Plasmodesmal-mediated cell-to-cell transport in wheat roots is modulated by anaerobic stress

Summary Cell-to-cell transport of small molecules and ions occurs in plants through plasmodesmata. Plant roots are frequently subjected to localized anaerobic stress, with a resultant decrease in ATP. In order to determine the effect of this stress on plasmodesmal transport, fluorescent dyes of increasing molecular weight (0.46 to 10 kDa) were injected into epidermal and cortical cells of 3-day-old wheat roots, and their movement into neighboring cells was determined by fluorescence microscopy. Anaerobiosis was generated by N₂ gas or simulated by the presence of sodium azide, both of which reduced the ATP levels in the tissue by over 80%. In the absence of such stress, the upper limit for movement, or size exclusion limit (SEL), of cortical plasmodesmata was <1 kDa. The ATP analogue TNP-ADP (mw 681) moved across the plasmodesmata of unstressed roots, indicating that plasmodesmata may be conduits for nucleotide (ATP and ADP) exchange between cells. Upon imposition of stress, the SEL rose to between 5 and 10 kDa. This response of plasmodesmata to a decrease in the level of ATP suggests that they are constricted by an ATP-dependent process so as to maintain a restricted SEL. When roots are subjected to anaerobic stress, an increase in SEL may permit enhanced delivery of sugars to the affected cells of the root where anaerobic respiration could regenerate the needed ATP.Abbreviations F-dextran fluorescein-coupled dextran - LYCH Lucifer Yellow CH - SEL size exclusion limit - TNP-ADP 2-O-(trinitrophenyl)adenosine-5-diphosphate     

The influence of elevated CO2 on non-structural carbohydrate distribution and fructan accumulation in wheat canopies

We grew 2.4 m² wheat canopies in a large growth chamber under high photosynthetic photon flux (1000 μmol m⁻² s⁻¹) and using two CO₂ concentrations, 360 and 1200 μmol mol⁻¹. Photosynthetically active radiation (400–700 nm) was attenuated slightly faster through canopies grown in 360μmol mol⁻¹ than through canopies grown in 1200μmol mol⁻¹, even though high-CO₂ canopies attained larger leaf area indices. Tissue fractions were sampled from each 5-cm layer of the canopies. Leaf tissue sampled from the tops of canopies grown in 1200μmol mol⁻¹ accumulated significantly more total non-structural carbohydrate, starch, fructan, sucrose, and glucose (p≤ 0.05) than for canopies grown in 360μmol mol⁻¹. Non-structural carbohydrate did not significantly increase in the lower canopy layers of the elevated CO₂ treatment. Elevated CO₂ induced fructan synthesis in all leaf tissue fractions, but fructan formation was greatest in the uppermost leaf area. A moderate temperature reduction of 10 °C over 5d increased starch, fructan and glucose levels in canopies grown in 1200μmol mol⁻¹, but concentrations of sucrose and fructose decreased slightly or remained unchanged. Those results may correspond with the use of fructosyl-residues and release of glucose when sucrose is consumed in fructan synthesis.     

Genotypic variation in the responsiveness to GA3 within tall (rht1) and semi-dwarf (Rht1) spring wheat

The effect of GA₃ on coleoptile-and first leaf elongation of tall (rht1) and semi-dwarf (Rht1) nearly-isogenic genotypes, within each of 25 random F₉ wheat families, was determined on seedlings grown in a growth room at 18 °C. Conspicuous and very significant inter-family variation in the response of the first leaf to GA₃ application was found in both the rht1 and Rht1 genotypes. The magnitudes of the response of the different families within genotypes to GA₃ were not related to the leaf length of their untreated seedlings. It is suggested that, under given environmental conditions, background genotypic effects, inducing inter-family variation in responsiveness to GA₃, regulate the elongation growth up to the limits set by the Rht alleles.     

Phytohormonal changes in intact shoots of wheat and oilseed rape treated with the acylcyclohexanedione growth retardant prohexadione calcium

The effects of the acylcyclohexanedione-type growth retardant prohexadione calcium on seedling growth and endogenous levels of immunoreactive phytohormone-like substances in shoots of wheat ( Triticum aestivum L. cv. Kanzler) and oilseed rape ( Brassica napus L. ssp. napus cv. Lirajet) were studied. After treatment of seedlings with increasing retardant concentrations in hydroponics, plant height and fresh weight of shoots were reduced by up to 40%. Concomitantly, the amount of immunoreactive gibberellins decreased, on a fresh weight basis, when compared with levels in the shoots of control plants. In contrast, the levels of abscisic acid and dihydrozeatin riboside and isopentenyladenosine-type cytokinins were considerably elevated by the growth retardant. The content of 3-indoleacetic acid decreased slightly. These results suggest that, in addition to its effect on gibberellin content, prohexadione calcium also influences the levels of endogenous abscisic acid and cytokinins.     

Gene transfer to inflorescence and flower meristems using ballistic micro-targeting

Direct gene transfer to floral meristems could contribute to cell-fate mapping, to the study of flower-specific genes and promoters, and to the production of transgenic gametes via the transformation of sporogenic tissues. Despite the wide potential of its applications, direct gene transfer to floral meristems has not been achieved so far because of the lack of suitable technology. We show in this paper that ballistic micro-targeting is the technique of choice for this purpose, and in this way, we were able to transfer genes efficiently into excised wheat immature spikes. Particle size was adjusted for optimal penetration into the L1 and L2 cell layers of the spikes with limited cell damage. Spikes at different developmental stages were shot either with a plasmid containing two genes involved in anthocyanin biosynthesis or with a plasmid bearing the uidA (-glucuronidase) gene. The transient expression of these marker genes was observed in the different developmental stages tested and in cells of both the L1 and the L2 layers. The transient expression of the uidA gene was significantly increased when the sucrose concentration in the culture medium was increased from 0.06 to 0.52 M. At the highest concentration, 100% of the targeted spikes expressed the uidA gene, with an average of 69 blue cells per spike. Twelve days after microtargeting, multicellular sectors showing transgene expression and containing up to 17 cells were found in 85% of the shot immature inflorescences. This indicated that targeted cells survived particle bombardment. Sectors were found in primordia of both vegetative and reproductive organs.     

A minichromosome-like structure in wheat embryos

A crude nuclear fraction of resting wheat embryos was used as the source of putative plant minichromosomes: unique DNA sequences the size of genes and flanked by telomere-type repeats. Preliminary separation of low-molecular-weight DNA species from chromosomal DNA (Hirt's method), velocity sedimentation, and isopycnic centrifugation were followed by PCR amplification of minichromosome-like sequences. The most abundant PCR product was cloned and sequenced. In addition to telomeric repeats (defined by a PCR primer), which were the expected sequences, the linear DNA molecule (637 pb) contained an ARS-like element, RAP1-binding site, and two relatively long ORFs. The whole sequence seems to represent a naturally occurring plant minichromosome.     

Sucrose and fructan metabolism in wheat roots at chilling temperatures

Sucrose and fructan metabolism were studied in wheat ( Triticuin aotiirum L. cv. Tribal 800) roots during a period at chilling temperature. Enzyme activities related to fructan and sucrose metabolism were measured. Sucrose-sucrose fructosyl transfer-ase (EC 2.4.1.99) activity increased more than 25-fold when plants were cooled to 4°C. Sucrose synthase (EC 2.4.1.13) and sucrose-phosphate synthase (EC 2.4.1.14) activities also increased, but low temperatures had no significant effect on invertaso (EC 3.2.1.26) or on fructan hydrolase (EC 3.2.1.26) activities. The accumulation pattern of fructan in roots was different to that in leaves. In roots chilling stimulated the synthesis of fructans of high degree of polymerization.     

Aluminum enhancement of plant growth in acid rooting media. A case of reciprocal alleviation of toxicity by two toxic cations

The generally rhizotoxic ion Al³⁺ often enhances root growth at low concentrations. The hypothesis that Al³⁺ enhances growth by relieving H⁺ toxicity was tested with wheat seedlings ( Triticum aestivum L.). Growth enhancement by Al³⁺ only occurred under acidic conditions that reduced root elongation. Al³⁺ increased cell membrane electrical polarity and stimulated H⁺ extrusion. Previous investigations have shown that Al³⁺ decreases solute leakage at low _pH and that the alleviation of H⁺ toxicity by cations appears to be a general phenomenon with effectiveness dependent upon charge (C³⁺>C²⁺>C^l+). Alleviation of one cation toxicity by another toxic cation appears to be reciprocal so that Al³⁺ toxicity is relieved by H⁺. It has been argued previously that this latter phenomenon accounts for the apparent toxicity of ALOH²⁺ and Al(OH)⁺₂. Reduction of cell-surface electrical potential by the ameliorative cation may reduce the cell-surface activity of the toxic cation.     

The control of boron accumulation by two genotypes of wheat

The mechanism of boron (B) uptake in wheat was studied using two genotypes with known differences in their ability to accumulate B. Influx and efflux of B was measured in the roots of intact 21 d old plants.Roots grown in 15 M B, when transferred to solutions containing 1mM B showed a rapid increase in B content for up to 60 min, after which no further increase was evident up to 4 h. No genotypic difference in B influx was apparent over these time periods. Roots grown in 1mM B for 7 d and then rinsed in B-free solutions quickly lost most of B that they contained within 1 hour; little further efflux was observed over the following three hours. As with the influx, no genotypic difference in B flux was evident.It is suggested that the lack of genotypic difference in the short-term B fluxes could be due to a masking effect of extracellular B bound in the cell walls of the roots.Department of Botany, University of Adelaide