Changes in the Protein Composition of the Shoot Apical Bud of Sinapis alba in Transition to Flowering

Vegetative plants of Sinapis alba L. grown in short days were induced to flower by expsoure to one or continuous long days. In both inductive conditions, the first flowers were initiated about 60 h after the start of the treatment. Soluble protein extracts were prepared from apical buds and just-expanded leaves of both vegetative and induced plants. Rabbit antisera were prepared using extracts from vegetative and reproductive buds. Immunodiffusion tests were performed. Analysis of the precipitin bands indicated that: (1) one antigenic protein was present in the vegetative buds and disappeared from the buds of induced plants between 96 and 240 h after the start of the inductive treatment; (2) the concentration of a another antigenic protein increased in buds of induced plants 30 h after the start of the inductive treatment; (3) the concentration of a third antigenic proteín increased in buds of induced plants at 96 h.     

Inhibition of growth and synchronised cell division in the shoot apex in relation to flowering in Silene

When plants of Silene coeli-rosa (L.) Godron were induced by seven long days, then exposed to darkness for 48 h before being returned to short days, they went on to initiate flowers with a delay of about 2 d. The synchronisation of cell division which normally occurs before flower initiation was suppressed, showing that it is not essential for flowering. Periods of darkness of up to 240 h inhibited apical growth and leaf initiation but did not prevent eventual flowering in short days. The commitment of the apex to flower was therefore maintained while apical growth was inhibited.Abbreviations SD short day(s) - LD long day(s)     

Appearance and disappearance of proteins in the shoot apical meristem of Sinapis alba in transition to flowering

Vegetative plants of Sinapis alba L. grown under short days were induced to flower by exposure to one long day or continuous long days. Irrespective of the number of long days, the first flower primordia were initiated by the shoot apical meristem 60 h after the start of the inductive treatment. An indirect histoimmunofluorescence technique was used to search in the apical meristem for three antigenic proteins which had been previously detected by immunodiffusion tests in the whole apical bud (Pierard et al. (1977) Physiol. Plant. 41, 254–258). One protein called protein A, present in the vegetative meristem, increased in concentration during the first 48 h following the start of the inductive treatment. It stayed constant up to 96 h and disappeared completely at a later time. Two other proteins called B and C, absent in the vegetative meristem, appeared in the meristem of induced plants between 30 and 36 h after the start of the inductive treatment and progressively accumulated at later times up to 240 h. These proteins appeared 8 h before the irreversible commitment of the meristem to produce flower primordia (point of no return) was reached and 24 h before start of flower production. These observations support an interpretation of floral evocation as consisting, at least partially, of an early and qualitative change in gene expression.Abbreviations AVB anti-vegetative-bud antiserum - ARB antireproductive-bud antiserum - IgG immunoglobulins G - TRITC tetramethylrhodamine isothiocyanate - GAR IgG goat antirabbit IgG - S₀ IgG non-immune rabbit IgG     

Changes in the carbohydrate content of the leaf and the apical bud of Sinapis during transition to flowering

Vegetative plants of Sinapis alba L. were induced to flower by a single long day of 20 h or by a single short day of 8 h starting at an unusual time of the 24-h cycle (displaced short day). The soluble sugar and starch contents of the just-expanded leaf and the apical bud were measured at various times after the start of each of these two photoinductive treatments. Associated with the induction of flowering there were temporary increases in the soluble sugar and starch contents of the leaf and of the bud. These increases were apparent 14 h after the start of the long day and 12 h after the start of the displaced short day. The starch content of the bud increased later. These results indicate that an increase of the soluble sugar content of the bud is required for its transition from the vegetative to the reproductive condition.     

Flower bud differentiation in Salix pentandra as affected by photoperiod, temperature and growth regulators

Flower bud initiation in seedlings and vegetatively propagated plants of Salix pentandra from different locations has been studied under controlled conditions. In mature plants flower bud formation was induced by 2-chloroethyltrimethylammoniumchloride (CCC) and by short day treatment. The effect of CCC was antagonized by GA₃. The critical photoperiod for flower bud formation was about 18 h for a southern clone (from 49°48'N), but cuttings of a northern ecotype (from 69°39'N) formed flower buds even at 24 h photoperiod. Generally, flower bud formation occurred simultaneously with apical growth cessation. However, apical growth cessation was not a prerequisite for floral initiation and flower buds were also found in elongating plants. Seedlings up to 60 days old did not form flower buds in growth chamber studies. The length of the juvenile phase has not been studied in detail, but cuttings taken from seedlings approximately 20 cm high and 60 days old were able to develop flower buds when treated with CCC. A gradual transition from the juvenile to the mature phase was obtained by repeated pruning of seedlings grown at 18°C and 24 h photoperiod.     

Synchronisation of cell division in the shoot apex of Silene in relation to flower initiation

In plants of Silene coeli-rosa, induced to flower by 7 LD, synchronisation of cell division in 20 per cent or more of the cells in the shoot apical dome was found on the 8th and 9th days after the beginning of induction, during the plastochron before sepal initiation. Synchronisation was inferred from the changes in the proportions of cells with the 2C and 4C amounts of DNA, and changes in mitotic index and labelling index. From the peaks of mitotic index a cell cycle of 10 h was measured for the synchronised cells, half that of cells in the apices of uninduced plants in short days. The faster cell cycle and synchronisation in the induced plants was associated with a shortening, of both G1 and G2, suggesting two control points, while S and M remained unchanged. These results are compared with those from other plants in which synchronisation occurs at the beginning rather than the end of evocation.Abbreviations LD long day(s) - SD short day(s) - S DNA synthesis phase of cell cycle - G1 pre-S interphase - G2 post-S interphase - M mitosis     

Changes in RNA levels in the shoot apex of Silene during the transition to flowering

Changes in RNA concentration in the shoot apical meristem during induction and the transition to flowering were measured histochemically in Silene coeli-rosa (L.) Godron, a long-day plant. In the apices of plants induced by 7 long days the RNA concentration increased to about 25 per cent higher than in non-induced plants. Three long days did not induce flowering but resulted in a transient rise in RNA concentration. When plants were given long days interrupted by varying numbers of short days successful induction was accompanied by a sustained increase in RNA concentration but those treatments which were not inductive gave only transient increases in RNA. Gibberellic acid had no effect on induction or apical growth rates but increased the RNA concentration by 50 per cent or more in both induced and non-induced plants. Plants induced to flower at 13° C had the same RNA concentration and growth rate at the apex as in non-induced plants at 20° C. Since changes in RNA concentration in the apex could occur without changes in growth rate and without flowering, and induction could occur without a change in RNA concentration or growth rate, it is suggested that the increase in RNA and growth rate which normally occur at the transition to flowering might not be essential for the formation of a flower but may be more closely related to the rapid growth associated with the formation of the inflorescence.Abbreviations LD long day - SD short-day     

Sucrose increase during floral induction in the phloem sap collected at the apical part of the shoot of the long-day plant Sinapis alba L.

Sinapis alba L., a long-day plant, has been induced to flower either by a single 22-h-long photoperiod or by an 8-h short photoperiod displaced by 10 h in a 24 h cycle. The ehtylenediametetraacetate method previously used for leaf exudation was modified to collect phloem sap at the apical part of the shoot. Carbohydrates in the phloem sap have been analysed comparatively in vegetative and induced plants, using high-performance liquid chromatography and refractometry. Sucrose was the major sugar detected. A dramatic increase of its flux in the apical sap occurred early and transiently during the floral transition in plants induced by both long days and displaced short days. These results indicate a message-like role for sucrose since they fit nicely with previous observations indicating that an early event in the floral transition in S. alba is the accumulation of sucrose in the meristem.Abbreviations DSD displaced short day - LD long day - SD short day This work was supported by the Fonds pour la Recherche Fondamentale et Collective (FRFC) of Belgium (Grant n 2.9009.87) and the University of Liège (Action de Recherche Concertée 88/93-29). One of us (P.L.) is grateful to the I.R.S.I.A. for the award of a research fellowship.     

Jasmonates Inhibit Flowering in Short-Day Plant Pharbitis nil

The role of jasmonates in the photoperiodic flower induction of short-day plant Pharbitis nil was investigated. The plants were grown in a special cycle: 72 h of darkness, 24 h of white light with lowered intensity, 24-h long inductive night, 14 days of continuous light. At 4 h of inductive night the cotyledons of non-induced plants contained about two times the amount of endogenous jasmonates (JA/JA-Me) compared to those induced. A 15-min long pulse of far red light (FR) applied at the end of a 24-h long white light phase inhibited flowering of P. nil. The concentration of jasmonates at 2 and 4 h of inductive night in the cotyledons of the plants treated with FR was similar. Red light (R) could reverse the effect of FR. R light applied after FR light decreased the content of jasmonates by about 50%. Methyl jasmonate (JA-Me) applied to cotyledons, shoot apices and cotyledon petioles of P. nil inhibited the formation of flower buds during the first half of a 24-h long inductive or 14-h long subinductive night. Application of JA-Me to the cotyledons was the most effective. None of the plants treated with JA-Me on the cotyledons in the middle of the inductive night formed terminal flower buds. The aspirin, ibuprofen and phenidone, jasmonates biosynthesis inhibitors partially reversed the effect of FR, stimulating the formation of axillary and terminal flower buds. Thus, the results obtained suggests that phytochrome system control both the photoperiodic flower induction and jasmonates metabolism. Jasmonates inhibit flowering in P. nil.     

Induction and floral determination in the terminal bud of Nicotiana tabacum L. cv. Maryland Mammoth,a short-day plant

Floral determination in the terminal bud of the short-day plant Nicotiana tabacum cv. Maryland Mammoth has been investigated. Plants grown continuously in short days flowered after producing 31.4±1.6 (SD) nodes while plants grown continuously in long days did not flower and produced 172.5±9.5 nodes after one year. At various ages, expressed as number of leaves that were at least 1.0 cm in length above the most basal 10-cm leaf, one of three treatments was performed on plants grown from seed in short days: 1) whole plants were shifted from short days to long days, 2) the terminal bud was removed and then rooted and grown in long days, and 3) the terminal bud was removed and then rooted and grown in short days. Whole plants flowered only when shifted from short days to long days at age 15 or later. Only rooted terminal buds from plants at age 15 or older produced plants that flowered when grown in long days. Only terminal buds from plants at age 15 or older that were rooted and grown in short days produced the same number of nodes as they would have produced in their original locations while buds from younger plants produced more nodes than they would have in their original locations. Thus, determination for floral development in the terminal bud, as assayed by rooting, is simultaneous with the commitment to flowering as assayed by shifting whole plants to non-inductive conditions.Abbreviations LD long day(s) - SD short day(s) - DN dayneutral     

Inhibition imposed by developing flowers on further flower-bud initiation in Chamelaucium uncinatum Schau

In Chamelaucium uncinatum, an Australian woody perennial, flower initiation ceases under continuous inductive short-day (SD) conditions after the first flowering flush. The developing flowers were found to be the prime cause of the cessation in flower initiation. Removal of flowering shoots or lowers as soon as the buds appeared resulted in continuous flower formation. Pruning the plants below the young flower buds at the same stage also caused increased flower formation at the tips of the new growth. If pruning was delayed until flower buds were approx. 3 mm in diameter, however, nor further flower initiation took place and the plants, although still under inductive conditions, shifted to vegetative growth. The inhibiting factor is translocated from one branch to another. At least a six-week rest period (a vegetative growth period under long-day conditions) is needed before the plants are able to respond to further SD stimuli.Abbreviations LD long day - SD short day - SE standard error     

Burst Potential Characterisation by Capacity for Nucleotide Accumulation in Rhododendron Catawbiense Apical Buds

Rhododendron catawbiense cv. Album propagated in vitro were transferred ex vitro and grown in a greenhouse, under long or short days. Under long days, the rhythmic growth led to an acrotonous development. In contrast, under short days, the upper buds were unable to burst, allowing basitony. In both photoperiodic conditions, the apical buds were sampled at different stages of the experiment. Growth capacities of the isolated buds were estimated by measuring their abilities to increase and diversify their non-adenylic nucleotide pool (NTP) after supplying adenosine as a precursor. Under long days, during the growth pause, the apical buds were able to increase and diversify their NTP pool. Under short days, adenosine was used to produce important quantities of ATP, while NTP pool increase became weaker. Nevertheless, during this long growth pause, apical bud tissues retained capacities to increase their NTP pool until the basal shoots developed.     

Early effects of floral induction on cell division in the shoot apex of Silene

The changes in cell number, the relative proportions of interphase nuclei with different amounts of DNA, mitotic index and labelling index have been investigated in the shoot apex of Silene coeli-rosa L. (a long-day plant) during the first long day of photoinduction, and compared with the corresponding changes in plants in short days. 3 h after the start of induction the proportion of nuclei in the G2 phase of the cell cycle had increased, the mitotic index tended to be higher, and the labelling index was lower than in plants in short days. 8–9 h later the values for plants in the long day had become similar to those for plants in short days. No evidence was obtained for a synchronisation of cells in one phase of the cell cycle as a result of photoinduction. The results obtained were consistent with a temporary shortening of the cell cycle in the induced apices over the first long day which resulted in a greater increase in cell number by the end of the first day of photoinduction than in plants in short days.Abbreviations LD long day - SD short day     

Cytokinin Fluxes during Floral Induction in the Long Day Plant Sinapis alba L

Sinapis alba is a long-day (LD) plant that can be induced to flower by a single LD. A number of changes normally occurring in the meristem of plants subjected to the LD can be produced in short day by a single application of a cytokinin to the apical bud. However, flower buds are not produced indicating that evocation by the cytokinin is only partial. In this work, the cytokinin content of root exudate, obtained under vacuum, and of leaf exudate, obtained by the EDTA-method, has been analyzed comparatively in vegetative and induced plants, using reversed-phase HPLC coupled to the Amaranthus bioassay. The results show that, as early as 16 hours after the start of the LD, there is an increase of cytokinin activity in both the root and leaf exudates of induced plants. These observations fit nicely with previous results obtained on Sinapis, and they indicate that cytokinins are part of the floral stimulus in this species.     

Effect of Photoperiod and Chlorogenic Acid on Morphogenesis in Leaf Discs of Streptocarpus Nobilis

Leaf discs from vegetative plants greatly increase their phenolic content when cultivated in vitro. Under long days the values remained constant, and were higher when compared with short days cultures. Under short days total phenolics decreased after 10 d, corresponding to the induction and expression of in vitro flowering. The effect of photoperiod and chlorogenic acid (0.01 mM) on leaf discs cultured from induced and non-induced plants, were analyzed regarding the neo-formation of roots, as well as vegetative and flower buds. Chlorogenic acid enhances the regeneration of roots in all treatments tested, with the highest stimulation on induced leaf discs cultivated in short days. The flowering was not affected by chlorogenic acid, but an inhibitory effect was observed on the neo-formation of vegetative buds in non-induced explants maintained in short days. Vegetative buds were reduced by 50% in flower-induced leaf discs cultivated under short days.     

Studies on the Photoperiodic Responses in Short-Day Plant Helianthus tubevotus

Plants of Helianthus tuberosus, variety white tuber, were treated with various daylengths of 4, 6, 8, 10, 11, 12, 13, 14, 15, 18 h. for 25 days as soon as six leaves on a plant appeard Irradiation for 6–13 h per day induced the plants to form flower buds and flowering, daylength with 14 h or longer kept the plants in vegetative growth. The experiments showed that this variety of Helianthus tuberosus required short days for flowering and the critical daylength was about 13 h. The plants were treated with short days for different durations. At least 17 days were required, Formation of flower buds and flowering had positive correlation with the number of short days over 17 days. After short-day induction, the shorter the daylength is, the more the flower buds inverted. Long-day treatment after an appropriate period of short days wouid reduce the number of flower and induce new vegetative branches from flowering granehes.     

Apical Growth Cessation and Shoot Tip Abscission in Salix

Time course of apical shoot growth and shoot tip abortion in northern ecotypes (lat. 69°39′N, long. 18°37′E) of Salix pentandra and S. caprea have been investigated. In trees more than 15 years old growing under natural climatic conditions apical growth cessation and shoot tip abortion normally occurred in June-July when the day length still was 24 h. Application of GA₃, in spring to the apex effectively delayed growth cessation and shoot tip abortion. Application of kinetin was without effect. First-year seedlings of both species grew continuously at temperatue of 9 to 24°C in 24 h photoperiod. Short days induced apical growth cessation, but two to four (S. pentandra) or three to five (S. caprea) weeks of 12 h photoperiod were required to stop the elongation growth. The results indicated that the critical photoperiod for apical growth cessation in the used ecotype of S. pentandra was 16 to 18 h at 18°C. Short days had a minor effect only on the formation of apical leaf primordia in small seedlings. Development of axillary buds and radial growth were stimulated by short days when compared with long days. Small seedlings of both species (3 to 8 cm high at the start) formed terminal buds in short days, but in large seedlings (more than about 15 cm high) apical growth cessation was accompanied by shoot tip abortion. Abscisic acid applied to the apex or through a leaf did not induce growth cessation in S. pentandra seedlings grown in continuous light. The growth retardants CCC, B-9 and Phosphon D reduced growth rate under continuous light and induced shoot tip abortion in some plants. The effect of CCC was counteracted by GA₃. Apical growth cessation in short days was significantly delayed by a single GA₁ application.     

Cell division and morphological changes in the shoot apex of Arabidopsis thaliana during floral transition

Eight-week-old vegetative plants of Arabidopsis thaliana, ecotype Columbia, were induced to flower by a single long day (LD). In this experimental system, it is known that the last component of the floral stimulus moves from the leaves to the apex 24-36 h after the start of the LD, and the first floral meristem is initiated by the shoot apical meristem (SAM) at 44-56 h (Corbesier et al., 1996, The Plant Journal 9: 947-952). Here we show that the rate of cell division is increased at floral transition in all SAM parts but not in the sub-apical pith cells. Mitotic activity starts to increase 24 h after the start of the LD and is two- to three-fold higher at peak times than that in non-induced plants. This activation is followed by the start of SAM enlargement at 44 h, SAM doming at 48 h, and the elongation of apical internodes (bolting) at 52 h.     

RNA synthesis in the cells of the apical meristem of Sinapis alba during transition from the vegetative to the reproductive condition

Summary Vegetative plants of Sinapis alba, a long-day species, were induced to flower by exposure to a single 20-hr long day. RNA synthesis in the apical meristem of vegetative (control) and induced plants was investigated by using ³H-uridine and autoradiography of sections.Light-microscope autoradiographs showed a sharp increase in total RNA synthesis per cell in induced meristems. This increase occurred as early as 18 hr after the start of the long day, i.e. at the presumed time of the arrival of the floral stimulus at the meristem. At the same time, electron-microscope autoradiographs showed that there were changes in the pattern of RNA synthesis in the meristematic cells. The ratio of the number of grains in the nucleus to that in the cytoplasm slightly decreased and the ratio of the number of grains in the chromatin to that in the nucleolus greatly increased.Experiments with 2-thiouracil (2-TU), a pyrimidine analogue which was shown to inhibit RNA synthesis in Sinapis, indicated that this compound was most inhibitory to floral induction between the 12th and the 20th hour after the start of the long day, i.e. at the same time as important quantitative and qualitative changes in RNA synthesis were detected in induced meristems by autoradiographic methods. It was thus assumed that 2-TU inhibits floral induction via its effect on these (or on one of these) changes.     

Cytokinin levels in leaves, leaf exudate and shoot apical meristem of Arabidopsis thaliana during floral transition

Understanding the complete picture of floral transition is still impaired by the fact that physiological studies mainly concern plant species whose genetics is poorly known, and vice versa. Arabidopsis thaliana has been successfully used to unravel signalling pathways by genetic and molecular approaches, but analyses are still required to determine the physiological signals involved in the control of floral transition. In this work, the putative role of cytokinins was investigated using vegetative plants of Arabidopsis (Columbia) induced to flower synchronously by a single 22 h long day. Cytokinins were analysed in leaf extracts, leaf phloem exudate and in the shoot apical meristem at different times during floral transition. It was found that, in both the leaf tissues and leaf exudate, isopentenyladenine forms of cytokinins increased from 16 h after the start of the long day. At 30 h, the shoot apical meristem of induced plants contained more isopentenyladenine and zeatin than vegetative controls. These cytokinin increases correlate well with the early events of floral transition.