In vitro control of floral transition in tomato (Lycopersicon esculentum Mill.), the model for autonomously flowering plants, using the late flowering uniflora mutant.

In vitro control of floral transition in tomato (Lycopersicon esculentum Mill.), the model plant for autonomously flowering species has been investigated using the late flowering mutant uniflora (uf). Apices collected from truly vegetative plants were cultivated on solid media supplemented with different combinations of growth regulators and chemicals. Several chemical factors implicated in the promotion of floral transition of the uf mutant have been identified: sucrose, cytokinins and nitrogenous nutrients have all to be supplied at optimal concentrations. In contrast, gibberellic acid was found to be inhibitory. These results are discussed in relation to knowledge accumulated on the nature of the flowering signals circulating, at floral transition, in other plants, especially in photoperiodic species. This study suggests that tomato could constitute an adequate model to investigate the genetic and physiological control of floral transition and contribute in unravelling pathways which are constitutively regulating this important step of plant life cycle.     

Polyamine metabolism,floral initiation and floral development in Chrysanthemum (Chrysanthemum morifolium Ramat.)

In the short-day plant Chrysanthemum (Chrysanthemum morifolium Ramat. variety Pavo) putrescine and spermidine conjugates appeared in the apical bud before the first observable transformation of the meristem into floral structures. These compounds accumulated on floral initiation and well before floral evocation. Spermidine conjugates were predominant during floral initiation whereas free amines did not accumulate to any significant extent. Different associations of amides were observed during floral initiation as compared with the reproductive phase. 3,4-Dimethoxyphenethylamine conjugates (water-insoluble compounds) were the predominant amine conjugates observed during flower development. These compounds decreased drastically after fertilization. In vegetative buds from plants grown in long days polyamine conjugates were very low and appeared as plants aged. We present evidence that ornithine decarboxylase (ODC) regulates putrescine biosynthesis during floral initiation and floral development. When ODC action was blocked by DFMO (-DL-difluoromethylornithine, a specific, irreversible inhibitor of ODC), flowering was inhibited, and free and conjugated polyamines were not detected. This treatment led to a slight enhancement of ADC activity. When putrescine was added, polyamine titers and flowering were restored. A similar treatment with DFMA (-DL difluoromethylarginine, a specific, irreversible inhibitor of ADC) did not affect flowering and the polyamine titers. The results suggest that ODC and polyamine conjugates are involved in regulating floral initiation in Chrysanthemum.Abbreviations ADC arginine decarboxylase - ODC ornithine decarboxylase - DFMA -DL-difluoromethylarginine - DFMO -DL-difluoromethylornithine     

Epigenetic changes in the expression of the maize A1 gene in Petunia hybrida: role of numbers of integrated gene copies and state of methylation.

Summary ThePetunia hybrida mutant RL01 is white flowering due to a genetic block in the anthocyanin pathway. The introduction of the maize Al cDNA under the control of the CaMV 35S RNA promoter leads to the production of pelargonidin derivatives, resulting in a brick red flower phenotype. Among the transgenic petunia plants the pigmentation of the petals exhibited different expression patterns which could be categorized into the red, the variegated, and the white phenotype. This system proved to be especially suitable for the investigation of gene expression by simply looking at the pigmentation pattern of the petals. The uniformity of floral pelargonidin pigmentation is inversely correlated with the number of integrated Al copies. Furthermore, a correlation was found between the methylation status of the 35S RNA promoter and the instability of the floral pelargonidin coloration. The status of promoter methylation controlling the expression of the Al gene seems to be influenced by the copy number and the chromosomal position of the transferred gene.     

‘Leaf tip hooding’ — an induced mutation inN. Tabacum,var. Delcrest

A leaf tip hooded mutant was isolated in the progenies of the irradiated seeds of a commercial tobacco variety, namely Delcrest. The mutant character was determined as monogenic recessive. Along with leaf tip hooding, certain other characters such as increase in size of leaves, number and clasping arrangement of leaves on stem, and delayed flowering, were found to be closely associated and this was attributed to the pleiotropic effect of the leaf tip hooding gene.     

Characterization of tomato (Solanum lycopersicum L.) mutants affected in their flowering time and in the morphogenesis of their reproductive structure

The impact of the season on flowering time and the organization and morphogenesis of the reproductive structures are described in three tomato mutants: compound inflorescence (s), single flower truss (sft), and jointless (j), respectively, compared with their wild-type cultivars Ailsa Craig (AC), Platense (Pl), and Heinz (Hz). In all environmental conditions, the sft mutant flowered significantly later than its corresponding Pl cultivar while flowering time in j was only marginally, but consistently, delayed compared with Hz. The SFT gene and, to a lesser extent, the J gene thus appear to be constitutive flowering promoters. Flowering in s was delayed in winter but not in summer compared with the AC cultivar, suggesting the existence of an environmentally regulated pathway for the control of floral transition. The reproductive structure of tomato is a raceme-like inflorescence and genes regulating its morphogenesis may thus be divided into inflorescence and floral meristem identity genes as in Arabidopsis. The s mutant developed highly branched inflorescences bearing up to 200 flowers due to the conversion of floral meristems into inflorescence meristems. The S gene appears to be a floral meristem identity gene. Both sft and j mutants formed reproductive structures containing flowers and leaves and reverting to a vegetative sympodial growth. The SFT gene appears to regulate the identity of the inflorescence meristem of tomato and is also involved, along with the J gene, in the maintenance of this identity, preventing reversion to a vegetative identity. These results are discussed in relation to knowledge accumulated in Arabidopsis and to domestication processes.     

Changes of the diurnal and circadian (endogenous) mRNA oscillations of the chlorophyll a/b binding protein in tomato leaves during altered day/night (light/dark) regimes

Characteristic steady-state mRNA level oscillations were monitored for the chlorophyll a/b-binding (cab) protein in tomato plants grown under the natural day/night (light/dark) regime as well as under constant environmental conditions. This typical expression pattern was altered when plants were transferred to different light/dark regimes. For example, by shifting the light phase by six hours, a change of the time points of maximum and minimum of expression level was monitored, while the principal oscillation pattern remained the same. It appeared that the transition from dark to light is involved in determining the time points of minima and maxima of mRNA accumulation.After exposing tomato plants to an abnormal light/dark periodicity (e.g. six hours of alternating light/dark) an altered oscillation pattern was determined: within 24 hours two maxima of cab mRNA levels were detected. However, this entrained abnormal rhythm was not manifested at the molecular level and the circadian pattern reappeared under constant environmental conditions (e.g. darkness). This result favours the hypothesis that the oscillation pattern of the cab mRNA in tomato plants is not only endogenous but also hereditary.     

Fluctuation of free IAA under inductive and non-inductive photoperiods in Chenopodium rubrum

Fluctuation in levels of endogenous free IAA has been followed in the SD plant Chenopodium rubrum under photoperiodic conditions inductive or not inductive of flowering. Endogenous IAA was measured fluorimetrically as -pyrone. The level of IAA shows little fluctuation under continuous illumination. An endogenous rhythm of IAA fluctuation was found in plants transferred from light to continuous darkness, with a natural period of 30 hrs. The troughs of minimum IAA level within the endogenous rhythm coincided with the peaks in the endogenous rhythm of flowering response, which possessed the same period length. The concentration of IAA in the shoot always decreased at the end of cycles of dark period that induce flowering. The results are discussed in relation to the role of IAA in flowering of SD plants.     

Light-stimulated proton transport into the vacuoles of leaf mesophyll cells does not require energization by the tonoplast pyrophosphatase

Photosynthetic characteristics of transgenic tobacco (Nicotiana tabacum L.) plants with a soluble pyrophosphatase in the cytosol of their leaf cells were compared to those of wild-type plants. Although the development of the transgenic plants was somewhat retarded compared to the wild type, as shown by stunted growth and delayed flowering, photosynthetic responses were comparable in transgenic and wild-type leaves of similar physiological age. In particular, light-dependent proton transport into the vacuoles of leaf mesophyll cells was not decreased in leaves of the transgenic plants, which did not contain pyrophosphate in the cytosol owing to the presence of a soluble pyrophosphase. This shows that light-stimulated proton pumping did not require the pumping activity of the tonoplast pyrophosphatase. Apparently, light-stimulated proton pumping can be based solely on the activity of the tonoplast ATPase.Abbreviation CDCF 5-(and 6-)arboxy-2,7-dichlorofluorescein This work was supported within the Sonderforschungsbereiche 176 and 251 of the University of Würzburg.     

The leaf as the site of gibberellin action in flower formation in Bryophyllum daigremontianum

Summary The long-short-day plant Bryophyllum daigremontianum can be induced to flower by transfer from long to short days (LDSD), or by gibberellin (GA) application under SD. Application of GA to mature leaves of intact or partially defoliated plants induces flowering more effectively than when applications are made to the youngest leaf pair and the shoot tip.Mature leaves on de-budded plants in SD are induced to produce floral stimulus by GA application, as demonstrated by grafting LD receptor scions onto the debudded plants, or by grafting SD leaves treated with GA onto receptor stocks in LD. This shows that GA applied to Bryophyllum in SD exerts its flower-promoting effect in the leaves.The minimal number of SD necessary for flower formation in Bryophyllum is approximately 15, both in case of photoinduction by the shift LDSD, and after GA treatment in SD. It is concluded that the LD part of photinduction establishes a high level of endogenous GAs in the leaves which is a prerequisite for production of floral stimulus under subsequent SD.Work supported by the United States Atomic Energy Commission, Contract No. AT(11-1) 1338.     

Prevalence of common mutations in the arylsulphatase A gene in metachromatic leukodystrophy patients diagnosed in Britain

The frequency of two common disease-associated mutations in the arylsulphatase A (ASA) gene, and of a mutation causing ASA pseudodeficiency, have been established in metachromatic leukodystrophy patients diagnosed in our laboratory. A total of 37 mutant genes have been analysed. The GA change destroying the splice donor site of exon 2 is generally associated with more severe disease and was found in 43.2% of mutant ASA genes. The CT transition causing a proline to leucine substitution at position 426 in exon 8 (P426L) is associated with later onset disease, and was found in 16.2% of mutant genes. The AG transition leading to loss of a polyadenylation signal associated with ASA pseudodeficiency was present at a frequency of 7.5% in the patients and heterozygotes studied.     

Selection for spontaneous tomato haploids using a conditional lethal marker

We describe a method for the isolation of spontaneous haploid tomato plants from greenhousegrown seedlings obtained from crosses involving a transgenic parental line in which a counter-selectionable chimeric gene has been introduced. Transgenic seeds transformed with the aux2 gene, a gene of Agrobacterium rhizogenes that transforms naphthalene acetamide (NAM) into naphthalene acetic acid (NAA), did not develop roots in the presence of NAM, whereas wildtype tomato seeds developed a normal rooting system in its presence. Transgenic plants homozygous for aux2 (cv UC82b) were used to pollinate male-sterile (ms322) tomato plants (cv Apedice). Using NAM as a toxic substrate to kill heterozygous diploid plants carrying aux2, we selected for three maternal haploid plants resulting from the development of the female nucleus without fertilization. Maternal haploid selection using the aux2 marker was less efficient than the visual screening of haploid plants displaying recessive morphological markers of the female parent, but provided evidence for the feasibility of haploid selection in species for which no morphological markers are available.     

In search of the best correlates for local organismal biodiversity in cultivated areas

Based on a transect consisting of 19 identical trap stations in cultivated areas and seminatural habitats, the correlation of species numbers of higher taxonomic groups with total species numbers of flowering plants and arthropods per trap site was calculated. A total of 191214 invertebrate specimens and 2221 species of plants and animals were analysed. Considering the value of the correlation coefficient R2 as well as the effort for sorting and identification, a top twenty list of indicator groups favours Heteroptera, flowering plants, Symphyta and aculeate Hymenoptera as the best choice for biodiversity evaluation. In general, flight traps rated better than pitfall traps. In most taxonomic groups, diversity indices such as the Shannon and the Simpson index were only weakly correlated with local species diversity.     

Effects of sugar on vegetative development and floral transition in Arabidopsis

Although sugar has been suggested to promote floral transition in many plant species, growth on high concentrations (5% [w/v]) of sucrose (Suc) significantly delayed flowering time, causing an increase in the number of leaves at the time of flowering in Arabidopsis. The effect of high concentrations of Suc seemed to be metabolic rather than osmotic. The delay of floral transition was due to extension of the late vegetative phase, which resulted in a delayed activation of LFY expression. In addition, growth on low concentrations (1% [w/v]) of Suc slightly inhibited flowering in wild-type plants. This delay resulted from effects on the early vegetative phase. This inhibition was more pronounced in tfl1, an early flowering mutant, than in the wild type. Although 1% (w/v) Suc was reported to promote floral transition of late-flowering mutants such as co, fca, and gi, floral transition in these mutants was delayed by a further increase in Suc concentration. These results suggest that sugar may affect floral transition by activating or inhibiting genes that act to control floral transition, depending on the concentration of sugars, the genetic background of the plants, and when the sugar is introduced. Growth on 1% (w/v) Suc did not restore the reduced expression levels of FT and SOC1/AGL20 in co or fca mutants. Rather, expression of FT and SOC1/AGL20 was repressed by 1% (w/v) Suc in wild-type background. The possible effects of sugar on gene expression to promote floral transition are discussed.     

Regulation of expression of a wound-inducible tomato inhibitor I gene in transgenic nightshade plants

A wound-inducible proteinase Inhibitor I gene from tomato containing 725 bp of the 5 region and 2.5 kbp of the 3 region was stably incorporated into the genome of black nightshade plants (Solanum nigrum) using an Agrobacterium Ti plasmid-derived vector. Transgenic nightshade plants were selected that expressed the tomato Inhibitor I protein in leaf tissue. The leaves of the plants contained constitutive levels of the inhibitor protein of up to 60 g/g tissue. These levels increased by a factor of about two in response to severe wounding. Only leaves and petioles exhibited the presence of the inhibitor, indicating that the gene exhibited the same tissue specificity of expression found in situ in wounded tomato leaves. Inhibitor I was extracted from leaves of wounded transformed nightshade plants and was partially purified by affinity chromatography on a chymotrypsin-Sepharose column. The affinity-purified protein was identical to the native tomato Inhibitor I in its immunological reactivity and in its inhibitory activity against chymotrypsin. The protein exhibited the same M _r of 8 kDa as the native tomato Inhibitor I and its N-terminal amino acid sequence was identical to that of the native tomato inhibitor I, indicating that the protein was properly processed in nightshade plants. These expriments are the first report of the expression of a member of the wound-inducible tomato Inhibitor I gene family in transgenic plants. The results demonstrate that the gene contains elements that can be regulated in a wound-inducible, tissuespecific manner in nightshade plants.     

Polyamines,floral induction and floral development of strawberry (Fragaria ananassa Duch.)

In the short-day plant, strawberry (Fragaria ananassa Duch.), polyamines (putrescine, spermidine and spermine), conjugated spermidine (water-insoluble compounds) and bound amines (putrescine, spermidine, phenylethylamine, 3-hydroxy, 4-methoxyphenylethylamine) accumulated in the shoot tips during floral induction and before floral emergence. Different associations of free amines and conjugated amines were observed during floral induction, as compared with the reproductive phase. During the whole period of floral development, phenylethylamine (an aromatic amine) was the predominant amine, representing 80 to 90% of the total free amine pool. Phenylethylamine conjugates (water-insoluble compounds) were the predominant amides observed prior to fertilization. These substances decreased drastically after fertilization. In vegetative shoot tips from plants grown continously under long days, free polyamines (putrescine, spermidine) and bound polyamines (putrescine, spermidine) were low and no change was observed. Free amines (spermine and phenylethylamine), bound aromatic amines (phenylethylamine, 3-hydroxy, 4-methoxyphenylethylamine), conjugated spermidine and phenylethylamine did not appear. Male-sterile flowers were distinguished by their lack of conjugated spermidine and phenylethyalamine and by a decrease in free phenylethylamine. In normal and sterile strawberry plants -DL-difluoromethylornithine (DFMO), a specific irreversible inhibitor of ornithine decarboxylase (ODC), caused inhibition of flowering and free and polyamine conjugates. When putrescine was added, polyamine titers and flowering were restored. A similar treatment with -DL-difluoromethylarginine (DFMA), a specific, irreversible inhibitor of arginine decarboxylase (ADC), did not affect flowering and polyamine titers. These results suggest that ornithine decarboxylase (ODC) and polyamines are involved in regulating floral initiation in strawberry. The relationship between polyamines, aromatic amines, conjugates, floral initiation and male sterility is discussed.Abbreviations ADC arginine decarboxylase - ODC ornithine decarboxylase - DFMA -DL-difluoromethylarginine - DFMO -DL-difluoromethylornithine - Put putrescine - Spd spermidine - Spm spermine - Phen phenylethylamine - 3H4M Phen 3-hydroxy, 4-methoxyphenylethylamine     

A genetically related response to iron deficiency stress in muskmelon

A mutant muskmelon (Cucumis melo L.) with characteristic Fe-deficiency chlorosis symptoms was compared to related cultivars in its ability to obtain Fe via the widely known Fe-stress response mechanisms of dicotyledonous plants. The three cultivars (fefe, the Fe-inefficient mutant; Mainstream and Edisto, both Fe efficient plants) were grown in nutrient solution in either 0 or 3.5 mg L^-1 Fe as FeCl₃. None of the three cultivars released reductants or phytosiderophores, but both Edisto and Mainstream produced massive amounts of H+ ions to reduce and maintain the pH of nutrient solutions below pH 4.0. The roots of these two Fe-efficient cultivars were also capable of reducing Fe³⁺ to Fe²⁺. These responses maintained green plants, resulted in high leaf Fe in both Edisto and Mainstream, and produced Mn toxicity in Mainstream. The lack of Fe-deficiency stress response in fefe not only affected leaf Fe concentration and chlorosis, but also resulted in reduced uptake of Mn. The importance of reduced Fe (Fe²⁺) to the Fe-efficient cultivars was confirmed by growing the cultivars with BPDS (4, 7-diphenyl-1, 10-phenanthroline disulfonic acid, a ferrous chelator) and EDDHA [ethylene-diamine di (0-hydroxphenylacetic acid)] (a ferric chelator), and observing increased chlorosis and reduced Fe uptake in BPDS grown plants. The Fe-deficiency response observed in these cultivars points out the diversity of responses to Fe deficiency stress in plants. The fefe mutant has a limited ability to absorb Fe and Mn and perhaps could be used to better understand Mn uptake in plants.     

High levels of ripening-specific reporter gene expression directed by tomato fruit polygalacturonase gene-flanking regions

The 1.4 kb 5 polygalacturonase (PG) gene-flanking region has previously been demonstrated to direct ripening-specific chloramphenicol acetyl transferase (CAT) expression in transgenic tomato plants. The steady state level of CAT mRNA in these plants was estimated to be less than 1% of the endogenous PG mRNA. Further constructs containing larger PG gene-flanking regions were generated and tested for their ability to direct higher levels of reporter gene expression. A 4.8 kb 5-flanking region greatly increased levels of ripening-specific reporter gene activity, while a 1.8 kb 3 region was only shown to have a positive regulatory role in the presence of the extended 5 region. Transgenic plants containing the CAT gene flanked by both of these regions showed the same temporal pattern of accumulation of CAT and PG mRNA, and steady-state levels of the transgene mRNA were equivalent to 60% of the endogenous PG mRNA on a per gene basis. The proximal 150 bp of the PG promoter gave no detectable CAT activity. However, the distal 3.4 kb of the 4.8 kb 5 PG promoter was shown to confer high levels of ripening-specific gene expression when placed in either orientation upstream of the 150 bp minimal promoter. The DNA sequence of the 3.4 kb region revealed a 400 bp imperfect reverse repeat, and sequences which showed similarity to functionally significant sequences from the ripening-related, ethylene-regulated tomato E8 and E4 gene promoters. The possible roles of the flanking regions in regulating PG gene expression are discussed.     

The Interaction of Plant Growth Regulators and Vernalization on the Growth and Flowering of Cauliflower (Brassica oleracea var. botrytis)

The growth and flowering response of a cold-requiring cauliflower (Brassica oleracea var. botrytis cv. 60 day) to a range of temperatures under 10 h photoperiod and to growth regulator application were investigated. Endogenous gibberellin A₁(GA₁) concentrations were also assessed under these treatments. Flowering and growth of the inflorescence stalk were correlated with plant developmental stage at the time of a vernalizing cold treatment. Temperature and its duration also affected flowering and inflorescence development. The most effective temperature for inflorescence induction was 10 °C. Flowering did not occur in non-vernalized plants (25 °C) even though they had been treated with GA₃. Application of GA₃ promoted inflorescence stalk elongation greatly in vernalized plants (10 °C), but less so in partially vernalized plants (15 °C or 20 °C). Paclobutrazol (PP₃₃₃) sprayed at the 8–9 leaf stage significantly suppressed inflorescence stalk length and slightly delayed flower bud formation and anthesis. Vernalization at 10 °C increased endogenous GA₁ content in both leaves and the inflorescence stalk irrespective of GA₃ or PP₃₃₃ treatment. Application of GA₃ tended to increase GA₁ levels, while PP₃₃₃ significantly reduce GA₁, both irrespective of vernalization. Vernalization is an important factor for flowering, but not curd formation in this cauliflower cv. 60 day and GA₁ is likely a causal factor in inflorescence stalk elongation.     

The ichneumon wasp Venturia canescens: Oviposition and avoidance of superparasitism

The parthenogenetic ichneumon wasp Venturia (= Nemeritis( canescens (Grav.) carries a single egg in readiness for oviposition in a cavity right at the tip of the long ovipositor. The egg is passed into the cavity during a flexing or cocking movement of the abdomen, and this behaviour can be used to determine whether or not Venturia has deposited an egg into its host. The cocking movement shows that Venturia tends to avoid superparasitism.

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Résumé l'Ichneumonide parthenogénétique Nemeritis canescens porte un seul uf prêt à être pondu dans une cavité fusiforme, juste à la pointe de son long ovipositeur. L'uf est éventuellement émis quand l'ovipositeur pénètre dans une chenille hôte. L'uf chemine le long de l'ovipositeur, et est mis en place dans la cavité apicale pendant un mouvement de flexion très caractéristique de l'abdomen (Cocking Fig. 2) qui est bien distinct du mouvement accompli au moment même de la piqûre de l'hôte (Stabbing Fig. 2).Le mouvement dit d'armement (Cocking) est toujours effectué juste après la ponte dans un hôte sain, assurant ainsi la mise en place d'un nouvel uf pour une autre piqûre. L'absence du mouvement de Cocking entre deux actes de piqûre dans deux hôtes successifs indique que la piqûre dans le premier hôte n'a pas été suivie de ponte.En utilisant le mouvement de Cocking comme indicateur de ponte on peut démontrer que Nemeritis reconnaît la présence d'un uf dans une hôte cinq minutes après que celui-ci a été parasité, et évite le superparasitisme en effectuant une piqûre non suivie de ponte. La proportion de piqûres stériles s'accroît quand l'hôte héberge un uf dont le développement est plus avancé, l'évitement du superparasitisme atteint sa plus forte proportion 30 minutes seulement après le dépôt du premier uf (Fig. 3).