The promoter of the carotenoid cleavage dioxygenase 4a-5 gene of Chrysanthemum morifolium (CmCCD4a-5) drives petal-specific transcription of a conjugated gene in the developing flower

Abstract

Carotenoids comprise one of the major groups of pigments in flowers. Because carotenoids are physiologically indispensable pigments for all photosynthetic plants, their catabolism must be discretely regulated in photosynthetic organs and non-photosynthetic organs such as petals or fruits. In the chrysanthemum, carotenoid cleavage dioxygenase 4a (CmCCD4a), which is dominantly expressed in petals, cleaves carotenoid, leading to a white flower. CmCCD4a-5 was recently identified as a new member of the CmCCD4a family, but its detailed expression profile in plant tissues has not yet been established. In this study, we sequenced a 1094-bp region upstream of CmCCD4a-5 and assessed its petal-specific promoter activity. To evaluate the activity of this gene, we constructed two types of transgenic Arabidopsis thaliana that possessed, respectively, a fusion gene of a 1090-bp or 505-bp segment of the upstream region plus the β-d-glucuronidase (GUS) gene (1090bUR::GUS and 505bUR::GUS). GUS activity in the 505bUR::GUS strain was observed mainly in the anthers/pollen in flower buds, whereas GUS activity of the 1090bUR::GUS strain was observed in immature petals of the flower buds. Among the cis-acting elements located between positions -505 and -1090, no elements that have previously been reported to enhance the expression in petals or to suppress it in anthers/pollen were detected by PLACE analysis, indicating the existence of unknown cis-element(s). A semiquantitative reverse transcription-polymerase chain reaction analysis revealed that CmCCD4a-5 transcription was prominent in petals but was undetectable in roots, stems and leaves.

Keywords: 5-bromo-4-chloro-3-indolyl β-d-glucuronide; CCD; CCD4a; Carotenoid cleavage dioxygenase; Chrysanthemum morifolium; CmCCD4a; CmCCD4a-5; GUS; NCED; RT-PCR; X-Gluc; carotenoid cleavage dioxygenase; nine-cis-epoxycarotenoid; reverse transcription-polymerase chain reaction; β-d-glucuronidase.