Rates of Growth and Cell Division in the Shoot Apex of Silene During the Transition to Flowering

The growth rates of the shoot apex during and after floral inductionwere measured in Silene, a long-day plant. Plants were inducedto flower with 4 or more long days (LD) but not with 3 longdays or with short days (SD). The rate of increase of cell numberin the apical dome, above the youngest leaf pair, was exponentialand in plants given 3 LD remained the same as in plants in SD.In plants induced to flower with 7 LD, until the end of theinductive period the rate of increase of cell number in theapical dome remained the same as in plants in SD. Only whenthe apex began to enlarge as the first stage in the formationof the flower did the growth rate of the apical dome increase.The rates of increase of cell numbers in the apex correspondedto mean cell generation times of 20 to 33 h for plants in SD,for plants given 3 LD, and during the 7 days of induction forplants given 7 LD, and 6 to 8 h for induced plants when flowerformation was beginning. The distribution of cell division in the apex was examined bytreating plants with colchicine and noting in sections the positionsof the resulting metaphases. In vegetative apices and also inapices undergoing transition to flowering the whole of the apicaldome appeared to consist of cells dividing at a similar rate. The rate of leaf initiation during induction was the same asin vegetative, non-induced plants.     

Growth changes in the shoot apex of Sinapis alba during transition to flowering

The aim of the work was to report morphological changes whichoccur in the shoot apex during the morphogenetic switch to floweringin the model long day (LD) plant, Sinapis alba. During the floraltransition induced by 1 LD the growth rate of all componentsof the shoot apex is modified profoundly. The earliest changes,detected at 24 h after start of LD, include a decrease in plastochronduration and an increase of growth of leaf primordia. One daylater, the meristem dome starts to increase in volume, apicalinternodes have an increased height and there is a precociousoutgrowth of axillary meristems. All these changes precede initiationof flower primordia, which starts at about 60 h after the startof LD. Later changes include meristem doming, a decrease inthe plastochron ratio and a shift to a more complex phyllotaxis.All the changes, except the decreased plastochron ratio, arecharacteristics of an apex with an increased tempo of growth.The stimulation of longitudinal growth (height of apical intemodes)is more marked and occurs earlier than the reduction of radialgrowth (plastochron ratio). Key words: Axillary meristem, internode growth, leaf growth, plastochron ratio, plastochron duration     

Changes in mRNA level rhythmicity in the leaves ofSinapis alba during a lengthening of the photoperiod which induces flowering

A previous study has shown that mRNAs exhibit complex patterns of diurnal rhythms in their quantity in the leaves ofSinapis alba during an 8 h light/16 h dark short day (SD). In order to determine whether this situation is rapidly modified in plants subjected to an extended light treatment, we have usedin vitro translation and two-dimensional polyacrylamide gel electrophoresis, together with a strict gel comparison procedure giving aP=0.03 certitude level, to analyse the mRNA complement at different times during a 22 h light/2 h dark long day (LD).During this LD, complex changes affected about 10% of the mRNAs. Thirty-four different patterns were observed. Some diurnal rhythms present in SD are not modified by the lengthening of the light period, but most are affected. Moreover, we have shown that some mRNAs presenting a constant quantity under a SD regime show an increase or a decrease during the first hours of the photoperiod lengthening.InSinapis, this LD also induces flowering. All the changes in mRNA quantity detected thus parallel the photoperiodic induction of flowering in the leaves and are quantitative; no mRNA was shown to appear or to disappear.     

Changes to Proteins in the Shoot Meristem of Silene coeli-rosa during the Transition to Flowering

Changes in proteins were measured to test whether they werelinked to flowering, the cell cycle or both in Silene coeli-rosashoot apices. Seeds were germinated and grown at 20?C in shortdays (SD) of 8 h light from fluorescent and tungsten (F + T)/16hdarkness for 28 days (day 0). Plants were then exposed to: 7long days (LD)+2 SD (inductive), 7 LD + 48 h darkness (inductive)or 7 dark-interrupted (di) LD + 2SD (non-inductive), where eachLD and diLD comprised 8 h F + T/16 h T, 8 h F+T/l h darkness/15h T, respectively. There were no qualitative differences inpolypeptide composition in the LD and SD treatments on days0, 5, 6 and 7 but 60 new polypeptides were detected on LD₈ andsome modifications in the intensity of common spots also occurred.These qualitative and quantitative changes were not alteredfollowing 7 LD + 48 h darkness, which is known to suppress synchronisationof the cell cycle but not flowering. Thus, changes on day 8are linked to flowering. Transient increases in protein perunit area were detected in prophase cells on days 0, 3, 4 whilesustained increases occurred on days 7 and 8. The changes ondays O, 3 and 4 were suppressed by the diLD treatment. Thus,quantitative changes to proteins, co-inciding with known LD-inducedchanges to the cell cycle, are linked to flowering. (Received April 2, 1990; Accepted September 12, 1990)     

Purification, Characterization and cDNA Cloning of the 200 kDa Protein Induced by Cold Acclimation in Wheat

We have purified to homogeneity the 200 kDa protein inducedspecifically by low temperature in wheat (Triticum aestivumL.). The boiling solubility of the protein has been used asa main step in the purification procedure. Amino acid compositionindicates that the 200 kDa has a compositional bias for glycine(11.4%), threonine (13.3%), and alanine (22.0%). Using oligonucleotideprobes, we have isolated a clone (pWcs200) from a cold-acclimatedwinter wheat cDNA library. Northern analysis demonstrated thatthe expression of the corresponding gene was specifically upregulatedby low temperature. Southern analysis showed that the gene organizationand the relative copy number were identical in two cultivarsdiffering in their capacity to develop freezing tolerance. Proteinsequence and immunological analyses indicate that this proteinshares similar features with the 50 kDa protein induced duringcold acclimation of wheat. The two proteins are boiling-soluble,and possess similar repeated elements. These elements may beimportant for the development of freezing tolerance. We haveshown that the 200 kDa protein is the largest member of a familyof immunologically-related cold-induced proteins in wheat. Expressionof pWcs200 in E. coli yielded a product of around 200 kDa, indicatingthat the clone contains most of the coding region for this protein. (Received August 18, 1992; Accepted October 14, 1992)     

Changes in the pattern of protein synthesis during differentiation of the Ascomycete Sphaerostilbe repens

The vegetative mycelium of Sphaerostilbe repens Berkeley and Broome (strain CBS 275-60) gives rise, within 48 h, to aggregated organs composed of coremia and rhi-zomorphs. Developmental changes in polypeptide patterns were studied by one- and two-dimensional polyacrylamide gel electrophoresis after cells had been induced to undergo synchronized differentiation. One-dimensional gel electrophoresis revealed only minor changes during the morphogenesis. Of the 300 polypeptides resolved by two-dimensional gel electrophoresis, nearly 12% either increased or decreased during coremium and rhizomorph differentiation. Some polypeptides appeared to be unique to one or the other of the cell preparations and represented apparent qualitative differences. During the first 24 h of differentiation, about 20 polypeptide spots appeared, 6 were enhanced, 4 were reduced and 32 disappeared. Over the next 24 h changes in the population of proteins were less marked: 14 new proteins were revealed and 9 increased in intensity while 15 declined and 9 were no longer detectable. Five proteins which were present at a significant level only during the first stages of differentiation, may therefore, putatively be designated as aggregation-specific polypeptides.     

Seed Germination in Amaranthus retroflexus L. as Affected by the Photoperiod and Age During Flower Induction of the Parent Plants

Seed germination in Amaranthtis retroflexus, a facultative shortday plant, was affected by the parental photoperiodic conditions.Seeds from parents grown continuously in short days (SD, 8 h)had a higher dark germination and a greater response (at 30°C) to a short irradiation or low temperature pretreatmentthan seeds from plants grown continuously in long days (LD,16 h). Daily night breaks of 1 h in the middle of the long-nightinhibited the SD induction of flowering as well as the SD promotionof germinability. Germinability of seeds produced by plantsinduced to flower in LD by 1, 2, or 3 SD was lower than thatof seeds produced by plants grown continuously in SD, and decreasedwith the age of the parent plants at the time of flower induction.     

Cell Cycle Changes in the Shoot Apex of Silene coeli-rosa During the Second and Third Days of Floral Induction

The cell cycle in Silene coeli-rosa shoot apices was measuredto test whether or not early components of the floral stimulus,produced during the 2nd and 3rd long days (LD) of an inductiveLD treatment, resulted in an increase in the duration of G₂phase in constant 20–24 h cell cycles. Plants were grownat 20°C in short days (SD) of 8 h light and 16 h darknessfor 28 d (day 0). Starting on day 0, plants were given SD or3 LD each comprising an identical 8 h day and 16 h photo-extension,or 3 dark-interrupted (d.i.) non-inductive LD, interrupted at1700 h of each day with 1 h of darkness. The cell cycle (percentagelabelled mitoses method) and changes in cell number were determinedin the shoot apical meristem. During days 1–2 of the SDtreatment, the cell cycle and mean cell generation time (MCGT)was 18 and 32 h, respectively, giving a growth fraction of 56%.During days 2–3, the cell cycle and MCGT shortened to15 and 23 h, respectively (growth fraction = 65%). During days1–2 of the LD and d.i. LD treatments, cell cycles andMCGTs were 9–10 and 27–29 h, respectively, resultingin smaller growth fractions (about 33%). Thus, shortened cellcycles and altered growth fractions occurred regardless of whetheror not the treatment was inductive. The LD treatment resultedin a marked shortening of G₁ and, to a lesser extent, S-phase,whilst G₂ remained constant. These changes were consistent withincreases in the proportion of cells in G₂ during the photoextensionof each LD which were suppressed during the comparable periodsof the d.i. LD treatment. The latter treatment resulted in eachphase occupying virtually identical proportions of the cellcycle as in the SD treatment. Thus, the unique cell cycle responsesto the initial part of the inductive LD treatment were increasesin the proportion of cells in G₂ coupled with G₁ and G₂ beingof similar duration. Cell cycle, mean cell generation time, shoot apex, Silene coeli-rosa     

Early Increase in the Mitotic Index in the Shoot Apex of Lolium temulentum cv. Ceres During the Floral Transition

Six-week-old Lolium temulentum cv. Ceres plants were inducedto flower by a single long day (Day 1). Cell proliferation wasanalysed in the apex during the period extending from Day 1to Day 3. A stimulation of mitotic and DNA synthetic activitieswas observed in the apical summit at 8 h of the photoextensionperiod of the LD, i.e. before any apparent movement of the floralstimulus out of the leaves and well in advance of apex evocation. Key words: Cell cycle, flowering, Lolium, shoot apex     

Differences in Graft Transmission of the Floral Stimulus in Two Species of Kleinia

Graft transmission of the floral stimulus was studied in homograftsof Kleinia articulata (SDP) and heterografts between K. articulataand K. repens (LSDP). While receptor shoots of K. repens graftedonto induced donor plants of K. articulata flowered readilyin LD (16 h) as well as SD (8 h), graft-induced flowering failedto take place in LD receptors of K. articulata. Neither theinduced shoots and detached leaves from induced plants of K.articulata nor the induced shoots of K. repens could evoke theflowering response in the K. articulata receptors. Increasingthe donor pool of induced leaves even up to ten per receptoralso had no effect. It is known that the very young leaf primordiaof K. articulata are photoperiodically sensitive, and it seemsthat they may prevent the stimulus from reaching the apex. Key words: Kleinia, Flowering stimulus, Graft transmission     

Nuclear Proteins During the Onset of Cell Proliferation in Pea Root Meristems

The DNA: proteins ratio in nuclei of root meristems of pea (Pisumsativum L. cv. Lincoln) changed considerably during germinationas cells moved from a quiescent to an actively proliferatingstate with a higher protein content in nuclei of the latter.Electrophoretic patterns of nuclear proteins extracted at differenttimes of germination were used to examine qualitative changes.The various patterns presented a substantial similarity butthere were some proteins whose content increased or decreasedand others which disappeared or appeared during germination.The bulk of these variations occurred between 24 and 48 h ofgermination, suggesting that they might be correlated to thetransition from quiescence to the proliferating state. The patternof nuclear proteins obtained from adult differentiated roottissue was also examined. We tried to purify five differentnuclear protein components from intact nuclei by a multi-stepextraction procedure using a series of different buffers toascertain the nature of proteins presenting major interestingvariations. Most of these proteins purified with the nuclearsap or ribosomal components. Key words: Cell proliferation, electrophoresis, Pisum sativum L, root meristems     

Increased Pentose Phosphate Pathway Activity Linked to Floral Induction in Apices of Spinacia oleracea during Short Days

A quantitative cytochemical study of glucose-6-phosphate dehydrogenaseactivity as a marker of the pentose phosphate pathway (PPP)was made on shoot apices of Spinacia oleracea kept either continuouslyin short days for up to eight weeks or transferred for a singleperiod of 20 h to continuous light after between three and eightweeks in short days. By the time the apices kept in continuousshort days showed morphological changes relating to the floralstate, the PPP activity was already elevated. From four weeksonwards, the apices were more readily induced to the floralstate as evidenced by the increased PPP activity. In addition,the level of PPP activity achieved in the short-day apices asthey progressed to the floral state was as great as that observedin the apices induced to flower by a 20-h day. Spinacea oleracea, pentose phosphate pathway, shoot apex, cytochemistry, glucose-6-phosphate dehydrogenase, floral induction     

Periodicity of Pollen Development and Quantitative Cytochemistry of Exine and Intine Enzymes in the Grasses Lolium perenne L. and Phalaris tuberosa L.

Daily analysis of anther samples during flower development hasenabled an estimation of the duration of defined developmentalperiods in pollen of the grass Phalaris tuberosa. A similarsequence of pollen development has been established for ryegrass,Lolium perenne, where changes in activity of wall enzymes havebeen followed using quantitative cytochemical methods. Acidphosphatase, an intine enzyme, showed two periods of activity:during the vacuolate period corresponding to deposition of theintine polysaccharides; and in the maturation period correspondingto cytoplasmic activity. Non-specific esterase showed greatestactivity in the parietal tapetal cells until their dissolutionearly in the vacuolate period when an increase in pollen-associatedactivity occurred. These changes provide additional evidencefor the transfer of tapetal proteins to exine sites. Lolium perenne L., Phalaris tuberosa L., ryegrass, canary grass, pollen development, quantitative cytochemistry, enzyme activities, acid phosphatase, esterase     

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.     

Flowering requirements in Bromus inermis, a short-long-day plant

Smooth bromegrass plants ( Bromus inermis Leyss.) have a dual photoperiodic requirement for flowering. At temperatures ranging from 6 to 24°C, short days (SD) are necessary for primary induction while a transition to long days (LD) is required for initiation of flower primordia, culm elongation and flower development (secondary induction). Critical photoperiods for primary induction (50% flowering) were 13.5 h (15°C) and 12 h (24°C) in the American cv. Manchar and 14.5 and 13 h, respectively, in the Norwegian cv. Löfar. For the secondary induction the respective critical photoperiods were 14 and 15 h in 'Manchar' and 16 and 17.5 h in 'Löar', which also appeared to be better adapted to low temperatures. Low temperature vernalization in LD for up to 16 weeks at 3°C was unable to cause primary induction and temperatures below 12°C also strongly reduced the SD effect. At optimum temperature (15-2TC) 4 to 6 weeks of 8-10 h SD treatments were needed for optimal primary induction effect. A minimum of 8 LD cycles of 24 h were required for complete secondary induction in 'Manchar', while more than 16 cycles were needed in 'Löfar'. Seedlings grown in SD developed a rosette type of growth with shoots growing in a decumbent position, while those in LD grew upright and formed elongated vegetative culms. Rate of leaf initiation was enhanced by about 60% by LD while tillering was promoted by SD.     

Proteins Induced by Physical Stimuli in Root Tips of Arabidopsis thaliana Seedling

Proteins newly synthesized in cells of root tips of Arabidopsisseedlings after gravistimulation and photo-induced tactile stimulationwere analyzed by two-dimensional gel electrophoresis. Intensitiesof two out of about 600 protein spots were observed to increasetransiently when culture flasks in which seedlings has beengrown were kept on their sides. When the flasks were kept verticalon a rocking table and rocked continuously for 24 hours, intensitiesof ten protein spots increased, and four spots appeared forthe first time. Analysis of [³²P]-labeled proteins revealedthat the continuous rocking treatment enhanced the phosphorylationof proteins in two spots. When the seedlings in flasks wereilluminated from the front, and the roots bent towards the backwall of the flasks, three spots appeared for the first timeand intensities of nine spots were enhanced. Three of the twelvespots whose intensities were enhanced by the photo-induced tactilestimulation were also affected by continuous rocking treatment.The roles of protein synthesis and phosphorylation in the pathwaysbetween the stimuli and the responses are discussed. (Received June 18, 1992; Accepted December 16, 1992)     

Analysis of In Vivo Protein Synthesis and Histological Studies of Haustorial Formation in Root Cultures of Witch weed (Striga asiatica L Kuntze)

We recently described an in vitro approach that uses root culturesto study haustorial formation in Striga asiatica. Previous studieshave used haustoria formed on intact radicles of Striga seedlings.In vitro cultured roots formed haustoria that appeared morphologicallysimilar to those formed by Striga radicles, but were 5–10-foldlarger. In this study, we provide biochemical and histologicalevidence to support further the similarity of root culture haustoriato haustoria formed on radicles of seedlings. We examined invivo protein synthesis during haustorial development on rootcultures and radicles by 2-D PAGE. Four proteins increased inabundance in both root cultures and radicles after 6 h of haustorialinduction. All four proteins appeared transiently in root culturesand radicles, being more abundant at 6 h, and less abundantafter 24 h of haustorial induction. Only three of the four haustorial-specificproteins were more abundant in root cultures after 2 h of haustorialinduction; all four had decreased in abundance after 12 h ofhaustorial induction. Using light microscopic analysis we comparedthe ontogeny of root culture haustoria to that of haustoriaon radicles. These studies revealed that root culture haustoriaundergo developmental changes similar to those reported forradicle haustoria such as early expansion of cortical cells,the emergence of haustorial hairs from epidermal cells, andthe development of densely staining cells at the haustorialapex. In addition, these changes occurred within a similar time-frameand sequence in root culture and radicle haustoria. Finally,root culture haustoria were found to be capable of attachingto sorghum host roots. Key words: Striga asiatica L., Kuntze, haustoria, root cultures, proteins, histology, 2D-PAGE     

Changes in protein composition of the shoot meristem during floral evocation in Sinapis alba

Shoot apical meristcms of vegetative and induced plants of Sinapis alba L. were labelled with [³⁵S] methioninc for 2 h and the proteins were then separated by isoelectric focussing and polyacrylamide gel electrophoresis. Quantitative and possibly qualitative changes in the complement of proteins being synthesised during evocation were detected in the meristem, distal to the primordia, 50 to 52 h after the beginning of the inductive long day. This was before morphological changes in the meristem, and before the initiation of flower bud primordia.     

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     

Intergeneric Translocation of Floral Stimulus across a Graft in Monoecious Cucurbitaceae with Special Reference to the Sex Expression of Flowers

The effect of floral stimulus on flower sex expression in monoeciouscucurbits was examined using a qualitative short-day plant,Sicyos angulatus L. Sicyos was induced to flower not only bygrafting it onto a flower-induced plant of the same speciesbut also by intergeneric grafting Onto the day-neutral plantCucumis sativus L. or the quantitative short-day plant Luffacylindrica Roem under noninductive long-day conditions. Sicyosplant grafted onto various cucumber varieties having differentgenetic backgrounds for their sex expression developed bothstaminate and pistillate inflorescences with similar sex expression.When the availability of floral stimulus was restricted as inthe case of grafting of Sicyos onto young cucumber seedlingsat the cotyledonary stage, most inflorescences appearing onthe Sicyos were staminate ones. Pistillate flowers formed onthe cucumber receptors substantially increased when they weregrafted onto Sicyos donors which had a sufficient number ofleaves induced by short-days as compared with those graftedonto noninduced ones. These results suggest that the availabilityof floral stimulus participates in the sex expression of flowersin Cucurbitaceae. Undeveloped pistillate inflorescences, whichoccasionally appear on Sicyos scion, flowered normally whenN⁶-benzylaminopurine was directly applied to the inflorescence. (Received February 27, 1981; Accepted October 16, 1981)