Long-day Plant Silene Armeria Research Articles

Induction of flowering by DNA demethylation in Perilla frutescens and Silene armeria: Heritability of 5-azacytidine-induced effects and alteration of the DNA methylation state by photoperiodic conditions

The cytidine analogue 5-azacytidine (azaC), which causes DNA demethylation, induced flowering in the long-day plant Silene armeria and the short-day plant Perilla frutescens var. crispa under non-inductive photoperiodic conditions, suggesting that the expression of photoperiodic flowering-related genes is regulated by DNA methylation. The progeny of plants induced to flower by azaC did not flower under non-inductive photoperiodic conditions. The DNA gel blot hybridization analysis revealed no differences in the methylation states of genomic DNA from the azaC and control progeny in P. frutescens. These results suggest that the flowering-related genes activated by DNA demethylation were remethylated in the progeny. This remethylation system may allow the genes to be demethylated in response to inductive photoperiodic conditions in each generation. The azaC treatment also induced dwarfism in P. frutescens, and this characteristic was inherited by the progeny. Similar relationships between the azaC-induced dwarfism and the demethylation/remethylation system could not be established. Genome-wide methylation changes were examined by methylation-sensitive amplified fragment length polymorphism (MS-AFLP) analysis in P. frutescens, and the results showed that the DNA methylation state was altered by photoperiodic conditions.

Physiological Signals That Induce Flowering.

The timing of the transition from vegetative growth to flowering is of paramount importance in agriculture, horticulture, and plant breeding because flowering is the first step of sexual reproduction. Studies to understand how this transition is controlled have occupied countless physiologists during the past half century and have produced an almost unmanageably large amount of information (Bernier et al., 1981a; Halevy, 1985-1989; Bernier, 1988; Kinet, 1993). A majority of plants use environmental cues to regulate the transition to flowering because all individuals of a species must flower synchronously for successful outcrossing and because all species must complete their sexual reproduction under favorable externa1 conditions. Any environmental variables exhibiting regular seasonal changes are potential factors that control the transition to flowering. The major factors are photoperiod, temperature, and water availability. Plants that do not require a particular photoperiod or temperature to flower, i.e., the so-called “autonomous-flowering” plants, are usually sensitive to irradiance. The environmental factors are perceived by different parts of the plant. Photoperiod and irradiance are perceived mainly by mature leaves in intact plants. Temperature is perceived by all plant parts, although low temperature (vernalization) is often perceived mainly by the shoot apex. Water availability is perceived by the root system. There are strong interactions between these different factors, so that each factor can change the threshold value for the effectiveness of the others. Plants, as opportunists, will thus make use of a different critical factor in different environments. Melilotus officinalis, for example, is a biennial with a vernalization requirement in temperate zones and an annual long-day (LD) plant with no cold requirement in arctic regions. In photoperiodic species, such as the short-day (SD) plant Pharbitis nil and the LD plant Silene armeria, flowering in unfavorable photoperiods can be caused by changing temperature, irradiance, or nutrition or by removing the roots. Similarly, in some late-flowering mutants of Arabidopsis, vernalization and an increase in the proportion of far-red light in the light source can substitute for one another in promoting the transition to flowering (Martinez-Zapater and Somerville, 1990; Bagnall, 1992). Clearly, there are alternate pathways to flowering in most, if

Stem elongation and changes in the levels of gibberellins in shoot tips induced by differential photoperiodic treatments in the long-day plant Silene armeria

The effects of differential photoperiodic treatments applied to shoot tips and mature leaves of the long-day (LD) plant Silene armeria L. on growth and flowering responses, and on the levels of endogenous gibberellins (GAs), were investigated. Gibberellins were analyzed by gaschromatography-mass spectrometry and the use of internal standards. Exposure of mature leaves to LD, regardless of the photoperiodic conditions of the shoot tips, short days (SD), LD, or darkness, promoted elongation of the stems and of the immature leaves. Long-day treatment of the mature leaves modified the levels of endogenous GAs in shoot tips kept under LD, SD, or darkness. In shoot tips kept in LD or darkness the levels of GA53 were reduced, whereas the levels of GA19 and GA20 were increased. The contents of GA1 were increased in all three types of shoots: SD twofold, LD fivefold, and darkness eightfold. Dark treatment of the shoot tips on plants of which the mature leaves were grown in SD promoted elongation of the immature etiolated leaves and increased the GA1 content of the shoot tips threefold. However, this treatment did not cause stem elongation. The different photoperiodic treatments applied to the shoot tips did not change the levels of GAs in mature leaves. These results indicate that both LD and dark treatments result in an increase in GA1 in shoot tips. In addition, it is proposed that LD treatment induces the formation of a signal that is transmitted from mature leaves to shoot tips where it enhances the effect of GA on stem elongation.

Cellular changes induced by exogenous and endogenous gibberellins in shoot tips of the long-day plant Silene armeria

Stem elongation and flowering are two processes induced by long-day (LD) treatment in Silene armeria L. Whereas photoperiodic control of stem growth is mediated by gibberellins (GAs), the flowering response cannot be obtained by GA applications. Microscopic observations on early cellular changes in the shoot meristem following LD induction or GA treatment in short days (SD) were combined with GA analyses of stem sections at various distances below the shoot apex. The earliest effects of both LD and GA induction on the subapical meristem were an increase in the number of cells per cell file and a reduction of cell length in the meristematic tissue approx. 1.0-3.0 mm below the shoot apex. Within 8 d after the beginning of LD induction or after GA application, the cells in the subapical meristem were oriented in long files. In induced tips, cellulose deposition occurred mostly in longitudinal walls, indicating that many transverse cell divisions had taken place which, in turn, increased the length of the stem. In contrast to LD induction, GA treatments did not promote the transition from the vegetative to the floral stage. Endogenous GAs were analyzed by selected ion monitoring (SIM), using labeled internal standards, in extracts from transverse sections of the tip at various distances below the apical meristem. In control plants, the levels of the six 13-hydroxy GAs studied (GA53, GA44, GA19, GA20, GA1, and GA8) decreased as the distance from the apical meristem increased. Except for GA53, GA levels were higher in tips of LD-induced plants, particularly in the meristematic zone approx. 0.5-1.5 mm below the apical meristem. In comparison with SD, the highest increase observed was for GA1, the content of which increased 30-fold in the zone 0.5-3.5 mm below the shoot apex. These data indicate a spatial correlation between the accumulation of GA1 and its precursors, and the enhanced mitotic activity which occurs in the subapical meristem of elongating Silene apices.

Gibberellins and Stem Growth as Related to Photoperiod in Silene armeria L.

Stem growth and flowering in the long-day plant Silene armeria L. are induced by exposure to a minimum of 3 to 6 long days (LD). Stem growth continues in subsequent short days (SD), albeit at a reduced rate. The growth retardant tetcyclacis inhibited stem elongation induced by LD, but had no effect on flowering. This indicates that photoperiodic control of stem growth in Silene is mediated by gibberellins (GA). The objective of this study was to analyze the effects of photoperiod on the levels and distribution of endogenous GAs in Silene and to determine the nature of the photoperiodic after-effect on stem growth in this plant. The GAs identified in extracts from Silene by full-scan combined gas chromatography-mass spectrometry (GC-MS), GA(12), GA(53), GA(44), GA(17), GA(19), GA(20), GA(1), GA(29), and GA(8), are members of the early 13-hydroxylation pathway. All of these GAs were present in plants under SD as well as under LD conditions. The GA(53) level was highest in plants in SD, and decreased in plants transferred to LD conditions. By contrast, GA(19), GA(20), and GA(1) initially increased in plants transferred to LD, and then declined. Likewise, when Silene plants were returned from LD to SD, there was an increase in GA(53), and a decrease in GA(19), GA(20), and GA(1) which ultimately reached levels similar to those found in plants kept in SD. Thus, measurements of GA levels in whole shoots of Silene as well as in individual parts of the plant suggest that the photoperiod modulates GA metabolism mainly through the rate of conversion of GA(53). As a result of LD induction, GA(1) accumulates at its highest level in shoot tips which, in turn, results in stem elongation. In addition, LD also appear to increase the sensitivity of the tissue to GA, and this effect is presumably responsible for the photoperiodic after-effect on stem elongation in Silene.

The Effect on Flower Formation in the Long-day Plant Silene armeria L. of Interrupting Long Photoperiods by Short Periods of Darkness

Interrupting long photoperiods by short periods of darkness — indicated as «Day Interruption» - resulted in an accelerated flower formation when applied early during the photophase as compared with late application, the total amount of light being the same in both cases. Day Interruption primarily influenced floral induction, whereas realization was influenced to a much smaller extent or not at all. It is not darkness as such but absence of light that is the determining factor. Different effects of Day Interruption according to the time of its application can be understood as different effects of the daily photoperiodic action, i.e. weak at the onset, but increasing sharply near the critical daylength. Suppression of the light action by darkness at a time when the effect of light is weak, results in earlier flower formation than suppression at a time when this effect is strong. The combined results of Day Interruption and Night Interruption lead to the conclusion that the photoperiodic action of light during a 24 h-cycle follows a circadian rhythm.

The Mechanism of Vernalization in Silene armeria L.

Vernalization by low temperature (5 °C) in the long-day plant Silene armeria L. is only effective when applied during the dark phases of short-day cycles. This is similar to the effect of temperatures above 30 °C. Presumably, chromosomal reblocking of the floral-hormone forming gene in darkness takes place at medium temperatures only, and neither at extremely low nor at extremely high temperatures.

The Direct Action of the Floral Hormone in Silene armeria L.

The difference in the time of flowering between an early line, «Early», and a very late line, «Late», of the long-day plant Silene armeria L. was studied. Juvenility, growth rate and leaf surface were not involved, whereas the intensity of the DNA-blocking in vegetative plants was: this blocking was relatively weak in «Early» and very strong in «Late», resulting in rapid and slow deblocking, in early and late flowering, respectively. When Early and Late were grafted on D(onor) Early in short day (SD), 100% simultaneous flowering occurred, so that the difference in time of flowering between ungrafted Early and Late had completely disappeared (Table 1). When, however, Early and Late were grafted on D Late in SD, no flowering followed. Upon transferring the plants to long day, very advanced and simultaneous flowering took place, so that again the difference in the time of flowering had completely disappeared (Table 2). These results are interpreted as a direct action of the floral hormone from the donors, leaving the blocking in the receptors Early and Late undisturbed. The remaining problem is the difference in action between the floral hormones from D Early and D Late. When «induction» is defined as deblocking, the term «indirect induction» must be considered as obsolete, since evidently no deblocking at all takes place as a result of grafting.

Influence of daylength on gibberellin metabolism and stem growth in Silene armeria

When radioactive gibberellin A5 ((3)H-GA5) was applied to the apices and surrounding young leaves of the long-day plant Silene armeria, it was partially converted to at least two other acidic substances. One of them was similar to GA3 in chromatographic, but not in biological properties. The other metabolite was more polar than GA3 and inactive in the dwarf d-5 corn assay.The rate of (3)H-GA5 conversion was influenced by the photoperiod under which Silene plants were grown. Exposure to 2 long days significantly increased (3)H-GA5 metabolism over that in control plants kept under short days. The increased conversion of (3)H-GA5 persisted for at least a few days after transferring Silene plants back from long to short days. Likewise, stem growth induced by long photoperiods continued for a considerable period of time under subsequent short days.Application of the growth retardant AMO-1618 to Silene reduced the levels of two endogenous GA-like substances, one of them with GA5-like properties, more under long than under short days. These results indicate that long photoperiods, which induce flower formation and stem elongation in Silene, increase the turnover of endogenous gibberellins.