Role of VIN3-LIKE 2 in facultative photoperiodic flowering response in Arabidopsis

In Arabidopsis, expression of FLC and FLC-related genes (collectively called FLC clade) contributes to flowering time in response to environmental changes, such as day length and temperature, by acting as floral repressors. VIN3 is required for vernalization-mediated FLC repression and a VIN3 related protein, VIN3-LIKE 1/VERNALIZATION 5 (VIL1/VRN5), acts to regulate FLC and FLM in response to vernalization.^1–3 VIN3 also exists as a small family of PHD finger proteins in Arabidopsis, including VIL1/VRN5, VIL2/VEL1, VIL3/VEL2 and VIL4/VEL3. We showed that the PHD finger protein, VIL2, is required for proper repression of MAF5, an FLC clade member, to accelerate flowering under non-inductive photoperiods. VIL2 acts together with POLYCOMB REPRESSIVE COMPLEX 2 (PRC2) to repress MAF5 in a photoperiod dependent manner.Key words: photoperiod, chromatin, floweringThe decision to flower is critical to the survival of flowering plants. Thus, plants sense environmental cues to initiate floral transition at a time that both ensures and optimizes their own reproductive fitness. Using a model plant, Arabidopsis thaliana, genetic studies have shown that the regulation of floral transition mainly consists of four genetic pathways: the inductive photoperiod pathway, the autonomous pathway, the vernalization pathway and the gibberellin pathway.⁴ In Arabidopsis, these four flowering pathways eventually merge into a group of genes called floral integrators, including FLOWERING LOCUS T (FT), SUPPRESSOR OF OVEREXPRESSION OF CONSTANS 1 (SOC1) and LEAFY (LFY). Based on the response to specific photoperiod conditions, the flowering behaviors of plants can be classified into three groups: long day (LD), short day (SD) and day neutral response.^5,6 Depending on the requirement of day length, plants show either obligate or facultative responses. For example, henbane, carnation and ryegrass are obligate long day (LD) flowering plants which flower under increasing inductive photoperiod but do not flower at all under non-inductive photoperiod.⁵ On the other hand, plants including Arabidopsis, wheat, lettuce and barley, are considered to be facultative flowering plants. Thus, these plants exhibit early flowering under LD and late-flowering under non-inductive short days (SD). Studies on photoperiodic flowering time mainly focus on the inductive LD-photoperiod pathway in Arabidopsis.     

In vitro flowering red miniature rose

Using aseptic plantlets obtained from stem node explants of hybrid red miniature rose (Rosa hybrida cv. Fairy Dance), the effects of shoot physiological status, medium ingredients, and culture thermoperiod on in vitro flowering were evaluated. Shoot height, subculture media for shoot multiplication, sucrose concentration, plant growth regulators (PGRs), mineral substances in media, and thermoperiod had a significant effect on the percentage of in vitro flowering. Shoots 3 ± 0.2 or 2 ± 0.2 cm in height cultured on Murashige and Skoog (MS) medium containing 2.0 mg dm^?3 6-benzyladenine (BA), 0.2 mg dm^?3 α-naphthaleneacetic acid (NAA), and 20 g dm^?3 sucrose were more suitable for in vitro flowering than shoots 4 ± 0.2, or 5 ± 0.2 cm in height. The most suitable sucrose concentration for in vitro flowering was 50 g dm^?3 and the most suitable PGRs were a combination of 3.0 mg dm^?3 BA and 0.1 mg dm^?3 NAA. Increasing the potassium nitrate to ammonium nitrate ratio or increasing the phosphate concentration in MS medium had a positive effect on in vitro flowering. The percentage of in vitro flowering was significantly higher at day/night temperature of 28/20 °C than at other constant temperatures. The percentage of in vitro flowering shoots reached 68.33 % despite the occurrence of abnormal flowers and some unique developmental patterns. It makes miniature rose a potentially new in vitro experimental platform for research on the molecular mechanisms of flowering ornamental plants.     

Floral stimulus movement in perilla and flower inhibition caused by noninduced leaves

Photoperiodic control of flowering in the short day plant Perilla involves the transmission of a floral stimulus from induced leaves to the shoot apex. We have studied the basipetal movement of this stimulus and of ¹⁴C-labeled assimilates in plants with an induced leaf (donor) grafted into the uppermost internode of a vegetative plant in which the axillary shoots at various nodes along the stem function as receptors.     

Identification of the Flower-inducing Factor Isolated from Aphid Honeydew as being Salicylic Acid

Honeydew produced by the aphid Dactynotus ambrosiae when feeding on flowering or vegetative plants of the short day plant Xanthium strumarium contains an active substance capable of inducing flowering in the long day plant Lemna gibba G3. In the present study, this active material has been identified as salicylic acid through the use of gas-liquid chromatography and mass, infrared, and ultraviolet spectrometry. Authentic salicylic acid induces flowering in L. gibba G3 under strict short day conditions with an optimal response at about 5.6 μm. The possible significance of salicylic acid for the control of flowering in Xanthium or L. gibba G3 is discussed.     

Miscanthus Clones Display Large Variation in Floral Biology and Different Environmental Sensitivities Useful for Breeding

A wider range of Miscanthus varieties is required to develop Miscanthus clones that are suitable for bioenergy production. For this reason, breeding programs need to be initiated using knowledge regarding the genetic influence on floral biological traits. The objective of the present study was to characterize the genotypic variation in flowering and panicle architecture traits in Miscanthus by studying (i) the clone effect on these traits and (ii) the clone sensitivity to environmental conditions. The flowering traits characterized were date of panicle emergence, date of flowering onset, and interval between these two traits. The panicle architecture traits characterized were total panicle length, longest panicle raceme size, raceme number per panicle, floral density, and total flower number per panicle. Eight clones were studied in a greenhouse under four environmental conditions including two day lengths (an 8-h short day length and a natural day length) and two temperature treatments (warm and cool). Miscanthus clones showed large differences in flowering and panicle architecture traits. Moreover, day length appeared to be the most important environmental factor creating differential clone sensitivities for the panicle emergence and the onset of flowering in contrast to temperature factor for the total flower number per panicle. In addition, the behavior of the clone Sacc was in contrast with that of the other clones for most of the traits studied. This knowledge will be useful to optimize the synchronization of flowering between Miscanthus clones for more successful breeding programs.     

Modulation of Ambient Temperature-Dependent Flowering in Arabidopsis thaliana by Natural Variation of FLOWERING LOCUS M

Plants integrate seasonal cues such as temperature and day length to optimally adjust their flowering time to the environment. Compared to the control of flowering before and after winter by the vernalization and day length pathways, mechanisms that delay or promote flowering during a transient cool or warm period, especially during spring, are less well understood. Due to global warming, understanding this ambient temperature pathway has gained increasing importance. In Arabidopsis thaliana, FLOWERING LOCUS M (FLM) is a critical flowering regulator of the ambient temperature pathway. FLM is alternatively spliced in a temperature-dependent manner and the two predominant splice variants, FLM-ß and FLM-δ, can repress and activate flowering in the genetic background of the A. thaliana reference accession Columbia-0. The relevance of this regulatory mechanism for the environmental adaptation across the entire range of the species is, however, unknown. Here, we identify insertion polymorphisms in the first intron of FLM as causative for accelerated flowering in many natural A. thaliana accessions, especially in cool (15°C) temperatures. We present evidence for a potential adaptive role of this structural variation and link it specifically to changes in the abundance of FLM-ß. Our results may allow predicting flowering in response to ambient temperatures in the Brassicaceae.     

Limitations on Leaf Nitrate Reductase Activity during Flowering and Podfill in Soybean

The objective of this study was to identify factors which limit leaf nitrate reductase (NR) activity as decline occurs during flowering and beginning seed development in soybean (Glycine max [L.] Merr. cv Clark). Level of NR enzyme activity, level of reductant, and availability of NO₃⁻ as substrate were evaluated for field-grown soybean from flowering through leaf senescence. Timing of reproductive development was altered within one genotype by (a) exposure of Clark to an artificially short photoperiod to hasten flowering and podfill, and (b) the use of an early flowering isoline. Nitrogen (N) was soil-applied to selected plots at 500 kilograms per hectare as an additional variable. Stem NO₃⁻ concentration and in vivo leaf NR activity were significantly correlated (R² = 0.69 with nitrate in the assay medium and 0.74 without nitrate in the medium at P = 0.001) across six combinations of reproductive and soil N-treatment. The supply of NO₃⁻ from the root to the leaf tissue was the primary limitation to leaf NR activity during flowering and podfill. Levels of NR enzyme and reductant were not limiting to leaf NR activity during this period.     

Regulation of flowering by green light depends on its photon flux density and involves cryptochromes

Photoperiodic lighting can promote flowering of long‐day plants (LDPs) and inhibit flowering of short‐day plants (SDPs). Red (R) and far‐red (FR) light regulate flowering through phytochromes, whereas blue light does so primarily through cryptochromes. In contrast, the role of green light in photoperiodic regulation of flowering has been inconsistent in previous studies. We grew four LDP species (two petunia cultivars, ageratum, snapdragon and Arabidopsis) and two SDP species (three chrysanthemum cultivars and marigold) in a greenhouse under truncated 9‐h short days with or without 7‐h day‐extension lighting from green light (peak = 521 nm) at 0, 2, 13 or 25 μmol m^?2 s^?1 or R + white (W) + FR light at 2 μmol m^?2 s^?1. Increasing the green photon flux density from 0 to 25 μmol m^?2 s^?1 accelerated flowering of all LDPs and delayed flowering of all SDPs. Petunia flowered similarly fast under R + W + FR light and moderate green light but was shorter and developed more branches under green light. To be as effective as R + W + FR light, saturation green photon flux densities were 2 μmol m^?2 s^?1 for LDP ageratum and SDP marigold and 13 μmol m^?2 s^?1 for LDP petunia. Snapdragon was the least sensitive to green light. In Arabidopsis, cryptochrome 2 mediated promotion of flowering under moderate green light, whereas both phytochrome B and cryptochrome 2 mediated that under R + W + FR light. We conclude that 7‐h day‐extension lighting from green light‐emitting diodes can control flowering of photoperiodic ornamentals and that in Arabidopsis, cryptochrome 2 mediates promotion of flowering under green light.     

Does zinc concentration in the substrate influence the onset of flowering in Arabidopsis arenosa (Brassicaceae)?

We investigated the impact of low zinc (Zn) concentrations in the substare on the onset of flowering in Arabidopsis arenosa (Brassicaceae). Experiments were carried out in controlled conditions using plants from four different populations. The research was aimed to verify experimentally the following hypotheses: (1) Zn content in the growth medium promote the onset of flowering in A. arenosa, (2) Changes in the onset of flowering induced by Zn depend on Zn concentration employed; (3) Zn-induced early onset of flowering is an universal plant response present within the species and is not an effect of stress or physiological adaptation to high Zn content in the environment. Investigated plants were subjected to four different Zn concentrations: 0.4 (control), 155, 775 and 1,550???M Zn²⁺. To asses stress level in investigated plants we calculated biomass accumulation and employed fluorometric methods. Zn content was estimated in shoots using atomic absorption spectroscopy. Differences in the onset of flowering were assessed using Kaplan?CMeier curves. Our results showed that Zn was transported form growth medium to roots and shoots of investigated plants and that the content of Zn increased with the increase of Zn concentration in the growth medium. We evidenced that apart from one (1,550???M Zn²⁺) applied Zn concentrations did not caused stress in investigated plants what was confirmed by two independent experimental approaches: measurement of biomass accumulation and chlorophyll a fluorescence. Flowering curves obtained on the basis of calculation of Kaplan?CMeier estimator showed that: (1) control plants originating from four different populations did not differ in terms of the onset of flowering, (2) plants from each population tested tends to enter flowering phase earlier in response to applied Zn concentrations than control plants, (3) plants treated with the lowest tested Zn concentration (155???M Zn²⁺) tend to flower earlier than plants treated with the higher concentration (775???M Zn²⁺), (4) the impact of Zn on the onset of flowering did not depend on the origin on the plant material used (Zn-rich or Zn-poor soils). Our results indicate that Zn ions present in the growth medium promote early flowering in A.arenosa and that this effect may depend on Zn concentration used. Zn-induced early flowering in A. arenosa seems to be an universal plant response present within the species and is not an effect of stress or physiological adaptation to high Zn content in the environment.     

Gibberellic Acid Reduces Flowering Intensity in Sweet Orange [Citrus sinensis (L.) Osbeck] by Repressing CiFT Gene Expression

In Citrus, gibberellic acid (GA₃) applied at the floral bud inductive period significantly reduces flowering intensity. This effect is being used to improve the fruit set of parthenocarpic cultivars that tend to flower profusely. However, the molecular mechanisms involved in the process remain unclear. To contribute to the knowledge of this phenomenon, adult trees of ‘Salustiana’ sweet orange were sprayed at the floral bud inductive period with 40?mg?L^?1 of GA₃ and the expression pattern of flowering genes was examined up to the onset of bud sprouting. Trees sprayed with paclobutrazol (PBZ, 2,000?mg L^?1), a gibberellin biosynthesis inhibitor, were used to confirm the effects, and untreated trees served as control. Bud sprouting, flowering intensity, and developed shoots were evaluated in the spring. GA₃ significantly reduced the number of flowers per 100 nodes by 72% compared to the control, whereas PBZ increased the number by 123%. Data of the expression pattern of flowering genes in leaves of GA₃-treated trees revealed that this plant growth regulator inhibited flowering by repressing relative expression of the homolog of FLOWERING LOCUS T, CiFT, whereas PBZ increased flowering by boosting its expression. The activity of the homologs TERMINAL FLOWER 1, FLOWERING LOCUS C, SUPPRESSOR OF OVEREXPRESSION OF CONSTANS 1, and APETALA1 was not affected by the treatments. The number of flowers per inflorescence, in both leafy and leafless inflorescences, was not altered by GA₃ but increased with PBZ; the latter paralleled LEAFY relative expression. These results suggest that GA₃ inhibits flowering in Citrus by repressing CiFT expression in leaves.     

Flowering patterns in three neotropical mangrove species: Evidence from a Caribbean island

The present study sought to identify the factors that drive flowering in the main neotropical mangrove species. We evaluated the effects of water regime variables and foliar meristematic activity on the flowering intensity of Rhizophora mangle, Laguncularia racemosa, and Avicennia germinans in three physiographic types of San Andres Island, Colombia. The results show that pore salinity regulates flowering intensity and periodicity in all three mangrove species. All species flowering showed significant correlations with water balance and air vapor pressure deficit (VPD). In the fringe and interior mangroves, R. mangle flowering was explained linearly by salinity (25%) and monthly change in salinity (47%), respectively. L. racemosa flowering was linked with stronger periods of foliar meristematic activity and occurred during months of relatively high water balance (54-233 mm) and low VPD (1.18-1.29 kPa). The flowering of A. germinans was triggered by water deficit conditions when the monthly pore salinity increased over 30 g L⁻¹ and, with a month delay response, when the water column height (WCH) was below ground. The flowering of A. germinans was also explained by these variables at 65% and 39%, respectively. The flowering patterns of the studied mangrove species indicate that reproduction within the neotropical mangrove community depends on seasonally contrasting water conditions on an annual basis.     

On the thermogenesis of the Titan arum (Amorphophallus titanum)

The Titan arum (Araceae) produces the largest bloom of all flowering plants. Its flowering period of two days is divided into a female flowering phase in the first night and a male flowering phase in the second night. Recently, we have documented thermogenesis in the spadix of the Titan arum during the female flowering phase. Here, we document a second thermogenic phase in which the male florets are heated during the male flowering phase. Obviously the two nocturnal thermogenic phases are linked with the two flowering periods. These observations now allow a more detailed understanding of the flowering behavior of the Titan arum.Key words: Amorphophallus titanum, araceae, thermogenesis, infrared thermography, pollinationThe Titan arum (Amorphophallus titanum) is one of the most spectacular flowering plants. It has drawn the attention of botanists and naturalists since its discovery in 1878. However, the species has been rare in cultivation and flowering events in botanical gardens were even rarer. Besides, flowering events observed in the plant''s natural habitat are very few.^1,2 Therefore, the knowledge on A. titanum that depended on exact observations and scientific experiments remained very limited. It was only in the late 90s when a monograph on the species, containing anatomical details and some first experimental hypotheses, has been published.¹The Botanical Gardens of the University of Bonn (Germany) have been cultivating Amorphophallus titanum for more that 70 years and obtained 14 flowerings between 1937 and 2009. These regular flowering events have been the prerequisite to study A. titanum in detail. Consequently the data and hypotheses in the monograph mentioned above were gained mainly from the A. titanum plants in the Botanical Gardens Bonn. As a result of three flowering events in 2006, we have recently documented for the first time, that the inflorescence undergoes thermogenesis in which the central column (spadix) heats up to 36°C. Meanwhile four additional flowering events yielded additional insights into the flowering behavior of A. titanum.The inflorescence of A. titanum consists of a thickened unbranched inflorescence axis bearing hundreds of small female and male florets which are spatially separated (Fig. 1A). The inflorescence axis is extended into an appendix (spadix) and enveloped by a large bract referred to as spathe. The spathe enclosing the florets forms the floral chamber. Since the whole inflorescence functions as a single unit in pollination is often referred to as a bloom or “flower”. A. titanum has two timely separated flowering phases: a female flowering phase during the first evening and night after opening of the spathe and a male flowering phase in the following night.Open in a separate windowFigure 1(A) Flash-light photograph of an Amorphophallus titanum flowering zones showing part of the appendix, the male florets and the female florets. (B) Thermographic image taken during the male flowering phase. The male florets are heated to the maximum temperature of 35.9°C whereas the other parts of the plant have largely ambient air temperature of 26°C.Thermogenesis plays an important role in the pollination ecology of Araceae^3,4 and therefore occurs in many genera.^4–7 Similarly in A. titanum, we have reported the thermogenic spadix during the female flowering phase. Based on our observations of now six inflorescences, the heat production is determined and begins around 20 h, the temperature maximum of 36–38°C being reached around midnight. The duration of heat production differs between individual plants but usually stops between 2 h and 4 h in the morning. The spathe begins to close the next day in the early morning hours or in the forenoon. The opening and closing of the spathe seems to be influenced by the hours of daylight but these might be different in European countries from the plants native habitat in the tropical rainforests of Sumatra. The flowering events in Bonn usually take place in summer, the spathe opens during a daytime when it is still very bright and is fully opened when the daylight is decreasing while it is already dark then in the tropics.Some authors have observed heating of the male florets prior to appendix heating in other Araceae species.^4,5,8,9 In A. titanum however, we could not find an evidence for this, as stated in our previous article. But a question that still remained open is: when exactly the male flowering phase begins and if there might be thermogenic activity during the male flowering phase. To study this, the male florets were made visible by removing a part of the spathe of two flowering A. titanum and observed right after opening of the spathe. The beginning of the male flowering phase is easily to determine since the pollen is shed in well visible string-like structures.The pollen dissemination began in the evening around 17:20 h. Thereupon we filmed the male florets with a thermographic camera (Flexcam, GORATEC) taking an image every five minutes. The male florets were clearly thermogenic reaching a temperature maximum of 35.9°C between 18:40 h and 20:00 h (Fig. 1B). They slowly cooled down to ambient air temperature (ca. 26°C) around midnight. To test whether the temperature in the floral chamber around the male florets increases while they are heated, we recorded the temperature within the spathe of three intact inflorescences with data loggers (Tinytag, Gemini Data Loggers). However, no warming within the chamber in comparison with ambient air temperature could be measured, so the heated florets seem not to affect the temperature within the floral chamber.The flowering behavior of A. titanum is summarised in Figure 2. The carrion-like odor and the thermogenic spadix attract pollinators in the female flowering phase, during the first evening and night of the flowering period. Heating of the male florets occurs when no more odor is produced and hence no olfactorical attraction of the pollinators can take place. As a consequence, there must be only one attraction time period which is more or less restricted to the female flowering phase and to the nighttime where pollinators can be successfully attracted. The pollinators, although not exactly known,¹⁰ hence must be active only in these evening hours and at night. Once attracted, the pollinators stay inside the inflorescence and most likely use it as mating site or as a place to stay during the following day. It has already been hypothesised that Araceae inflorescences forming floral chambers may offer mating sites or places to rest for insects and rather keep their pollinators inside the floral chamber instead of a second attraction phase.^{11, 12} Numerous insects inside a A. titanum inflorescence have indeed been observed in its natural habitat,¹³ although the author did not explain these observations, it provides evidence for our hypothesis that pollinators spend some time inside the inflorescence.Open in a separate windowFigure 2Scheme of the flowering behavior of Amorphophallus titanum over its two-days flowering period. The scheme is idealised but represents observation of seven A. titanum inflorescences that all behave highly similar. Deviations in the time when opening and closing of the spathe begin in individual plants are indicated with a dashed line.The male florets are heated while pollen is released. There is evidence that at least some insects are able to percept IR light and it has been hypothesised that infrared radiation itself could be an attractant for insects, most likely to locate food sources.¹⁵ Floral heat may also be a direct reward for pollinators, helping them increasing their body temperature and thus faster reaching their activity level.^11,14 Both may also apply to A. titanum—the insects that have spent the day within the flower chamber may use the heated surface of the male florets to warm themselves up and by this collect pollen or feed on pollen. Still, a verification of these hypotheses could only come from field observations.To draw a conclusion, the new observations reported here now allow us a good understanding of the flowering behavior of A. titanum. Its two thermogenic phases are clearly linked with the two flowering phases and the plant''s complex interaction with its pollinators.