Main conclusion
BpCYP76AD15 is involved in betaxanthin biosynthesis in callus, but not in bracts, in bougainvillea.
Abstract
Bougainvillea (Bougainvillea peruviana) is a climbing tropical ornamental tree belonging to Nyctaginaceae. Pigments that are conferring colorful bracts in bougainvillea are betalains, and that conferring yellow color are betaxanthins. In general, for red-to-purple betacyanin biosynthesis, α clade CYP76AD that has tyrosine hydroxylase and DOPA oxygenase activity is required, while for betaxanthin biosynthesis, β clade CYP76AD that has only tyrosine hydroxylase is required. To date, betaxanthin biosynthesis pathway genes have not been identified yet in bougainvillea. Since bougainvillea is phylogenetically close to four-O-clock (Mirabilis jalapa), and it was reported that β clade CYP76AD, MjCYP76AD15, is involved in floral betaxanthin biosynthesis in four-O-clock. Thus, we hypothesized that orthologous gene of MjCYP76AD15 in bougainvillea might be involved in bract betaxanthin biosynthesis. To test the hypothesis, we attempted to identify β clade CYP76AD gene from yellow bracts by RNA-seq; however, we could not. Instead, we found that callus accumulated betaxanthin and that β clade CYP76AD gene, BpCYP76AD15, were expressed in callus. We validated BpCYP76AD15 function by transgenic approach (agro-infiltration and over-expression in transgenic tobacco), and it was suggested that BpCYP76AD15 is involved in betaxanthin biosynthesis in callus, but not in bracts in bougainvillea. Interestingly, our data also indicate the existence of two pathways for betaxanthin biosynthesis (β clade CYP76AD-dependent and -independent), and the latter pathway is important for betaxanthin biosynthesis in bougainvillea bracts.
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Data availability
The data that support the findings of this study are openly available in the Genbank and Sequence Read Archive under the accession number PRJDB15488 (https://www.ncbi.nlm.nih.gov/bioproject/PRJDB15488). Accession numbers: BpCYP76AD15A cDNA (LC760651), BpCYP76AD15B cDNA (LC760652). RNA-seq: ‘California Gold’ bract at stage 1 (DRR452711); ‘Golden Gold’ bract at stage 1 (DRR452712).
Abbreviations
- ADH:
-
Arogenate dehydrogenase
- B6GT:
-
Betanidin-6-O-glucosyltransferase
- cDOPA5GT:
-
Cyclo-DOPA 5-O-glucosyltransferase
- CYP76AD:
-
Cytochrome p450 76AD
- DODA:
-
DOPA 4,5-dioxygenase
- HPLC:
-
High-performance liquid chromatography
- L-DOPA:
-
3,4-Dihydroxy-L-phenyalanine
- RACE:
-
Rapid amplification of cDNA ends
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Computations were partially performed on the NIG supercomputer at ROIS National Institute of Genetics (Japan).
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Ohno, S., Kokado, R., Makishima, R. et al. BpCYP76AD15 is involved in betaxanthin biosynthesis in bougainvillea callus. Planta 258, 47 (2023). https://doi.org/10.1007/s00425-023-04202-3
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DOI: https://doi.org/10.1007/s00425-023-04202-3