J. Bianco

Dormancy-breakage in seeds of Douglas fir (Pseudotsuga menziesii (Mirb.) Franco). Support for the hypothesis that LEA gene expression is essential for this process

Summary In Douglas fir seeds ( Pseudotsuga menziesii (Mirb.) Franco), dormancy-breakage is associated with quantitative and qualitative changes in gene expression. Specific examples of genes that are up-regulated during dormancy-breakage include those encoding LEA proteins (DF6, DF65 and DF77) (Jarvis et al., J. Plant Physiol. 147: 559–566, 1996). In particular, DF65, a dehydrin, was specifically expressed during stratification and DF6 and DF77 were significantly up-regulated. On examination of the tissue distribution of the LEA gene transcripts during chilling (which breaks dormancy) and warm-incubation, LEA transcripts were detected in both embryo and megagametophyte tissues. Expression of DF65 was lower, and the transcript appeared later, in the embryo than in the megagametophyte. With DF6 and DF77, transcripts reached maximal levels in the megagametophyte earlier than in the embryo. While de novo synthesis of ABA was important for the expression of dormancy, four weeks of chilling, only led to a 36% reduction in endogenous ABA and did not change its distribution between the embryo and megagametophyte, compared with warm-incubated seeds. However, following chilling, seeds showed a reduced sensitivity to applied ABA. To determine if there was a link between the expression pattern of LEA genes and the dormancy status of the seeds and/or abiotic stresses, seeds were treated with the plant growth regulators ABA and Me-JA, or subjected to osmotic, salt, and heat stresses. ABA treatment prevented germination of non dormant seeds, and did not induce expression of LEA genes. Me-JA stimulated dormant seeds to germinate and induced low level expression of all three LEA genes after 24 hours, but not after one week. Expression of all three LEA genes was detected in seeds which had been subjected to osmotic and salt stresses expression. However, under heat stress only DF6 and DF77 were expressed. The degree of induction by these treatments was much less than that induced by chilling. We conclude that the primary role of these gene products is connected with changes that occur during the cold, moist treatment that leads to dormancy-breakage in seeds.

Endogenous cytokinins in rose petals and the effect of exogenously applied cytokinins on flower senescence

In extracts from rose petals cytokinin activity was detected by Amaranthus bioassay in HPLC eluates corresponding to the standards: Z, ZR, 2iP and 2iPA; subsequently, the presence of two groups of endogenous cytokinins was confirmed by ELISA.Measurements of senesence indicators (cell sap osmolarity and conductivity) and observations of flower vase-life indicated that when the above cytokinins were applied as holding solutions they delayed flower senescence by 34–56% and prolonged rose longevity.

De novo ABA synthesis and expression of seed dormancy in a gymnosperm: Pseudotsuga menziesii

Seeds of dormant Douglas-fir seeds germinated poorly when they were cultivated at 20–23 °C while isolated embryos germinated fully within two weeks. Seed dormancy was therefore imposed on the embryo by its surrounding structures. This physiological behaviour was well correlated with changes in ABA level during culture. Indeed, the ABA level decreased in isolated embryos while it increased in both embryo and megagametophyte during culture of whole seeds. The origin of this increase was analysed and the different ways by which seed coats could interfere with ABA accumulation are discussed.

Alteration of hormonal levels and hormone sensitivity by ri T-DNA-transformation of apple cuttings

Summary Changes in hormone levels and sensitivity to hormones were studied in the apple rootstock M 26 and in the clone transformed by Agrobacterium rhizogenes strain A4. The transformation of apple microcuttings by insertion of both TL-DNA and TR-DNA led to changes in leaf levels of all plant hormones studied (auxin, cytokinins and abscisic acid). An important increase in auxin and a decrease in cytokinin levels were observed. The ABA level was substantially lowered by transformation, whatever the water status of the leaves. The sensitivity to hormones of both transformed and untransformed cuttings was compared on the basis of their root induction efficiency. No significant difference in auxin sensitivity could be observed on this long-term response. However, the transgenic clone was less sensitive to cytokinin-induced rooting inhibition and more sensitive to ABA-induced rooting inhibition than the untransformed clone. The consequences of the changes in both hormone levels and sensitivity to hormones are discussed.

ABA Involvement in the Psychrolabile Dormancy of Fagus Embryo

Embryos of Fagus sylvatica isolated from fruits at harvest were dormant. After a cold-treatment in restricted water conditions, embryos were able to germinate at a percentage which increased with chilling duration. Embryo ABA content decreased during the dormancy-releasing treatment at 4°C; it also decreased in the same proportions during embryo culture at 23°C, a temperature allowing dormancy to be strongly expressed. It thus appears that embryo ABA content was not correlated with the physiological potentialities of the embryo. The significant decrease in ABA level observed during culture at 23°C could be explained by a very rapid ABA metabolism, revealed by the results of [3H]ABA feeding experiments. Also, when fluridone, an inhibitor of carotenoid synthesis, was applied directly to axes, dormant embryos were able to germinate after a one-week culture. The comparison between axis ABA content in the absence or in the presence of fluridone after a 6-day culture gave an estimation of the axis capacity for a de novo ABA synthesis. Consequently, it appears that the psychrolabile dormancy of Fagus is associated with the ability of the axis to synthesize its own ABA.

Gibberellins in Dormant Embryos of Pyrus malus L. cv Golden delicious.

The aim of this work was to search for free gibberellins in dormant embryos of Pyrus malus L. cv Golden delicious using several extraction procedures. Quantitative estimations were made by biotest (lettuce hypocotyle test) and by GLC. Using an ethanolic extraction, only minute quantities of free gibberellins were detected. When extraction was carried out in the Tris buffer pH 7.2, the quantities of gibberellins detected were slightly higher. When the embryos were crushed in Tris buffer and treated with Triton X 100, very large amounts of gibberellins were detected, especially GA1 and GA4. These results indicate that gibberellins are present in dormant embryos before chilling treatment. The form in which the gibberellins exist and the origin of GA4 are discussed.

ABA CONTENTS AND THE REGENERATION ABILITY OF FRITILLARIA IMPERIALIS L. CULTURED IN VITRO

Correlation between ABA content in “mother” tissue and subsequent regeneration ability of Fritillaria imperialis cultured in vitro was investigated. In every experiment regeneration was always most efficient from plant material containing the lowest amount of free ABA: a leafy stem part, bulbs used for micropropagation in October and those stored two months at 30 °C prior to in vitro culture. However, no direct correlation between the absolute amounts of ABA and percentage of regeneration was found.