| Anthocyanin biosynthesis can be readily studied in flowers that change colors naturally and are controlled by predictable environmental triggers. Viola cornuta cv. Yesterday, Today and Tomorrow (YTT), is a useful model system to study anthocyanin biosynthetic steps because open flowers undergo a striking color change from white to purple over an eight-day period. I have experimentally determined that both light and pollination are necessary for floral color change in V. cornuta---flowers that are kept in the dark or are un-pollinated remain white. I have found that floral pH values do not vary significantly across color stages. Color change in V. cornuta is due to the gradual accumulation of the pigment malvidin, overlaying constant levels of the copigment myricetin.;Myricetin is an early product of the anthocyanin biosynthetic pathway, and malvidin is produced at later stages, thus, these results suggest that early steps of the anthocyanin biosynthetic pathway are constitutively expressed in white V. cornuta flowers and that later steps of the pathway are up-regulated to produce malvidin. In many plant systems, chalcone synthase (Chs) is the most commonly regulated early gene in the pathway, and dihdyroflavonol 4-reductase (Dfr) and anthocyanidin synthase (Ans) are regulated late genes in the pathway. I have determined that Chs and Dfr are expressed in stage I flowers, and their expression over ontogeny increases moderately and considerably, respectively. I have also determined that Ans is barely expressed in stage I flowers and is dramatically up-regulated in stage II and stage III flowers; thus, Ans is a key regulated step in V. cornuta color change. Furthermore, pollination and light are necessary for expression of Chs, Dfr and Ans, because all three genes are considerably down-regulated in the absence of these cues. Finally, I have investigated whether V. cornuta Chs, Dfr and Ans are encoded by multi-gene families in order to specify which gene family member is expressed. I have detected only one copy of Dfr and Ans in V. cornuta. V. cornuta Chs, though, is composed of a small multi-gene family with at least four members, and I have established that a single copy is expressed. |
