Flower Color Change Research Articles

POPULATION GENETICS, PLEIOTROPY, AND THE PREFERENTIAL FIXATION OF MUTATIONS DURING ADAPTIVE EVOLUTION

Ongoing debate centers on whether certain types of mutations are fixed preferentially during adaptive evolution. Although there has been much discussion, no quantitative framework currently exists to test for these biases. Here, we describe a method for distinguishing between the two processes that likely account for biased rates of substitution: variation in mutation rates and variation in the probability that a mutation becomes fixed once it arises. We then use this approach to examine the type and magnitude of these biases during evolutionary transitions across multiple scales: those involving repeated origins of individual traits (flower color change), and transitions involving broad suites of traits (morphological and physiological trait evolution in plants and animals). We show that fixation biases can be strong at both levels of comparison, likely due to differences in the magnitude of deleterious pleiotropy associated with alternative mutation categories. However, we also show that the scale at which these comparisons are made greatly influences the results, as broad comparisons that simultaneously analyze multiple traits obscure heterogeneity in the direction and magnitude of these biases. We conclude that preferential fixation of mutations likely is common in nature, but should be studied on a trait-by-trait basis.

Shoot regeneration from leaf explants of Brunfelsia calycina

A protocol for plantlet regeneration through shoot formation was developed for the neotropical shrub Brunfelsia calycina. This shrub is unique in its change in flower color from dark purple to white. Explants from young and mature leaves were incubated on MS medium (pH 5.7, 30 g/l sucrose, 7.5 g/l agar) with various combinations of Indole-3-acetic acid (IAA) and 6-Benzyladenine (BA) under a 16 h photoperiod at a constant temperature of 25°C. Shoot emergence was best at 4.44 μM BA and 2.85 μM IAA for young leaf explants, and at 8.88 μM BA, 2.85 μM IAA for mature leaf explants. When shoots were transferred to MS medium supplemented with 1.23–2.46 μM indole butrytic acid (IBA), they developed roots.

Pigment composition patterns and expression analysis of flavonoid biosynthesis genes in the petals of evergreen azalea ‘Oomurasaki’ and its red flower sport

With 2 figures and 3 tables Abstract To obtain basic information about flower colour sports in azalea breeding, the petals of Rhododendron × pulchrum‘Oomurasaki’ and its red flower sport were compared in terms of pigment constitution, expression of eight genes related to flavonoid synthesis and sequences of the flavonoid 3′,5′-hydroxylase (F3′5′H) gene, which is a key gene that synthesizes the pigments of the delphinidin series. According to the HPLC analysis, ‘Oomurasaki’ contained anthocyanidins from both the cyanidin and delphinidin series, whereas the red flower sport contained only pigments from the cyanidin series. Real-time PCR analysis showed that F3′5′H transcript levels in the red flower sport were 0.14-fold those of ‘Oomurasaki’. Compared with the amino acid sequences of F3′5′H1-A of ‘Oomurasaki’, both F3′5′H1-B and F3′5′H1-C sequences of the red flower sport were changed by only one amino acid. These results suggested that low transcript levels of the F3′5′H gene caused no accumulation of delphinidin series pigments in flower colour change between ‘Oomurasaki’ and its red flower sport.

Bidirectional Flower Color and Shape Changes Allow a Second Opportunity for Pollination

Flowers act as "sensory billboards" with multiple signals (color, morphology, odor) attracting and manipulating potential pollinators. Many use changing signals as indicators that visitation and/or pollination have occurred). Floral color change is commonly used to transmit this information (often correlated with reduced nectar reward) and can be specifically triggered by pollination or visitation. By retaining color-changed flowers, plants benefit from larger floral displays but also indicate at close range which flowers are still rewarding (and still unpollinated), so that visitors forage more efficiently. However, the legume Desmodium setigerum shows a unique ability, if inadequately pollinated, to reverse its flowers' color and shape changes. Single visits by bees mechanically depress the keel and expose stigma and anthers (termed "tripping"); visits also initiate a rapid color change from lilac to white and turquoise and a slower morphological change, the upper petal folding downwards over the reproductive parts. But flowers receiving insufficient pollen can partially reopen, re-exposing the stigma, with a further color change to deeper turquoise and/or lilac. Thus, most flowers achieve pollination from one bee visit, but those with inadequate pollen receipt can reverse their signals, earning a "second chance" by eliciting attention from other potential pollinators.

The Birds, the Bees, and the Virtual Flowers: Can Pollinator Behavior Drive Ecological Speciation in Flowering Plants?

Biologists have long assumed that pollinator behavior is an important force in angiosperm speciation, yet there is surprisingly little direct evidence that floral preferences in pollinators can drive floral divergence and the evolution of reproductive (ethological) isolation between incipient plant species. In this study, we expose computer-generated plant populations with a wide variation in flower color to selection by live and virtual hummingbirds and bumblebees and track evolutionary changes in flower color over multiple generations. Flower color, which was derived from the known genetic architecture and phenotypic variance of naturally occurring plant species pollinated by both groups, evolved in simulations through a genetic algorithm in which pollinator preference determined changes in flower color between generations. The observed preferences of live hummingbirds and bumblebees were strong enough to cause adaptive divergence in flower color between plant populations but did not lead to ethological isolation. However, stronger preferences assigned to virtual pollinators in sympatric and allopatric scenarios rapidly produced ethological isolation. Pollinators can thus drive ecological speciation in flowering plants, but more rigorous and comprehensive behavioral studies are required to specify conditions that produce sufficient preference levels in pollinators.

The Birds, the Bees, and the Virtual Flowers: Can Pollinator Behavior Drive Ecological Speciation in Flowering Plants?

Biologists have long assumed that pollinator behavior is an important force in angiosperm speciation, yet there is surprisingly little direct evidence that floral preferences in pollinators can drive floral divergence and the evolution of reproductive (ethological) isolation between incipient plant species. In this study, we expose computer‐generated plant populations with a wide variation in flower color to selection by live and virtual hummingbirds and bumblebees and track evolutionary changes in flower color over multiple generations. Flower color, which was derived from the known genetic architecture and phenotypic variance of naturally occurring plant species pollinated by both groups, evolved in simulations through a genetic algorithm in which pollinator preference determined changes in flower color between generations. The observed preferences of live hummingbirds and bumblebees were strong enough to cause adaptive divergence in flower color between plant populations but did not lead to ethological isolation. However, stronger preferences assigned to virtual pollinators in sympatric and allopatric scenarios rapidly produced ethological isolation. Pollinators can thus drive ecological speciation in flowering plants, but more rigorous and comprehensive behavioral studies are required to specify conditions that produce sufficient preference levels in pollinators.

Adaptive Floral Colour Change in Erysimum scoparium (Brassicaceae) and Pollinator Behaviour of Anthophora alluadii on Tenerife (Hymenoptera: Apidae)

[Article] Flowering plants manipulate the behaviour of their pollinators in a wide range of ways, including exploiting associative learning by changing the colour of flowers as they age and at the same time withdrawing the nectar reward. This study investigated the adaptive function of the floral colour change from white to purple in Erysimum scoparium (Brouss. ex Willd) Wettst in relation to its main pollinator, Anthophora alluadii (Perez 1895). Both species are endemic to the Canary Islands. Field observations of nectar production and bee behaviour, plus inflorescence and whole plant manipulation experiments at sites in Tenerife in 2003,2004 and 2006, provided support for three hypotheses. Hypothesis 1: The flower colour change in £ scoparium serves as a visual signal, indicating to pollinators that the nectar reward of the flower has changed. Late phase purple flowers provide much less (and more frequently zero) nectar compared to early phase white flowers, and pollinators make fewer visits of shorter duration to purple flowers. Hypothesis 2: Removal of purple flowers from inflorescences reduced the rate of pollinator visitation to white flowers on those inflorescences. Retention of late phase, purple flowers therefore increased the visual attraction of individual inflorescences, and this shon-distance (up to tens of centimetres) visual signal directs pollinators to newly opened, rewarding flowers on that inflorescence potentially reducing geitonogamy. Hypothesis 3: Finally, evidence was found that retention of late phase, purple flowers on the plant acted as a long-distance visual signal to pollinators, highlighting the presence of plants over scales of metres to tens of metres. Removal of all purple flowers from individual plants did not result in reduced overall visitation rates to those plants, but it did change the relationship between plant size and bee visitation rate, from a positive correlation to no significant relationship. In relation to experimentally testing the function of the floral colour change, previous studies have obtained contrasting results. This may be because whole flower colour changes have evolved as medium and long range signals, whereas colour change of isolated parts of flowers may more often be a short range signal.

Modification of a viral satellite DNA-based gene silencing vector and its application to leaf or flower color change in Petunia hybrida

Virus-induced gene silencing offers a powerful reverse-genetic tool for the study of gene function in plants. We have previously reported effective gene silencing of plant genes using a viral satellite DNA associated with Tomato yellow leaf curl China virus (TYLCCNV). In this study, we further modified the viral satellite DNA-based vector. The modified vector can induce sulfu (Su) gene silencing as effective as the original vector in Nicotiana benthamiana plants, but the new system simplifies procedures for construction of vector derivative. Furthermore, a fragment of petunia Su or chalcone synthase (CHS) endogenous gene was inserted into the modified vector. When petunia plants were agroinoculated with the modified vector carrying a Su or CHS gene, the Su silenced plants started to appear yellowing in veins of systemically infected upper leaves two weeks after agroinoculation, while the CHS silenced plants started to show flower color change one month after agroinoculation and later single-color flowers became mosaic.

FLOWER ABSCISSION AND ANTHOCYANIN DEVELOPMENT IN CUT PHLOX FLOWER HEADS: EFFECT OF ETHYLENE INHIBITORS AND SUCROSE

Phlox paniculata `John Fanick' produces long lasting, dense terminal flower heads and has potential as a specialty cut flower. Quality and postharvest display life of cut flower heads depends primarily on ethylene-induced flower abscission, flower bud opening, and maintenance and development of flower color during vase life. Late events, such as flower and leaf senescence may also be detrimental to flower quality. In the control treatment, the initial red-pink and purple flower color changes to violet blue in 3 to 4 days, and may lose >50% of initial anthocyanins. Incorporating sucrose (SUC) in the vase solution not only maintained >75% of the initial floral pigments, but also promoted opening of additional flowers and anthocyanin development. Although both ethylene biosynthesis (AOA, ReTain, a.i. AVG) and action inhibitors (STS, 1-MCP) delayed flower abscission, STS and 1-MCP were relatively more effective than AOA and AVG. As in the control, newly opened flowers remained very small when treated with ethylene inhibitors, did not develop red-pink color, and exhibited only shades of violet blue color. Sucrose antagonized the effect of ethylene inhibitors. As such, the flowers in SUC+ethylene inhibitors treatments enlarged in size and developed a reddish-pink blue color. However, the flower quality in SUC alone was much superior than those in SUC+ethylene inhibitors. These results indicate that ethylene inhibitors, alone and in combination with SUC, were not of any additional value in improving postharvest performance and display life of cut phlox flower heads.

Clonal fidelity of chrysanthemum regenerated from long term cultures

Morphological characteristics of flowers of long term regenerated chrysanthemum, cv. "White Spider", after ten years of micropropagation are investigated. Shoot cultures are established and maintained more than ten years by stem segment culture on MS medium supplemented with BAP and NAA (1.0, 0.1 mgL-1, respectively). Rooting of shoots (100 %) has done on MS medium without hormones and it was very successful after ten years, as well as, after two or eight years of micropropagation. Acclimation of rooted chrysanthemum plantlets at greenhouse conditions was excellent and after appropriate photoperiod "in vitro" plants flowered 90.3 % and have the same flower color, shape and size as mother plants. Flower color changes of "in vitro" plants are observed during another flowering cycle one year after acclimatization. Observed variations of chrysanthemum flowers could be attributed to epigenetic factors.

Effects of high temperature on flower colour and anthocyanin content in pink flower genotypes of greenhouse chrysanthemum (Chrysanthemum morifolium Ramat.)

SummaryThe relationship between the intensity of flower colour and changes in the content of the main anthocyanins under various controlled temperatures was examined in order to clarify the effects of high temperature on flower colouration in six pink flower genotypes of greenhouse chrysanthemum (Chrysanthemum morifolium Ramat.). Poor colouration of flowers was observed at 30°C in all genotypes except ‘Chatoo’. This genotype showed little difference in flower colour between different temperature treatments. The degree of change in flower colour differed depending on the genotype, whereas no clear differences in flower colouring were observed between Summer – Autumn flowering and Autumn-flowering genotypes. All genotypes showed lower contents of the two anthocyanins tested [cyanidin 3-O-(6’’-O-monomalonyl- -glucopyranoside) and cyanidin 3-O-(3’’,6’’-O-dimalonyl- -glucopyranoside)] at higher temperatures. Therefore, flower colour changes were attributable to changes in these two main anthocyanins. Differences in colouration between genotypes and temperature conditions were also detectable in values that were measured using a colorimeter. Changed parameters that were visually verifiable were the a* value, representing the degree of red colour, and the C* value, representing chroma. For ‘Sei-Monako’, which showed visually greater differences between temperature treatments, the a* and C* values were low under high temperature conditions. On the other hand, in ‘Chatoo’, the differences detected by eye and those in a* and C* values between temperature treatments were small. In addition, the present results indicate that mean temperature is more important than either day or night temperature in determining the degree of flower colouration.

Molecular characterization of the flavonoid biosynthesis of Verbena hybrida and the functional analysis of verbena and Clitoria ternatea F3'5'H genes in transgenic verbena

Homologues of the flavonoid 3′,5′-hydroxylase (F3′5′H) gene, a key gene determining flower color, were obtained from a Verbena hybrida (verbena) cultivar Temari Violet, verbena cultivar Tapien Pink, and Clitoria ternatea (butterfly pea). The expression of the Temari and butterfly pea homologues in yeast confirmed that they encoded F3′5′H. The two genes under the control of an enhanced cauliflower mosaic virus 35S promoter were introduced into verbena Temari Sakura. Some of the transgenic verbena plants had elevated delphinidin contents and flower color altered toward violet. Interestingly, the butterfly pea F3′5′H gene yielded more delphinidin and gave clearer flower color change than the verbena Temari gene in the transgenic verbena plants. The results indicate that the choice of the gene source should be considered to obtain strong phenotypic changes, even if the genes encode the same enzymatic activity. We also cloned some flavonoid biosynthetic genes from verbenas. The potential usefulness of verbena in the phytomonitoring of environmental pollutants is also discussed.

Flavonoid components and flower color change in transgenic tobacco plants by suppression of chalcone isomerase gene

A cDNA encoding chalcone isomerase (CHI) was isolated from the petals of Nicotiana tabacum and the effect of its suppression on flavonoid biosynthesis was analyzed in transgenic tobacco plants. CHI-suppression by RNA interference (RNAi) showed reduced pigmentation and change of flavonoid components in flower petals. The plants also accumulated high levels of chalcone in pollen, showing a yellow coloration. Our results first demonstrated that suppression of CHI by genetic transformation is possible in higher plants. This suggests that CHI plays a major part in the cyclization reaction from chalcone to flavanone, and that spontaneous reactions are few, if any, in tobacco plants.

(283) Petal Color Changes in Carnation Plants Transformed with an Antisense DFR and a CHI Gene

To produce transgenic carnation plants expressing modified flower colors, cultivars of red-flowering carnation (`Desio' and `Garnet') were transformed with a antisense DFR gene encoding dihydroflavonol-4-reductase, and yellow-flowering carnations (`Pallas' and `Keumbyul') were introduced with a CHI gene encoding chalcone synthease isomerase. Explants of leaves, stems, and shoot tips were inoculated with Agrobacterium and grown overnight in induction broth supplemented with 0.2 mm acetosyringone. The cocultures of carnation explants were maintained in the light at 24 °C on one-half strength MS medium solidified with 0.3% Gelite and supplemented with 1.0 mg·L-1 BA for 10 days, and then were transferred to a selection medium containing 250 mg·L-1 cefotaxime/carbenicillin, 500 mg·L-1 kanamycin. They were subcultured every 2 weeks for over six times and regenerated. In `Garnet' and `Desio' carnation cultivars, a total of 146 plants was transformed with anti-sense DFR gene in pGA748/LBA4404. In yellow flowering cultivars `Pallas' and `Keumbyul', transformed with CHI gene in pGA748/LBA4404, 20 putative transformants obtained. The introduction of the transgenes were confirmed through PCR and flower color changes. Rates of phenotypic expression of the transgene, antisense DFR gene, vary among the transformants such as lighter pigmentation or chimeric discoloration, more prominently in outer petals of the flowers. Transgenic plants of chi gene bloomed flowers of lighter yellow petals in general. The most prominent phenotypic changes were discoloration of red strips on petals, which are typical characteristics of the cultivars.

Flower Color Development and Nano‐Science

AbstractFor Abstract see ChemInform Abstract in Full Text.

Erratum: Engineering of flower color in forsythia by expression of two independently-transformed dihydroflavonol 4-reductase and anthocyanidin synthase genes of flavonoid pathway

Flower color was modified in forsythia (Forsythia x intermedia cv ‘Spring Glory’) by inducing anthocyanin synthesis in petals through sequential Agrobacterium-mediated transformation with dihydroflavonol 4-reductase from Antirrhinum majus (AmDFR) and anthocyanidin synthase from Matthiola incana (MiANS) genes. This is the second report of flower color modification of an ornamental shrub after rose, and the first time an ANS gene is used for this purpose. Double transformants (AmDFR+MiANS) displayed a novel bronze-orange petal color, caused by the de novo accumulation of cyanidin-derived anthocyanins over the carotenoid yellow background of wild type (wt), and intense pigmentation of vegetative organs. Transformation with single genes (either AmDFR or MiANS) produced no change in flower color, showing a multistep control of late anthocyanin pathway in petals of forsythia. Analysis of relevant late flavonoid pathway genes – an endogenous flavonoid glycosyltransferase (FiFGT) and transformed DFR and ANS genes – showed appropriate expression in flower organs. Functional characterization of FiFGT expressed in E. coli revealed its ability to metabolize both flavonols and anthocyanidin substrates, a prerequisite for effective anthocyanin accumulation in petals of plants transformed with constructs leading to anthocyanidin synthesis. Biochemical analyses of flavonoid compounds in petals and leaves showed that, besides anthocyanin induction in petals of double transformants, the accumulation pattern of flavan-3-ols was quantitatively and qualitatively modified in petals and leaves of transformants, in agreement with the most recent model proposed for flavan-3-ol synthesis. On the other hand, phenylpropanoid, flavone and flavonol pools were not quantitatively affected, indicating a tight regulation of early flavonoid pathway.

花の色とナノサイエンス

Red, purple and blue flower color is mainly due to anthocyanins. The color development and stability of anthocyanin are effected by co-existence of co-pigment and metal ions, and pH. Our chemical studies elucidated that the color is stabilized and varied by formation of a supramolecule. Especially beautiful blue flower colors, found in blue dayflower and blue salvia, are developed by a metalloanthocyanin, a stoichiometric supramolecular metal complex pigment.The metalloanthocyanins are composed with six molecules of anthocyanins, six molecules of glycosylflavones, and 2 atoms of metal ion and the diameter is ca. 3 nano meter. Under formation of metalloanthocyanin fine chiral molecular recognition occurs. To clarify the mechanism we synthesized chiral glycosylflavones derivatives according to our newly invented method of direct C-and O-glycosylation to the flavonoid nucleus. Reproduction experiment from the components (anthocyanin, flavone and metal ion) including chiral isomer of the flavones bearing L-glucose was carried out. As a result, the 4'-D-glucoside of the flavone is essential for formation of metalloanthocyanin, but the L-sugar is not.As the petal tissue is not homogenous, both the epidermides are colored. For elucidation of true flower color development in a living petal cell, various analytical methodologies including nano-technology must be applied. For this purpose one cell analysis by pH-microelectrode, a nano-HPLC, a pico mole metalanalysis and microscopic photometry were carried out. The mech-anism of flower color change of morning glory, Ipomoea tricolor, from purple to blue during blooming, and color variation of hydrangea was studied.

Flavonoid composition related to petal color in different lines of Clitoria ternatea

Flavonoids in the petals of several C. ternatea lines with different petal colors were investigated with LC/MS/MS. Delphinidin 3- O-(2″- O-α-rhamnosyl-6″- O-malonyl)-β-glucoside was newly isolated from the petals of a mauve line (wm) together with three known anthocyanins. They were identified structurally using UV, MS, and NMR spectroscopy. Although ternatins, a group of 15 (poly)acylated delphinidin glucosides, were identified in all the blue petal lines (WB, BM-1, ‘Double Blue’ and ‘Albiflora’), WM accumulated delphinidin 3- O-(6″- O-malonyl)-β-glucoside instead. The white petal line (WW) did not contain anthocyanins. Quantitative data showed that the total anthocyanin contents in WB and ‘Double Blue’ were ca. 8- and 10-fold higher than that in BM-1, a bud mutant of ‘Double Blue’, respectively. The total anthocyanin content in ‘Albiflora’ was less than 2×10 −3 times those in WB or ‘Double Blue’. While all the lines contained the same set of 15 flavonol glycosides in similar relative ratios, the relative ratio of myricetin glycosides in ww and ‘Albiflora’ was ca. 30–70 times greater than those in the other lines. The change in flower color from blue to mauve was not due to a change in the structure of an anthocyanidin from delphinidin, but to the lack of (polyacylated) glucosyl group substitutions at both the 3′- and 5′-positions of ternatins. This implies that glucosylation at the 3′- and 5′-positions of anthocyanin is a critical step in producing blue petals in C. ternatea.

How does insect visitation trigger floral colour change?

Abstract. 1. Visitation by the key pollinator, Bombus terrestris, was implicated in inducible flower colour change in Lupinus pilosus. Behaviour at the flower and rate of visitation by these bumble bees had specific effects; exclusion of this flower visitor led to retarded onset, and reduced rate, of colour change.2. The foraging behaviour of B. terrestris was influenced by floral colour change in L. pilosus. Choice of pre‐change flowers was greater than random in relation to the proportion of colour phases available within the plant population.3. Levels of floral manipulation that mimicked the flower handling characteristics of visiting bumble bees confirmed that triggering of the pollen release mechanism is necessary for the instigation of colour change.4. Moreover, this study suggests that, in L. pilosus, an aspect of pollination (pollen deposition by bees and/or subsequent pollen tube growth within the style) is linked with colour change and may act as the trigger for such change.

Genetic engineering of the anthocyanin biosynthetic pathway with flavonoid-3′,5′-hydroxylase: specific switching of the pathway in petunia

Flavonoid-3′,5′-hydroxylase (F3′5′H) is the key enzyme in the synthesis of 3′,5′-hydroxylated anthocyanins, which are generally required for the expression of blue or purple flower color. It has been predicted that the introduction of this enzyme into a plant species that lacks it would enable the production of blue or purple flowers by altering the anthocyanin composition. We present here the results of the genetic engineering of petunia flower color, pigmentation patterns and anthocyanin composition with sense or antisense constructs of the F3 ′ 5 ′ H gene under the control of the CaMV 35S promoter. When sense constructs were introduced into pink flower varieties that are deficient in the enzyme, transgenic plants showed flower color changes from pink to magenta along with changes in anthocyanin composition. Some transgenic plants showed novel pigmentation patterns, e.g. a star-shaped pattern. When sense constructs were introduced into blue flower petunia varieties, the flower color of the transgenic plants changed from deep blue to pale blue or even pale pink. Pigment composition analysis of the transgenic plants suggested that the F3 ′ 5 ′ H transgene not only created or inhibited the biosynthetic pathway to 3′,5′-hydroxylated anthocyanins but switched the pathway to 3′,5′-hydroxylated or 3′-hydroxylated anthocyanins.