Abstract. The effects of growing seedlings of red oak (Quercus rubra) and red ash (Fraxinus pennsylvanica) with Hoagland solutions containing five N-regimes, differing in the N-forms (NH4, NO3) and concentrations (High and Low), in relation to light intensity were investigated by the utilization of enzymatic markers of the N assimilation pathway, nitrate reductase (NR) and glutamine synthetase (GS). Red oak and red ash showed different patterns of N-assimilation. Red oak seedlings assimilated NO3 in low amounts in their roots and leaves, whereas red ash seedlings assimilated high amounts of NO3, mostly in the leaves. A significant amount of constitutive NR activity was found in red oak seedlings supplied with NH4 N-regime. This could be characteristic of a species adapted to soils that are poor in nitrogen. Root GS activity was lower in red oak seedlings than in red ash seedlings, indicating that the rate of NH4 assimilation differed in these two hardwood species. Low irradiance reduced growth of both hardwood species, but greatly affected the specific leaf area of red ash and reduced NO3 assimilation (when data are expressed per leaf area). Both species reacted similarly to N-regimes in terms of relative growth rate.

Chromatin DNA-dependent RNA polymerases and RNases activities were measured in winter and spring varieties to understand the overall regulation of RNA synthesis during cold acclimation. We found that total RNA polymerase activities were significantly higher in chromatin isolated from winter wheat compared to the spring wheat during the acclimation period. This increase was parallel to the increase in protein and RNA contents during hardening. The ratio of RNA polymerase I to RNA polymerase II activity was higher than 2 in winter wheat after 30 days of hardening compared, to a ratio of 0.90 under the nonhardening conditions. The increase in activity and the ratio of polymerase I to polymerase II was maintained after the separation of the enzymes from the template, suggesting that RNA synthesis is regulated in part at the enzyme level. On the other hand, the chromatin associated RNase activity decreased in both varieties during acclimation, indicating a nonspecific inhibition caused by low temperature rather than a selective genetic response associated with cold acclimation.The enhancement of RNA and protein synthesis is one of the most important physiological changes in the hardy plant cell during the development of cold tolerance. It has been suggested that increased protein synthesis may result in the synthesis of cryoprotective substance(s) which increase membrane resistance to freezing and induce old hardiness (3,4,7,10,13,14,16,27,29). The mechanism by which cold hardening conditions regulate protein and RNA synthesis in hardy varieties is not fully known, but there is general agreement that frost hardening is associated with de novo protein synthesis. To understand the nature of this metabolic change, we studied the regulation of protein synthesis at the transcriptional level as a first step to elucidating the overall mechanism of regulation of cold hardiness. For this purpose, the functional activity of the chromatin was determined by measuring the change in DNA-dependent RNA polymerases and in RNase activities during hardening of two varieties of wheat differing in their degree of cold resistance. [3H]thymidine (20 Ci/mol) from New England Nuclear were used to label the RNA and DNA, respectively. The seedling was carefully removed with the root system intact and washed thoroughly before incubation in the nutrient solution containing the radioactive uridine or thymidine at a concentration of I ,uCi/ml. Gramicidin D, 6 Ag/ml (Calbiochem) was added to prevent bacterial growth during the experiment. MATERIALS AND METHODSExtraction of nucleic acids was carried out as described above and aliquots of RNA and DNA fractions were counted in a Nuclear Chicago ISOCAP 300 liquid scintillation spectrometer to determine incorporation of the radioactive precursor.Preparation of Chromatin. The tissues were homogenized in a Waring Blendor in 3 volumes per unit weight of: 0.5 M sucrose, 50 mM Tris-HCl (pH 8.0), 10 mM MgCI2, 4 mM ,B-mercaptoethanol, and 6 ,ug/ml gramicidin D. The homogenate was filtered through eigh...

A cryoselection protocol has been developed that provides freezing-tolerant callus that, in turn, can regenerate plants with enhanced cold hardiness. Tolerant calli were selected from spring wheat (Triticum aestivum L.) callus by immersion in liquid nitrogen without addition of cryoprotectants. Less than 15% of the calli survived the initial challenge, whereas 30 to 40% of previously selected calli survived subsequent exposure. Seed progeny from five of 11 regenerant (R2) lines tested exhibited significantly enhanced tolerance to freezing at -120C. Thus, cryoselection appears to involve at least in part, selection for genetic rather than epigenetic variants. Analysis of one callus line indicated that cryoselection did not induce significant alterations in lipid composition, adenylate energy charge, or freezing point. An increase in the soluble sugar component was detected. Changes were also detected in the protein complement of microsomal membrane and soluble protein extracts of cryoselected callus. In all, seven unique proteins ranging from 79 to 149 kilodaltons were identified. The results demonstrate that freezing tolerant callus can be isolated from a heterogeneous population by cryoselection, and factors that contribute to hardiness at the callus level are biologically stable and can contribute to tolerance at the whole plant level.esses occur where extraorgan freezing and intracellular solute accumulation are important strategies for survival. Several reports indicate that, at least within a species, the relative water content of meristematic regions is an important factor in survival (6, 15). Both factors limit the availability of freezable water and therefore, reduce the likelihood of intracellular ice formation.It is clear that variability in response to freezing stress extends even to the level ofa single cell line (7). As an extreme example, in a previous study from this laboratory, Chen and associates (4, 5) reported that 7.2% of Catharanthus roseus and 2.5% of Triticum aestivum suspension cells survived immersion in liquid nitrogen without the addition of cryoprotectants. From this observation it was speculated that severe freezing stress might be used to isolate freezing-tolerant variants from a heterogeneous population of cells.The present investigation was undertaken to determine: the extent of survival of spring wheat callus following freezing in LN22 without the addition of cryoprotectants; the ability of such "cryoselected" callus to demonstrate enhanced LN2 freezing tolerance after a second challenge; the potential of the cryoselected callus to regenerate plants with enhanced (conventional) freezing tolerance; and the biochemical alterations, if any, that might correlate with survival.Plant species exhibit varying degrees oftolerance to freezing stress. For example, while in temperate grasses the minimum temperature for survival is in the -15°to -30°C range, some woody species, in the fully hardened state, can survive immersion in liquid nitrogen (12). This variability is also reflect...

The effects of high O3 (200 nl l-1 during the light period) and high CO2 (650 &mgr;l l-1 CO2, 24 h a day) alone and in combination were studied on 45-day-old sugar maple (Acer saccharum Marsh.) seedlings for 61 days in growth chambers. After 2 months of treatment under the environmental conditions of the experiment, sugar maple seedlings did not show a marked response to the elevated CO2 treatment: the effect of high CO2 on biomass was only detected in the leaves which developed during the treatment, and assimilation rate was not increased. Under high O3 at ambient CO2, assimilation rate at days 41 and 55 and Rubisco content at day 61 decreased in the first pair of leaves; total biomass was reduced by 43%. In these seedlings large increases (more than 2-fold) in glucose 6-phosphate dehydrogenase (G6PDH, EC 1.1.1.49) activity and in anaplerotic CO2 fixation by phosphoenolpyruvate carboxylase (PEPC, EC 4.1.1.31) were observed, suggesting that an enhanced reducing power and carbon skeleton production was needed for detoxification and repair of oxidative damage. Under high O3 at elevated CO2, a stimulation of net CO2 assimilation was observed after 41 days but was no longer observed at day 55. However, at day 61, the total biomass was only reduced by 21% and stimulation of G6PDH and PEPC was less pronounced than under high O3 at ambient CO2. This suggests that high CO2 concentration protects, to some extent, against O3 by providing additional carbon and energy through increased net assimilation.

Through a survey of the literature and an analysis of selected national and regional statistics, a review of the incidence of osteoporosis and its consequences was performed. Results illustrate that the consequences of osteoporosis are preventable and should sensitize clinicians to the importance of early detection and the identification of risk factors for disease prevention and to early treatment once disease has been established. A marked increase in the annual incidence of hip fractures in all Canadians is noted. Whereas the incidence was less than 20,000 in 1981, the incidence of hip fracture grew to 27,342 in 1995, with 73% occurring in women. It is estimated that in Canada at least one in four women older than 50 years will have one or more osteoporosis-related fractures in their lifetimes. The consequences of these fractures are considerable, both for patients and healthcare services. Only half of all victims regain total autonomy, and the total direct costs in Canada stemming from osteoporosis are estimated to be $1.3 billion per year.