First published online September 23, 2005; 10.1104/pp.105.067827
Plant Physiology 139:674-689 (2005)
© 2005 American Society of Plant Biologists
BIOCHEMICAL PROCESSES AND MACROMOLECULAR STRUCTURES
Implications of Carotenoid Biosynthetic Genes in Apocarotenoid Formation during the Stigma Development of Crocus sativus and Its Closer Relatives1
Raquel Castillo,
José-Antonio Fernández and
Lourdes Gómez-Gómez*
Sección de Biotecnología, Instituto de Desarrollo Regional, Escuela Técnica Superior Ingenieros Agrónomos, Universidad de Castilla-La Mancha, Campus Universitario, Albacete E–02071, Spain
Crocus sativus is a triploid sterile plant characterized by its long red stigmas, which produce and store significant quantities of the apocarotenoids crocetin and crocin, formed from the oxidative cleavage of zeaxanthin. Here, we investigate the accumulation and the molecular mechanisms that regulate the synthesis of these apocarotenoids during stigma development in C. sativus. We cloned the cDNAs for phytoene synthase, lycopene- -cyclase, and -ring hydroxylase from C. sativus. With the transition of yellow undeveloped to red fully developed stigmas, an accumulation of zeaxanthin was observed, accompanying the expression of CsPSY, phytoene desaturase, and CsLYCb, and the massive accumulation of CsBCH and CsZCD transcripts. We analyzed the expression of these two transcripts in relation to zeaxanthin and apocarotenoid accumulation in other Crocus species. We observed that only the relative levels of zeaxanthin in the stigma of each cultivar were correlated with the level of CsBCH transcripts. By contrast, the expression levels of CsZCD were not mirrored by changes in the apocarotenoid content, suggesting that the reaction catalyzed by the CsBCH enzyme could be the limiting step in the formation of saffron apocarotenoids in the stigma tissue. Phylogenetic analysis of the CsBCH intron sequences allowed us to determine the relationships among 19 Crocus species and to identify the closely related diploids of C. sativus. In addition, we examined the levels of the carotenoid and apocarotenoid biosynthetic genes in the triploid C. sativus and its closer relatives to determine whether the quantities of these specific mRNAs were additive or not in C. sativus. Transcript levels in saffron were clearly higher and nonadditive, suggesting that, in the triploid gene, regulatory interactions that produce novel effects on carotenoid biosynthesis genes are involved.
1 This work was supported by the Consejería de Ciencia y Tecnología of the Junta de Communidades de Castilla-La Mancha (grant no. PAI–02–026) and by the Ministerio de Ciencia y Tecnologia Competitive Grants Program (grant no. BIO2003–05259).
Article, publication date, and citation information can be found at www.plantphysiol.org/cgi/doi/10.1104/pp.105.067827.
* Corresponding author; e-mail marialourdes.gomez{at}uclm.es; fax 34–967599233.
Received June 29, 2005;
returned for revision August 2, 2005;
accepted August 16, 2005.
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