Skip to main content
SpringerLink
Log in

Unraveling new genes associated with seed development and metabolism in Bixa orellana L. by expressed sequence tag (EST) analysis

Molecular Biology Reports Aims and scope Submit manuscript

Abstract

The tropical tree Bixa orellana L. produces a range of secondary metabolites which biochemical and molecular biosynthesis basis are not well understood. In this work we have characterized a set of ESTs from a non-normalized cDNA library of B. orellana seeds to obtain information about the main developmental and metabolic processes taking place in developing seeds and their associated genes. After sequencing a set of randomly selected clones, most of the sequences were assigned with putative functions based on similarity, GO annotations and protein domains. The most abundant transcripts encoded proteins associated with cell wall (prolyl 4-hydroxylase), fatty acid (acyl carrier protein), and hormone/flavonoid (2OG-Fe oxygenase) synthesis, germination (MADS FLC-like protein) and embryo development (AP2/ERF transcription factor) regulation, photosynthesis (chlorophyll a–b binding protein), cell elongation (MAP65-1a), and stress responses (metallothionein- and thaumatin-like proteins). Enzymes were assigned to 16 different metabolic pathways related to both primary and secondary metabolisms. Characterization of two candidate genes of the bixin biosynthetic pathway, BoCCD and BoOMT, showed that they belong, respectively, to the carotenoid-cleavage dioxygenase 4 (CCD4) and caffeic acid O-methyltransferase (COMT) families, and are up-regulated during seed development. It indicates their involvement in the synthesis of this commercially important carotenoid pigment in seeds of B. orellana. Most of the genes identified here are the first representatives of their gene families in B. orellana.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price includes VAT (France)

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4

References

  1. Lawrence BM, Hogg JW (1973) Ishwarane in Bixa orellana leaf oil. Phytochemistry 12:2995

    Article  CAS  Google Scholar 

  2. Harbone JB (1975) Flavonoid bisulfates and their co-occurrences with ellagic acid in the Bixaceae, Frankeniaceae, and related families. Phytochemistry 14:1331–1337

    Article  Google Scholar 

  3. Jondiko IJO, Pattenden G (1989) Terpenoids and an apocarotenoid from seeds of Bixa orellana. Phytochemistry 28:3159–3162

    Article  CAS  Google Scholar 

  4. Frega N, Mozzon M, Bocci F (1998) Identification and estimation of tocotrienols in the annatto lipid fraction by gas chromatography mass spectrometry. J Am Oil Chem Soc 75:1723–1727

    Article  CAS  Google Scholar 

  5. Pino JA, Correa MT (2003) Chemical composition of the essential oil from annatto (Bixa orellana L.) seeds. J Essent Oil Res 15:66–67

    CAS  Google Scholar 

  6. Shilpi JA, Taufiq-Ur-Rahman Md, Uddin SJ, Alam Md S, Sadhu SK, Seidel V (2006) Preliminary pharmacological screening of Bixa orellana L. leaves. J Ethnopharmacol 108:264–271

    Article  PubMed  Google Scholar 

  7. Giuliano G, Al-Babili S, Von Lintig J (2003) Carotenoid oxygenases: cleave it or leave it. Trends Plant Sci 8:145–149

    Article  CAS  PubMed  Google Scholar 

  8. Mercadante AZ, Pfander H (1998) Carotenoids from annatto: a review. Recent Res Devel Agric Food Chem 2:79–91

    Google Scholar 

  9. Lu C, Wallis JG, Browse J (2007) An analysis of expressed sequence tags of developing castor endosperm using a full-length cDNA library. BMC Plant Biol 7:42

    Article  PubMed  Google Scholar 

  10. Crowhurst RN, Gleave AP, MacRae EA et al (2008) Analysis of expressed sequence tags from Actinidia: applications of a cross species EST database for gene discovery in the areas of flavor, health, color and ripening. BMC Genomics 9:351

    Article  PubMed  Google Scholar 

  11. Yoshida K, Nishiguchi M, Futamura N, Nanjo T (2007) Expressed sequence tags from Cryptomeria japonica sapwood during the drying process. Tree Physiol 27:1–9

    CAS  PubMed  Google Scholar 

  12. Hocher V, Auguy F, Argout X, Laplaze L, Franche C, Bogusz D (2006) Expressed sequence-tag analysis in Casuarina glauca actinorhizal nodule and root. New Phytol 169:681–688

    Article  PubMed  Google Scholar 

  13. Jako C, Coutu C, Roewer I, Reed DW, Pelcher LE, Covello PS (2002) Probing carotenoid biosynthesis in developing seed coats of Bixa orellana (Bixaceae) through expressed sequence tag analysis. Plant Sci 163:141–145

    Article  CAS  Google Scholar 

  14. Rodrigues SM, Soares VLF, Oliveira TM, Gesteira AS, Otoni WC, Costa MGC (2007) Isolation and purification of RNA from tissues rich in polyphenols, polysaccharides, and pigments of annato (Bixa orellana L.). Mol Biotechnol 37:220–224

    Article  CAS  PubMed  Google Scholar 

  15. Sambrook J, Fritsch EF, Maniatis T (1989) Molecular cloning: a laboratory manual, 2nd edn. Cold Spring Harbor Laboratory Press, Plainview, New York

    Google Scholar 

  16. Conesa A, Götz S, García-Gomez JM, Terol J, Talón M, Robles M (2005) Blast2GO: a universal tool for annotation, visualization and analysis in functional genomics research. Bioinformatics 21:3674–3676

    Article  CAS  PubMed  Google Scholar 

  17. Altschul SF, Gish W, Miller W, Myers EW, Lipman DJ (1990) Basic local alignment search tool. J Mol Biol 215:403–410

    CAS  PubMed  Google Scholar 

  18. Labarga A, Valentin F, Anderson M, Lopez R (2007) Web services at the European Bioinformatics Institute. Nucleic Acids Res 35:w6–w11

    Article  PubMed  Google Scholar 

  19. Thompson JD, Higgins DG, Gibson TJ (1994) CLUSTAL W: improving the sensitivity of progressive multiple sequence alignment through sequence weighting, position-specific gap penalties and weigh matrix choice. Nucleic Acids Res 22:4673–4680

    Article  CAS  PubMed  Google Scholar 

  20. Saitou N, Nei M (1987) The neighbor-joining method: a new method for reconstructing phylogenetic trees. Mol Biol Evol 4:406–425

    CAS  PubMed  Google Scholar 

  21. Livak KJ, Schmittgen TD (2001) Analysis of relative gene expression data using real-time quantitative PCR and the 2−ΔΔCt method. Methods 25:402–408

    Article  CAS  PubMed  Google Scholar 

  22. Cruz AC, Costa MGC, Otoni WC (2008) Aspectos gerais da cultura e beneficiamento da semente de urucum. In: Stringheta PC, Silva PI (eds) Pigmentos de urucum: extração, reações químicas, usos e aplicações. Suprema, Viçosa, pp 11–23

    Google Scholar 

  23. Ma J, Morrow D, Fernandes J, Walbot V (2006) Comparative profiling of the sense and antisense transcriptome of maize lines. Genome Biol 7:R22

    Article  PubMed  Google Scholar 

  24. Terol J, Conesa A, Colmenero JM et al (2007) Analysis of 13000 unique Citrus clusters associated with fruit quality, production and salinity tolerance. BMC Genomics 8:31

    Article  PubMed  Google Scholar 

  25. Horn R, Lecouls AC, Callahan A et al (2005) Candidate gene database and transcript map for peach, a model species for fruit trees. Theor Appl Genet 110:1419–1428

    Article  PubMed  Google Scholar 

  26. Vecchietti A, Lazzari B, Ortugno C, Bianchi F, Malinverni R, Caprera A, Mignani I, Pozzi C (2009) Comparative analysis of expressed sequence tags from tissues in ripening stages of peach (Prunus persica L. Batsch). Tree Genet Genome 5:377–391

    Article  Google Scholar 

  27. Keskiaho K, Hieta R, Sormunen R, Myllyharju J (2007) Chlamydomonas reinhardtii has multiple prolyl 4-hydroxylases, one of which is essential for proper cell wall assembly. Plant Cell 19:256–269

    Article  CAS  PubMed  Google Scholar 

  28. Koski MK, Hieta R, Hirsilã M, Könka A, Myllyharju J, Wierenga RK (2009) The crystal structure of an algal prolyl 4-hydroxylase complexed with a proline-rich peptide reveals a novel buried tripeptide binding motif. J Biol Chem 2484:25290–25301

    Article  Google Scholar 

  29. Asif MH, Trivedi PK, Misra P, Nath P (2009) Prolyl-4-hydroxylase (5AtP4H1) mediates and mimics low oxygen response in Arabidopsis thaliana. Funct Integr Genomics 9:525–535

    Article  CAS  PubMed  Google Scholar 

  30. Suh MC, Schultz DJ, Ohlrogge JB (1999) Isoforms of acyl carrier protein involved in seed-specific fatty acid synthesis. Plant J 17:679–688

    Article  CAS  PubMed  Google Scholar 

  31. Buchanan BB, Gruissem W, Jones RL (2002) Biochemistry and molecular biology of plants. American Society of Plant Physiologists, Rockville

    Google Scholar 

  32. Chiang GCK, Barua D, Kramera EM, Amasino RM, Donohue K (2009) Major flowering time gene, FLOWERING LOCUS C, regulates seed germination in Arabidopsis thaliana. Proc Natl Acad Sci USA 106:11661–11666

    Article  CAS  PubMed  Google Scholar 

  33. Aravind L, Koonin E (2001) The DNA-repair protein AlkB, EGL-9, and leprecan define new families of 2-oxoglutarate- and iron-dependent dioxygenases. Genome Biol 2:1–8

    Article  Google Scholar 

  34. Lukacin R, Britsch L (1997) Identification of strictly conserved histidine and arginine residues as part of the active site in Petunia hybrida flavanone 3b-hydroxylase. Eur J Biochem 249:748–757

    Article  CAS  PubMed  Google Scholar 

  35. Hedden P, Phillips AL (2000) Gibberellin metabolism: new insights revealed by the genes. Trends Plant Sci 5:523–530

    Article  CAS  PubMed  Google Scholar 

  36. Wang KL, Li H, Ecker JR (2002) Ethylene biosynthesis and signaling networks. Plant Cell 14:S131–S151

    CAS  PubMed  Google Scholar 

  37. Nakano T, Suzuki K, Fujimura T, Shinshi H (2006) Genome-wide analysis of the ERF gene family in arabidopsis and rice. Plant Physiol 140:411–432

    Article  CAS  PubMed  Google Scholar 

  38. Boutilier K, Offringa R, Sharma VK et al (2002) Ectopic expression of BABY BOOM triggers a conversion from vegetative to embryonic growth. Plant Cell 14:1737–1749

    Article  CAS  PubMed  Google Scholar 

  39. Chan MG, Smertenko A, Hussey PJ, Naldrett M, Bottrill A, Lloyd CW (2003) Identification of a MAP65 isoform involved in directional expansion of plant cells. FEBS Lett 534:161–163

    Article  CAS  PubMed  Google Scholar 

  40. Smertenko AP, Kaloriti D, Chang HY, Fiserova J, Opatrny Z, Hussey PJ (2008) The C-terminal variable region specifies the dynamic properties of Arabidopsis microtubule-associated protein MAP65 isotypes. Plant Cell 20:3346–3358

    Article  CAS  PubMed  Google Scholar 

  41. Sawano M, Shimmen T, Sonobe S (2000) Possible involvement of 65 kDa MAP in elongation growth of azuki bean epicotyls. Plant Cell Physiol 41:968–976

    Article  CAS  PubMed  Google Scholar 

  42. Vasák M, Hasler DW (2000) Metallothioneins: new functional and structural insights. Curr Opin Chem Biol 4:177–183

    Article  PubMed  Google Scholar 

  43. Bilecen K, Ozturk UH, Duru AD et al (2005) Triticum durum metallothionein: isolation of the gene and structural characterization of the protein using solution scattering and molecular modeling. J Biol Chem 280:13701–13711

    Article  CAS  PubMed  Google Scholar 

  44. Andrews GK (2000) Regulation of metallothionein gene expression by oxidative stress and metal ions. Biochem Pharmacol 59:95–104

    Article  CAS  PubMed  Google Scholar 

  45. Brouwer M, Syring R, Brouwer TH (2002) Role of a copper-specific metallothionein of the blue crab, Callinectes sapidus, in copper metabolism associated with degradation and synthesis of hemocyanin. J Inorg Biochem 88:228–239

    Article  CAS  PubMed  Google Scholar 

  46. Ebadi M, Leuschen MP, El Refaey H, Hamada FM, Rojas P (1996) The antioxidant properties of zinc and metallothionein. Neurochem Int 29:159–166

    Article  CAS  PubMed  Google Scholar 

  47. Dunaeva M, Adamska I (2001) Identification of genes expressed in response to light stress in leaves of Arabidopsis thaliana using RNA differential display. Eur J Biochem 268:5521–5529

    Article  CAS  PubMed  Google Scholar 

  48. Oztur ZN, Talame V, Deyholos M, Michalowski CB, Galbraith DW, Gozukirmizi N, Tuberosa R, Bohnert HJ (2002) Monitoring large-scale changes in transcript abundance in drought- and salt-stressed barley. Plant Mol Biol 48:551–573

    Article  PubMed  Google Scholar 

  49. Vitali A, Pacini E, De Mori P, Pucillo L, Maras B, Botta B, Brancaccio A, Giardina B (2006) Purification and characterization of an antifungal thaumatin-like protein from Cassia didymobotrya cell culture. Plant Physiol Biochem 44:604–610

    Article  CAS  PubMed  Google Scholar 

  50. van der Wel H, Loeve K (1972) Isolation and characterization of thaumatin I and II, the sweet-tasting proteins from Thaumatococcus daniellii Benth. Eur J Biochem 4:221–225

    Article  Google Scholar 

  51. Abad LR, D’Urzo MP, Liu D et al (1997) Antifungal activity of tobacco osmotin has specificity and involves membrane permeability. Plant Sci 118:11–23

    Article  Google Scholar 

  52. Min K, Ha SC, Hasegawa PM, Bressan RA, Yun DJ, Kim KK (2004) Crystal structure of osmotin, a plant antifungal protein. Proteins 54:170–173

    Article  CAS  PubMed  Google Scholar 

  53. Ng TB (2004) Antifungal proteins and peptides of leguminous and non-leguminous origins. Peptides 25:1215–1222

    Article  CAS  PubMed  Google Scholar 

  54. Garcia-Casado C, Collada C, Allona I, Soto A, Casado R, Rodriguez-Cerezo E, Gomez L, Aragoncillo C (2000) Characterization of an apoplastic basic thaumatin-like protein from recalcitrant chestnut seeds. Physiol Plant 110:172–180

    Article  CAS  Google Scholar 

  55. Martin A, Lee J, Kichey T et al (2006) Two cytosolic glutamine synthetase isoforms of maize are specifically involved in the control of grain production. Plant Cell 18:3252–3274

    Article  CAS  PubMed  Google Scholar 

  56. Tabuchi M, Sugiyam K, Ishiyama K, Inoue E, Sato T, Takahashi H, Yamaya T (2005) Severe reduction in growth rate and grain filling of rice mutants lacking OsGS1;1, a cytosolic glutamine synthetase1;1. Plant J 42:641–651

    Article  CAS  PubMed  Google Scholar 

  57. Hirel B, Le Gouis J, Ney B, Gallais A (2007) The challenge of improving nitrogen use efficiency in crop plants: towards a more central role for genetic variability and quantitative genetics within integrated approaches. J Exp Bot 58:2369–2387

    Article  CAS  PubMed  Google Scholar 

  58. Lea PJ, Azevedo RA (2007) Nitrogen use efficiency. 2. Amino acid metabolism. Ann Appl Biol 151:269–275

    Article  CAS  Google Scholar 

  59. Ohmiya A, Kishimoto S, Aida R, Yoshioka S, Sumitomo K (2006) Carotenoid cleavage dioxygenase (CmCCD4a) contributes to white color formation in chrysanthemum petals. Plant Physiol 142:1193–1201

    Article  CAS  PubMed  Google Scholar 

  60. Huang FC, Molnár P, Schwab W (2009) Cloning and functional characterization of carotenoid cleavage dioxygenase 4 genes. J Exp Bot 60:3011–3022

    Article  CAS  PubMed  Google Scholar 

  61. Ytterberg AJ, Peltier JB, van WiJk KJ (2006) Protein profiling of plastoglobules in chloroplasts and chromoplasts. A surprising site for differential accumulation of metabolic enzymes. Plant Physiol 140:984–997

    Article  CAS  PubMed  Google Scholar 

  62. Rubio A, Rambla JL, Santaella M, Gómez MD, Orzaez D, Granell A, Gómez-Gómez L (2008) Cytosolic and plastoglobule-targeted carotenoid dioxygenases from Crocus sativus are both involved in β-ionone-release. J Biol Chem 283:24816–24825

    Article  CAS  PubMed  Google Scholar 

  63. Bouvier F, Dogbo O, Camara B (2003) Biosynthesis of the food and cosmetic plant pigment bixin (annatto). Science 300:2089–2091

    Article  CAS  PubMed  Google Scholar 

  64. Bouvier F, Suire C, Mutterer J, Camara B (2003) Oxidative remodeling of chromoplast carotenoids: identification of the carotenoid dioxygenase CsCCD and CsZCD genes involved in Crocus secondary metabolite biogenesis. Plant Cell 15:47–62

    Article  CAS  PubMed  Google Scholar 

  65. Kato OR, Figueirêdo FJC, Belfort AJ, Nogueira OL, Barbosa WC (1992) Época de colheita de sementes de urucu: emergência e teor de corantes. Pesq Agropec Bras 27:1291–1302

    Google Scholar 

  66. Han Y, Gasic K, Korban SS (2007) Multiple-copy cluster-type organization and evolution of genes encoding o-methyltransferases in apple. Genetics 176:2625–2635

    Article  CAS  PubMed  Google Scholar 

  67. Wu S, Watanabe N, Mita S, Ueda Y, Shibuya M, Ebizuka Y (2003) Two o-methyltransferases isolated from flower petals of Rosa chinensis var. spontanea involved in scent biosynthesis. J Biosci Bioeng 96:119–128

    CAS  PubMed  Google Scholar 

  68. Christensen AB, Gregersen PL, Olsen CE, Collinge DB (1998) A flavonoid 7-O-methyltransferase is expressed in barley leaves in response to pathogen attack. Plant Mol Biol 36:219–227

    Article  CAS  PubMed  Google Scholar 

  69. He X, Reddy JT, Dixon RA (1998) Stress responses in alfalfa (Medicago sativa L.). XXII. cDNA cloning and characterization of an elicitor-inducible isoflavone 7-O-methyltransferase. Plant Mol Biol 36:43–54

    Article  CAS  PubMed  Google Scholar 

  70. Ross JR, Nam KH, D’Auria JC, Pichersky E (1999) S-Adenosyl-l-methionine: salicylic acid carboxyl methyltransferase, an enzyme involved in floral scent production and plant defense, represents a new class of plant methyltransferases. Arch Biochem Biophys 367:9–16

    Article  CAS  PubMed  Google Scholar 

  71. Ogawa M, Herai Y, Koizumi N, Kusano T, Sano H (2001) 7-Methylxanthine methyltransferase of coffee plants. Gene isolation and enzymatic properties. J Biol Chem 276:8213–8218

    Article  CAS  PubMed  Google Scholar 

  72. Mizuno K, Okuda A, Kato M, Yoneyama N, Tanaka H, Ashihara H, Fujimura T (2003) Isolation of a new dual-functional caffeine synthase gene encoding an enzyme for the conversion of 7-methylxanthine to caffeine from coffee (Coffea arabica L.). FEBS Lett 534:75–81

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgments

This research was supported by BNB (Research grant no. 2004-1-22 to MGCC) and CNPq (Research grant no. 473619/04-04 to WCO). We gratefully acknowledge the PhD scholarships to V.L.F. Soares and S.M. Rodrigues by FAPESB (Fundação de Amparo à Pesquisa do Estado da Bahia) and FAPEMIG (Fundação de Amparo à Pesquisa do Estado de Minas Gerais), respectively. T.M de Oliveira and T.O. de Queiroz are recipients of PIBIC/CNPq and IC/FAPESB fellowships, respectively. We thank Dr. Claudia Fortes Ferreira (Embrapa, Brazil) and Dr. Nicolas Carels (Fiocruz, Brazil) for helpful advice and critical reading of the manuscript.

Author information

Authors and Affiliations

  1. Departamento de Ciências Biológicas, Universidade Estadual de Santa Cruz, Ilhéus, BA, Brazil

    Virgínia L. F. Soares, Tahise M. de Oliveira, Talisson O. de Queiroz, Braz T. Hora-Júnior, Fabienne Micheli, Júlio C. M. Cascardo & Marcio G. C. Costa

  2. Departamento de Biologia Vegetal, Universidade Federal de Viçosa, Viçosa, MG, Brazil

    Simone M. Rodrigues & Wagner C. Otoni

  3. Centro de Pesquisas do Cacau, CEPLAC, Ilhéus, BA, Brazil

    Lívia S. Lima & Karina P. Gramacho

  4. CIRAD-BIOS, UMR DAP, Montpellier, France

    Fabienne Micheli

  5. Embrapa Mandioca e Fruticultura Tropical, Cruz das Almas, BA, Brazil

    Abelmon S. Gesteira

Authors
  1. Virgínia L. F. Soares
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Simone M. Rodrigues
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Tahise M. de Oliveira
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Talisson O. de Queiroz
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Lívia S. Lima
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Braz T. Hora-Júnior
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Karina P. Gramacho
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Fabienne Micheli
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. Júlio C. M. Cascardo
    View author publications

    You can also search for this author in PubMed Google Scholar

  10. Wagner C. Otoni
    View author publications

    You can also search for this author in PubMed Google Scholar

  11. Abelmon S. Gesteira
    View author publications

    You can also search for this author in PubMed Google Scholar

  12. Marcio G. C. Costa
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Marcio G. C. Costa.

Electronic supplementary material

Rights and permissions

Reprints and permissions

About this article

Cite this article

Soares, V.L.F., Rodrigues, S.M., de Oliveira, T.M. et al. Unraveling new genes associated with seed development and metabolism in Bixa orellana L. by expressed sequence tag (EST) analysis. Mol Biol Rep 38, 1329–1340 (2011). https://doi.org/10.1007/s11033-010-0234-8

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s11033-010-0234-8

Keywords

Search

Navigation