Growth-active gibberellins overcome the very slow shoot growth of Hancornia speciosa, an important fruit tree from the Brazilian “Cerrado”

Shoot elongation of Hancornia speciosa, an endangered tree from the Brazilian savannah “Cerrado”, is very slow, thus limiting nursery production of plants. Gibberellins (GAs) A₁, A₃, and A₅, and two inhibitors of GA biosynthesis, trinexapac-ethyl and ancymidol were applied to shoots of Hancornia seedlings. GA₁ and GA₃ significantly stimulated shoot elongation, while GA₅ had no significant effect. Trinexapac-ethyl and ancymidol, both at 100 μg per seedling, inhibited shoot elongation up to 45 days after treatment, though the effect was statistically significant only for ancymidol. Somewhat surprisingly, exogenous GA₃ more effectively stimulated shoot elongation in SD-grown plants, than in LD-grown plants. The results from exogenous application of GAs and inhibitors of GA biosynthesis imply that Hancornia shoot growth is controlled by GAs, and that level of endogenous growth-active GAs is likely to be the limiting factor for shoot elongation in Hancornia. Application of GAs thus offer a practical method for nursery production of Hancornia seedlings for outplanting into the field.     

Effects of Yariv dyes, arabinogalactan-protein binding reagents, on the growth and viability of Brazilian pine suspension culture cells

Arabinogalactan-proteins (AGPs) are a family of highly glycosylated hydroxyproline-rich glycoproteins implicated in several aspects of plant growth and development. (β-d-glucosyl)₃ Yariv phenylglycoside (β-GlcY), commonly known as Yariv reagent, selectively binds AGPs. We treated cell suspension cultures of Araucaria angustifolia, the Brazilian pine, with β-GlcY and observed inhibition of biomass increase in a culture medium with 50 μM β-GlcY. However, the growth was not inhibited by (α-d-galactosyl)₃ Yariv phenylglycoside (α-GalY) which does not bind AGPs. Fluorescein diacetate staining of cells indicated that β-GlcY severely affected cell viability. However, cell swelling, bursting and release of cellular contents, all characteristics of necrotic cell death, were not observed in β-GlcY-treated cells. Instead, programmed cell death (PCD) structural changes such as cytoplasmic shrinkage and condensation were observed in β-GlcY-treated cells. In addition, callose accumulation, which is another marker of PCD, was also observed in β-GlcY-treated cells. The use of both, Ac-VEID-CHO, an inhibitor of caspase-like proteolytic activity related to PCD, and phenyl methyl sulphonyl fluoride (PMSF), a protease inhibitor known to suppress PCD, in the culture medium did not reverse the growth inhibition caused by β-GlcY. These data indicate that the β-GlcY-induced inhibition of Araucaria cell’s growth is related to AGP perturbation, and also that this growth inhibition is due to increased cell death not driven by necrosis.     

Characterization of cold-induced changes in the fatty acids profile of rice seedlings

Rice, a staple food for more than one half of the world’s population, is one of the most cold-sensitive cereals. Breeding programs aimed at increasing rice production are expected to reduce cold-imposed grain losses. Several reports have demonstrated that cold induce differential effects on the fatty acids profile of membranes in chilling-sensitive and chilling-tolerant plants. In this work, we evaluated changes in fatty acid (FA) composition as a potential screening tool to evaluate chilling sensitivity of rice accessions. Cold exposure led to the preferential accumulation of the polyunsaturated linolenic and linoleic FAs and reduction of palmitic and stearic FAs, besides showing increased lignoceric acid content in roots of the variety. Similarly, roots of cold-exposed line Quila 66304 also presented preferential accumulation of linolenic and linoleic FAs and reduction of palmitic and stearic FAs. Cold exposure also led to enhanced levels of palmitic acid in shoots of Amaroo and, in a smaller extent, in shoots of Quila 66304. Linolenic acid was reduced in the shoots of both Amaroo and Quila 66304, while oleic acid content was reduced in shoots of Amaroo and slightly increased in shoots of Quila 66304. Double-bond index analysis indicated that 18 carbons FAs DBI for roots might be a good screening tool for cold response in rice. Results in this report demonstrate that cold-induced changes in FA profile represent a useful screening tool for early identification of differences in cold acclimation potential among rice accessions.     

Brassinosteroid-stimulated branch elongation in the marubakaido apple rootstock

Brassinosteroids (BRs) comprise a specific class of low-abundance plant steroids now recognized as a new class of phytohormones. In this paper, we demonstrate that a fluoro derivative of 28-homocastasterone (5F-HCTS) stimulates branch elongation in in vitro-grown shoots of Malus prunifolia, the marubakaido apple rootstock. In addition to that, we show that this BR-stimulated branch elongation is paralleled by an increase in ethylene release. However, either the presence of 1-amino-cyclopropane-1-carboxylic acid (ACC), the immediate precursor of ethylene in higher plants, in the culture medium or an ethylene-enriched atmosphere resulted in inhibition of branch elongation, indicating that the stimulation of branch elongation observed for 5F-HCTS-treated shoots in this study was not, at least directly, related to the BR-induced enhancement in ethylene release rate. Besides its positive effect on the marubakaido shoot growth, i.e. branch elongation, the 5F-HCTS-driven enhancement of branch elongation found in this study is potentially useful to improve micropropagation techniques for other plant species as well, especially woody species, in which branch elongation is typically a constraint for efficient micropropagation.     

In vitro propagation ofHancornia speciosa,a tropical fruit tree

Summary Hancornia speciosa fruit is highly desired for the juice and ice cream industry in tropical regions. A rapid reduction in germination ability ofH. speciosa seeds has been a problem for its large-scale cultivation. This paper describes anin vitro technique that may lead to an alternative propagation method forH. speciosa. Shoot apices and nodal segments from aseptically germinated young embryos were cultivatedin vitro on. Murashige and Skoog (1962) medium supplemented with growth regulators. Shoot multiplication was maintained by sequential subculture of shoot tips and nodal segments. N⁶-benzyladenine was the most effective cytokinin for the induction of shoot growth. N⁶-furfurylamino-purine, at various concentrations, yielded multiplication rates sevenfold lower than the highest multiplication rate found with N⁶-benzyladenine. Increased root initiation rate and root elongation was observed with the presence of γ-(indole-3) butyric acid in the half-strength Murashige and Skoog culture medium, especially at 10μM. N⁶-benzyladenine strongly inhibited rooting, even in the presence of γ-(indole-3) butyric acid. Thein-vitro-raised rooted plantlets were acclimatized to greenhouse environment through progressive reduction in relative humidity and later transplanted to the field.     

Arabinogalactan-proteins from cell suspension cultures of Araucaria angustifolia

Arabinogalactan-proteins (AGPs), found in the culture medium of suspension cells of Araucaria angustifolia grown in plant growth regulator-free and plant growth regulator-containing BM media, BM0 and BM2, respectively, were evaluated quantitatively and qualitatively. The concentrated extracellular fractions (CEFs), obtained from suspension cell cultures grown for 20 days in BM0 and BM2 media yielded two fractions, CEF-0 and CEF-2, respectively. CEF-0 and CEF-2 was submitted to selective precipitation using the β-glucosyl Yariv reagent (β-GlcY) to isolate AGPs for structural characterization; this yielded fractions designated CEF-0YPF and CEF-2YPF, respectively. The monosaccharide composition analysis established that samples were composed of Rha, Ara, Gal and uronic acid in a molar ratio 3:37:55:5 (CEF-0YPF) and 1:37:58:4 (CEF-2YPF), although trace amounts (<0.5 mol%) of Xyl were also found. Methylation analysis of CEF-YPF fractions showed similar results for both CEF-0YPF and CEF-2YPF, with non-reducing terminal units of Araf, Arap, Galp, Rhap and Xylp, as well as 3-O-substituted and 5-O-substituted Araf units and 3-O-substituted, 6-O-substituted and 3,6-di-O-susbtituted Galp units. The amino acid composition analysis established Ser, Ala, and Hyp as major amino acids in both samples. In conclusion, this investigation has shown that CEF-0YPF and CEF-2YPF contain macromolecules having typical AGP characteristics, including a Hyp/Ala/Ser-rich protein moiety, a (1 → 3) and/or (1 → 6) linked β-d-galactopyranosyl main chain substituted by Gal, Ara, Rha and Xyl residues, and binding affinity for β-GlcY and monoclonal anti-AGP antibodies.     

Shooting Control in Eucalyptus Grandis × E. urophylla Hybrid: Comparative Effects of 28-Homocastasterone and a 5α-Monofluoro Derivative

28-Homocastasterone (28-HCTS), a brassinosteroid, was used to treat in vitro-grown shoots of a hybrid between Eucalyptus grandis and E. urophylla. Treated shoots showed enhanced elongation and formation of new main shoots (the shoots originating directly from the initial explant) at low doses. Coincidently, there was reduced elongation and formation of primary lateral shoots (shoots originating from the main shoot). However, a 5α-monofluoro derivative of 28-HCTS (5F-HCTS) was unable to either stimulate elongation or formation of new main shoots, although it did stimulate elongation of primary lateral shoots. In conclusion, it is quite apparent that exogenously supplied brassinosteroids are able to change shooting patterns in Eucalyptus. These findings have practical biotechnological applications, for example on the improvement of micropropagation techniques for clonal propagation of woody angiosperms.     

Differential performance of marubakaido apple rootstock shoots grown in culture media containing different agar brands: dynamic rheological analysis

Shoots of the marubakaido apple rootstock grown in culture medium containing BBL agar presented significantly lower multiplication rate (MR) compared to MRs found for shoots grown in medium containing A-7002, A-7921, Select, and Phytagar as gelling agents. In addition, significant hyperhydricity was found for shoots grown in Phytagar and A-7921 agar-containing media. Analysis of elastic (G′) and viscous (G″) modulus showed that for all of the five agar brands used in this study, G′ was always much higher, i.e., typically one order of magnitude higher than G″, which characterizes a strong gel. G′ changed randomly with time for all of the agar brands studied, except for BBL, which presented progressive decline in G′ throughout the culture cycle. Examination of G′, within the same week, showed that Select agar always had the smallest G′, while Phytagar always had the highest G′. Analysis of the loss tangent (tan δ = G″/G′), a better indicator for gel behavior compared to G′ isolated, showed that tan δ for Select and Phytagar were always between tan δ values found for A-7002 and BBL. In addition, analysis of tan δ also indicated that BBL and Select agars showed a significantly weaker gel network, compared to Phytagar, A-7002 and A-7921 agars after the third week of culture. When seen together, these results indicate that shoot performance for the marubakaido rootstock is not related to agar gel strength. In addition, the high hyperhydricity rate found for shoots grown on agars A-7921 and Phytagar could not be related to agar gel strength, as well. Analysis of HPSEC profiles indicated that the best performance, i.e., multiplication rate, of marubakaido shoots in agars A-7002 and A-7921 is likely to be related to their lower polydispersity and/or smaller amount of high molecular weight fractions, compared to BBL, Phytagar, and Select agars.     

Effect of inhibitors of ethylene biosynthesis and signal transduction pathway on the multiplication of in vitro-grown Hancornia speciosa

Initiation and elongation of lateral buds is stimulated in in vitro-grown shoots of Hancornia speciosa(a tropical fruit tree) by high temperature (35 °C), which is associated with the plant's reduced ability to release ethylene. However, no increase in either lateral shoot elongation or multiplication rate of H. speciosa shoots growing in vitro at non-shoot inducing temperature (31 °C) was associated with the presence of inhibitors of ethylene biosynthesis, such as α-amino isobutyric acid and cobalt ions or (aminooxy) acetic acid. Likewise, no increase in the multiplication rate was associated with aminoethoxyvinylglycine (AVG), another inhibitor of ethylene biosynthesis. In addition, stimulation of lateral shoot elongation in H. speciosa shoots grown at non shoot-inducing temperature was achieved when two inhibitors of the ethylene signal transduction pathway, silver thiosulphate (5–10 μM Ag⁺⁺) and 1-methylcyclopropene (90 nl l⁻¹) were present in the culture medium or in the culture vessel-headspace, respectively. However, increased multiplication only occurred with the 1-methylcyclopropene-treated shoots. Thus, lateral bud development with 1-methylcyclopropene could be used to enhance H. speciosa multiplication in vitro. This revised version was published online in June 2006 with corrections to the Cover Date.