ARS | Publication request: THE CONCENTRATION OF TWO REGULATORY PROTEINS AFFECTS THE AMOUNTS OF ANTHOCYANIN PRODUCED IN FLOWERS

Submitted to: Journal of the American Society for Horticultural Science
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: August 13, 2007
Publication Date: January 4, 2008
Citation: Ma, H., Pooler, M.R., and Griesbach, R.J. 2008. Ratio of Myc and Myb transcription factors regulates antheoyanin production in orchid flowers. J. Amer. Soc. Hort. Sci. 133(1):133-138.

Interpretive Summary: Novel coloration and patterns of coloration add aesthetic appeal to ornamental plants and therefore have great commercial value to the floral and nursery industries. The red, purple, and blue colors of vegetative and floral organs are due to anthocyanins, the prominent pigments in higher plants. Anthocyanin synthesis is regulated by genetic factors, environmental stimuli and developmental cues. Two regulatory genes have been isolated from several species and have been expressed in a number of plant species in which they either enhanced the amount of anthocyanin produced, or activated de novo biosynthesis in unpigmented tissues. However, in the majority of those studies, both regulatory genes were overexpressed, which resulted in deleterious effects on plant growth. This effect could be due to either high levels of anthocyanin induced by the increased expression of the two regulatory genes or the accumulation of high levels of the two transcription factors. In our study, the regulatory genes were expressed in white orchid petals using either a strong promoter or a weak promoter, along with an internal transformation control of the green fluorescent protein. An in-vivo functional assay was developed to efficiently monitor the activity of the regulatory factors through anthocyanin production, which indicated that the two regulatory genes might play different roles in activating anthocyanin biosynthesis in terms of when, where, and how much anthocyanin is accumulated. This information could benefit future studies aimed at creating viable plants with new flower or foliage color.

Technical Abstract: Many studies have attempted to induce flavonoid pigmentation in colorless tissues by introducing anthocyanin regulatory genes of the R and C1 families. Expression of the two regulatory genes under the control of a strong promoter generally results in high levels of anthocyanin accumulation. However, such approaches usually have a negative effect on growth and development of the recovered plants. In this study, we employed two promoters of different strengths, a weak (potato polyubiquitin Ubi3) and a strong (double 35S) promoter, and generated two sets of expression constructs with the maize anthocyanin regulatory genes Lc and C1. A transient expression system was developed using biolistic bombardment of white Phalaenopsis amabilis flowers which we confirmed to be anthocyanin regulatory gene mutants. Transient expression of different combinations of the four constructs would generate three different ratios (Lc/C1>1, Lc/C1=1, and Lc/C1<1) of the two regulatory factors in P. amabilis. The enhanced green florescent protein gene (EGFP) was co-transformed as an internal control with the two anthocyanin regulatory gene constructs. Our results demonstrate that the ratio of the two regulatory factors had a significant influence on the amount of anthocyanin produced. The anthocyanin level induced was higher when C1 was under the control of the 35S promoter than the Ubi3 promoter, regardless of whether Lc expression was driven by the 35S or Ubi3 promoter.