Australian Centre for Plant Functional Genomics, The University of Adelaide, Glen Osmond, SA 5064, Australia.
C School of Botany, The University of Melbourne, Parkville, Vic. 3010, Australia.
D Corresponding author. Email: carolyn.schultz@adelaide.edu.au
Functional Plant Biology 36(12) 1098-1109 https://doi.org/10.1071/FP09104
Submitted: 8 May 2009 Accepted: 8 September 2009 Published: 3 December 2009
Abstract
Monoterpenoid indole alkaloids (MIA) are a diverse class of secondary metabolites important for plant protection and are drugs for treating human diseases. Arabidopsis thaliana (L.) is not known to produce MIAs, yet its genome has 15 genes with similarity to the periwinkle (Catharanthus roseus (L.) G. Don) strictosidine synthase (STR) gene. Phylogenetic analysis of strictosidine synthase-like (SSL) proteins reveals four well supported classes of SSLs in Arabidopsis. To determine if Arabidopsis produces active strictosidine synthase, Arabidopsis protein extracts were assayed for enzymatic activity and cDNAs were expressed in Escherichia coli. Arabidopsis protein extracts from leaves and hairy roots do not make strictosidine at levels comparable to C. roseus, but they metabolise one substrate, secologanin, a precursor of strictosidine in other plant species, and produce an ‘unknown’ compound proposed to be a dimer of secologanic acid. Recombinant Arabidopsis proteins expressed in E. coli were not active STRs. Quantitative PCR analysis was performed on class A Ssls and showed they are upregulated by salt, ultraviolet light and salicylic acid treatment. RNAi mutants of Arabidopsis with reduced expression of all four class A Ssls, suggest that class A SSL proteins can modify secologanin. Gene expression and metabolomics data suggests that class A Ssl genes may have a role in plant protection.
Additional keywords: gene duplication, monoterpenoid indole alkaloid, secologanin.
Acknowledgements
This project was supported by the Australian Centre for Plant Functional Genomics, funded by the Australian Research Council and the Grains Research and Development Corporation. The authors thank Kris Ford (ESI–MS/MS), Robert Asenstorfer, Max Tate (MS interpretation), Jelle Lahnstein (RP-HPLC) and Mark Livermore (E. coli expression) for their technical assistance, Ute Baumann for helpful discussions on phylogenetic analysis, and Robert Verpoorte and Magdi El-Sayed for their kind donation of the strictosidine standard.
Arase S,
Ueno M,
Toko M,
Honda Y,
Itoh K, Ozoe Y
(2001) Light-dependent accumulation of tryptamine in the rice Sekiguchi lesion mutant infected with Magnaporthe grisea. Journal of Phytopathology 149, 409–413.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
Baldauf SL
(2003) Phylogeny for the faint of heart: a tutorial. Trends in Genetics 19, 345–351.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
PubMed |
Burton RA,
Shirley NJ,
King BJ,
Harvey AJ, Fincher GB
(2004) The CesA gene family of barley. Quantitative analysis of transcripts reveals two groups of co-expressed genes. Plant Physiology 134, 224–236.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
PubMed |
Clough SJ, Bent AF
(1998) Floral dip: a simplified method for Agrobacterium-mediated transformation of Arabidopsis thaliana. The Plant Journal 16, 735–743.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
PubMed |
De Luca V
(2003) Biochemistry and molecular biology of indole alkaloid biosynthesis: the implication of recent discoveries. Recent Advances in Phytochemistry 37, 181–202.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
Dean JV,
Mohammed LA, Fitzpatrick T
(2005) The formation, vacuolar localization, and tonoplast transport of salicylic acid glucose conjugates in tobacco cell suspension cultures. Planta 221, 287–296.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
PubMed |
Edgar RC
(2004) MUSCLE: multiple sequence alignment with high accuracy and high throughput. Nucleic Acids Research 32, 1792–1797.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
PubMed |
Fabbri M,
Delp G,
Schmidt O, Theopold U
(2000) Animal and plant members of a gene family with similarity to alkaloid-synthesizing enzymes. Biochemical and Biophysical Research Communications 271, 191–196.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
PubMed |
Facchini PJ, De Luca V
(2008) Opium poppy and Madagascar periwinkle: model non-model systems to investigate alkaloid biosynthesis in plants. The Plant Journal 54, 763–784.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
PubMed |
Facchini PJ,
Bird DA, St-Pierre B
(2004) Can Arabidopsis make complex alkaloids? Trends in Plant Science 9, 116–122.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
PubMed |
Gerasimenko I,
Sheludko Y,
Unger M, Stockigt J
(2001) Development of an efficient system for the separation of indole alkaloids by high performance liquid chromatography and its applications. Phytochemical Analysis 12, 96–103.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
PubMed |
Imaoka A,
Ueno M,
Kihara J,
Kadowaki M, Arase S
(2008) Effect of a photo-synthetic inhibitor on tryptamine pathway-mediated sekiguchi lesion formation in lesion mimic mutant of rice infected with Magnaporthe grisea. Journal of Phytopathology 156, 522–529.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
Jacobs A,
Lunde C,
Bacic A,
Tester M, Roessner U
(2007) The impact of constitutive heterologous expression of a moss Na+ transporter on the metabolomes of rice and barley. Metabolomics 3, 307–317.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
Kopka J
(2006) Current challenges and developments in GC–MS based metabolite profiling technology. Journal of Biotechnology 124, 312–322.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
PubMed |
Kutchan TM
(1989) Expression of enzymatically active cloned strictosidine synthase from the higher plant Rauvolfia serpentina in Escherichia coli. FEBS Letters 257, 127–130.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
PubMed |
Kutchan TM,
Hampp N,
Lottspeich F,
Beyreuther K, Zenk MH
(1988) The cDNA clone for strictosidine synthase from Rauvolfia serpentina. FEBS Letters 237, 40–44.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
PubMed |
Leister D
(2004) Tandem and segmental gene duplication and recombination in the evolution of plant disease resistance genes. Trends in Genetics 20, 116–122.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
PubMed |
Ma X,
Panjikar S,
Koepke J,
Loris E, Stockigt J
(2006) The structure of Rauvolfia serpentina strictosidine synthase is a novel six-bladed β-propeller fold in plant proteins. The Plant Cell 18, 907–920.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
PubMed |
McCoy E,
Galan MC, O’Connor SE
(2006) Substrate specificity of strictosidine synthase. Bioorganic & Medicinal Chemistry Letters 16, 2475–2478.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
PubMed |
Munns R
(2002) Comparative physiology of salt and water stress. Plant, Cell & Environment 25, 239–250.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
PubMed |
Ober D
(2005) Seeing double: gene duplication and diversification in plant secondary metabolism. Trends in Plant Science 10, 444–449.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
PubMed |
Osbourn AE,
Qi X,
Townsend B, Qin B
(2003) Dissecting plant secondary metabolism – constitutive chemical defences in cereals. New Phytologist 159, 101–108.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
Ouwerkerk PBF,
Hallard D,
Verpoorte R, Memelink J
(1999) Identification of UV-B light responsive regions in the promoter of the tryptophan decarboxylase gene from Catharanthus roseus. Plant Molecular Biology 41, 491–503.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
PubMed |
Page RDM
(1996) TreeView: an application to display phylogenetic trees on personal computers. Computer Applications in the Biosciences 12, 357–358.
|
CAS |
PubMed |
Pennings EJM,
van den Bosch R,
van der Heijden R,
Stevens LH,
Duine JA, Verpoorte R
(1989) Assay of strictosidine synthase from plant cell cultures by high-performance liquid chromatography. Analytical Biochemistry 176, 412–415.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
PubMed |
Roessner U,
Patterson JH,
Forbes MG,
Fincher GB,
Langridge P, Bacic A
(2006) An investigation of boron toxicity in barley using metabolomics. Plant Physiology 142, 1087–1101.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
PubMed |
Schenk PM,
Kazan K,
Wilson I,
Anderson JP,
Richmond T,
Somerville SC, Manners JM
(2000) Coordinated plant defense responses in Arabidopsis revealed by microarray analysis. Proceedings of the National Academy of Sciences of the United States of America 97, 11 655–11 660.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
PubMed |
Sohani MM,
Schenk PM,
Schultz CJ, Schmidt O
(2009) Phylogenetic and transcriptional analysis of a strictosidine synthase-like gene family in Arabidopsis thaliana reveals involvement in plant defence responses. Plant Biology 11, 105–117.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
PubMed |
Stevens LH,
Giroud C,
Pennings EJM, Verpoorte R
(1993) Purification and characterization of strictosidine synthase from a suspension culture of Cinchona robusta. Phytochemistry 33, 99–106.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
Sumner LW,
Mendes P, Dixon RA
(2003) Plant metabolomics: large-scale phytochemistry in the functional genomics era. Phytochemistry 62, 817–836.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
PubMed |
Tester M, Davenport R
(2003) Na+ tolerance and Na+ transport in higher plants. Annals of Botany 91, 503–527.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
PubMed |
The Arabidopsis Genome Initiative
(2000) Analysis of the genome sequence of the flowering plant Arabidopsis thaliana. Nature 408, 796–815.
| Crossref | GoogleScholarGoogle Scholar | PubMed |
Treimer JF, Zenk MH
(1979) Purification and properties of strictosidine synthase, the key enzyme in indole alkaloid formation. European Journal of Biochemistry 101, 225–233.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
PubMed |
Ulm R,
Baumann A,
Oravecz A,
Mate Z,
Adam E,
Oakeley EJ,
Schafer E, Nagy F
(2004) Genome-wide analysis of gene expression reveals function of the bZIP transcription factor HY5 in the UV-B response of Arabidopsis. Proceedings of the National Academy of Sciences of the United States of America 101, 1397–1402.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
PubMed |
Wesley SV,
Helliwell CA,
Smith NA,
Wang M, Rouse DT , et al.
(2001) Construct design for efficient, effective and high-throughput gene silencing in plants. The Plant Journal 27, 581–590.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
PubMed |
Yamazaki Y,
Sudo H,
Yamazaki M,
Aimi N, Saito K
(2003a) Camptothecin biosynthetic genes in hairy roots of Ophiorrhiza pumila: cloning, characterization and differential expression in tissues and by stress compounds. Plant & Cell Physiology 44, 395–403.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
PubMed |
Yamazaki Y,
Urano A,
Sudo H,
Kitajima M,
Takayama H,
Yamazaki M,
Aimi N, Saito K
(2003b) Metabolite profiling of alkaloids and strictosidine synthase activity in camptothecin producing plants. Phytochemistry 62, 461–470.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
PubMed |
Zimmermann P,
Hennig L, Gruissem W
(2005) Gene-expression analysis and network discovery using Genevestigator. Trends in Plant Science 10, 407–409.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
PubMed |
