Nucleic Acid and protein changes in relation to cold acclimation and freezing injury of korean boxwood leaves

Quantitative and qualitative differences in nucleic acids of Korean boxwood (Buxus microphylla var. Koreana) leaves were determined by methylated albumin kieselguhr chromatography at different levels of cold hardiness. During cold acclimation there was an increase in RNA, mainly ribosomal RNA, with little or no change in DNA. The increase in ribosomal RNA was closely paralleled by an increase in water soluble and membrane bound proteins. As cold hardiness increased, ribonuclease activity declined.     

Leaf vascular systems in C(3) and C(4) grasses: a two-dimensional analysis

BACKGROUND AND AIMS: It is well documented that C(4) grasses have a shorter distance between longitudinal veins in the leaves than C(3) grasses. In grass leaves, however, veins with different structures and functions are differentiated: large longitudinal veins, small longitudinal veins and transverse veins. Thus, the densities of the three types of vein in leaves of C(3) and C(4) grasses were investigated from a two-dimensional perspective. METHODS: Vein densities in cleared leaves of 15 C(3) and 26 C(4) grasses representing different taxonomic groups and photosynthetic subtypes were analysed. KEY RESULTS: The C(4) grasses had denser transverse veins and denser small longitudinal veins than the C(3) grasses (1.9 and 2.1 times in interveinal distance), but there was no significant difference in large longitudinal veins. The total length of the three vein types per unit area in the C(4) grasses was 2.1 times that in the C(3) grasses. The ratio of transverse vein length to total vein length was 14.3 % in C(3) grasses and 9.9 % in C(4) grasses. The C(3) grasses generally had greater species variation in the vascular distances than the C(4) grasses. The bambusoid and panicoid C(3) grasses tended to have a denser vascular system than the festucoid C(3) grasses. There were no significant differences in the interveinal distances of the three vein types between C(4) subtypes, although the NADP-malic enzyme grasses tended to have a shorter distance between small longitudinal veins than the NAD-malic enzyme and phosphoenolpyruvate carboxykinase grasses. CONCLUSIONS: It seems that C(4) grasses have structurally a superior photosynthate translocation and water distribution system by developing denser networks of small longitudinal and transverse veins, while keeping a constant density of large longitudinal veins. The bambusoid and panicoid C(3) grasses have a vascular system that is more similar to that in C(4) grasses than to that in the festucoid C(3) grasses.     

Insect herbivory on C3 and C4 grasses

Summary This study tested the hypothesis that grasses with the C₄ photosynthetic pathway are avoided as a food source by insect herbivores in natural communities. Insects were sampled from ten pairs of C₃–C₄ grasses and their distributions analyzed by paired comparisons tests. Results showed no statistically significant differences in herbivore utilization of C₃–C₄ species. However, there was a trend towards heavier utilization of C₃ species when means for both plant groups were compared. In particular, Homoptera and Diptera showed heavier usage of C₃ plants. Significant correlations between insect abundances and plant protein levels suggest that herbivores respond to the higher protein content of C₃ grasses. ¹³C values for six of the most common grasshopper species in the study area indicated that three species fed on C₃ plants, two species fed on C₄ plants, and one species consumed a mixture of C₃ and C₄ tissue.Welder Wildlife Refuge Contribution Number 213     

The role of ABA in freezing tolerance and cold acclimation in barley

The role of ABA in freezing resistance in nonacclimated and cold‐acclimated barley ( Hordeum vulgare L.) was studied. Eleven nonacclimated cultivars differed in their LT₅₀, ranging from −10.8 to −4.8°C. Sugars, free proline, soluble proteins and ABA were analyzed in nonacclimated cultivars and during cold acclimation of one cultivar. There was an inverse correlation between LT₅₀ and both ABA and sucrose contents. Exogenous ABA caused a decrease in the freezing point of leaf tissue in the cultivar with the lowest level of endogenous ABA, but not in the cultivar with the highest level, suggesting that ABA in the latter may be near the optimum endogenous level to induce freezing tolerance. Plants of cv. Aramir treated with ABA or allowed to acclimate to cold temperature increased their soluble sugar content to a similar level. The LT₅₀ of leaves of cold‐acclimated cv. Aramir decreased from −5.8 to −11.4°C, with biphasic kinetics, accumulating proline and soluble sugars with similar kinetics. The biphasic profile observed during cold acclimation could be a direct consequence of cryoprotectant accumulation kinetics. ABA and soluble protein accumulation showed a single step profile, associated mainly with the second phase of the LT₅₀ decrease. Thus, a significant increase in endogenous ABA is part of the response of barley to low temperature and may be required as a signal for the second phase of cold acclimation. Endogenous ABA contents in the nonacclimated state may determine constitutive freezing tolerance.     

Effect of cold acclimation on the incidence of two forms of freezing injury in protoplasts isolated from rye leaves

The freezing tolerance and incidence of two forms of freezing injury (expansion-induced lysis and loss of osmotic responsiveness) were determined for protoplasts isolated from rye leaves (Secale cereale L. cv Puma) at various times during cold acclimation. During the first 4 weeks of the cold acclimation period, the LT₅₀ (i.e. the minimum temperature at which 50% of the protoplasts survived) decreased from −5°C to −25°C. In protoplasts isolated from nonacclimated leaves (NA protoplasts), expansion-induced lysis (EIL) was the predominant form of injury at the LT₅₀. However, after only 1 week of cold acclimation, the incidence of EIL was reduced to less than 10% at any subzero temperature; and loss of osmotic responsiveness was the predominant form of injury, regardless of the freezing temperature. Fusion of either NA protoplasts or protoplasts isolated from leaves of seedlings cold acclimated for 1 week (1-week ACC protoplasts) with liposomes of dilinoleoylphosphatidylcholine also decreased the incidence of EIL to less than 10%. Fusion of protoplasts with dilinoleoylphosphatidylcholine diminished the incidence of loss of osmotic responsiveness, but only in NA protoplasts or 1-week ACC protoplasts that were frozen to temperatures over the range of -5 to -10°C. These results suggest that the cold acclimation process, which results in a quantitative increase in freezing resistance, involves several different qualitative changes in the cryobehavior of the plasma membrane.     

植物抗寒及其基因表达研究进展

植物经过逐渐降低的温度从而提高抗寒能力 ,这个过程被人们称为低温驯化。植物低温驯化过程是一个复杂的生理、生化和能量代谢变化过程 ,这些变化主要包括膜系统的稳定性、可溶性蛋白的积累和小分子渗透物质 ,比如脯氨酸、糖等 ,这些变化中的一些是植物抗寒必需的 ,而另外一些变化不是必需的。主要对冷害和低温生理生化变化、低温诱导表达基因的功能和作用、低温驯化的调节机制及其信号转导方面进行了综述。通过差别筛选 c DNA文库的方法已经鉴定了许多低温诱导表达、进而提高植物抗寒能力的基因 ,其中有脱水素、COR基因和 CBF1转录因子等。低温信号的感受、转导和调节表达是低温驯化的关键环节 ,低温信号的转导过程与干旱胁迫之间具有一定的交叉 ,这为利用 ABA等来提高植物抗寒能力成为可能 ,相信不久的将来人们可以通过提高植物抗寒能力从而增加经济产量成为现实。     

The ecological distribution of C4 and C3 grasses in the Hawaiian Islands

Summary Isoenzyme analyses were carried out on the hypotetraploid Stellaria media and the diploid Stellaria pallida occurring on abandoned sandy fields in the coastal dunes of the Netherlands. Amylase isoenzymes in germinating seeds of Stellaria media showed a striking intrapopulational variability, which is in accordance with the germination behaviour of the different subpopulations. In contrast to these findings no intrapopulational variation nor developmental change in isoenzyme pattern could be detected on basis of 14 other analysed leaf enzymes. For five enzymes there was a difference in isoenzyme pattern between the species Stellaria media and Stellaria pallida. Of these five enzymes two showed an interpopulational variation in Stellaria media. The fact that there was almost no variability of the isoenzyme pattern in Stellaria pallida could be explained by the permanent autogamous (cleistogamous) state of the species. Effects of polyploidy on the isoenzyme pattern and its activity could not be demonstrated. Consequences of the extreme difference between the monomorphic pattern of the leaf enzymes as opposed to the polymorphism of the amylase are discussed in view of the difference in germination ecology and lifecycle strategy of Stellaria media subpopulations.     

Cryoprotective and osmotic responses to cold acclimation and freezing in freeze-tolerant and freeze-intolerant earthworms

In this paper we present the results of physiological responses to winter acclimation and tissue freezing in a freeze-tolerant Siberian earthworm, Eisenia nordenskioeldi, and two freeze-intolerant, temperate earthworm species, Lumbricus rubellus and Aporrectodea caliginosa. By analysing the physiological responses to freezing of both types we sought to identify some key factors promoting freeze tolerance in earthworms. Winter acclimation was followed by a significant increase in osmolality of body fluids in E. nordenskioeldi, from 197 mosmol kg⁻¹ in 10 °C-acclimated animals to 365 mosmol kg⁻¹ in animals acclimated to 0 °C. Cold acclimation did not cause any change in body fluid osmolality in the two freeze-intolerant species. As a response to ice formation in the body, the freeze-intolerant species produced copious amounts of slime and expulsion of coelomic fluids, and thereby lost 10–30% of their total water content. Contrary to this, the freeze-tolerant species did not lose water upon freezing. At temperatures down to −6.5 °C, the ice content in the freeze-tolerant E. nordenskioeldi was significantly lower than in L. rubellus. At lower temperatures there were no differences in ice content between the two species. Cold acclimated, but unfrozen, specimens of all three species had low levels of ammonia, urea, lactate, glycerol and glucose. As a response to ice formation, glucose levels significantly increased within the first 24 h of freezing. This was most pronounced in E. nordenskioeldi where a 153-fold increase of glucose was seen (94 mmol · l⁻¹). In L. rubellus and A. caliginosa a 19-fold and 17-fold increase in glucose was seen. This is the first study on physiological mechanisms promoting freeze tolerance in E. nordenskioeldi, or any other oligochaete. Our results suggest that the cryoprotective system of this species more closely resembles that of freeze-tolerant anurans, which synthesize cryoprotectants only after tissues begin to freeze, than that of cold-hardy invertebrates which exhibit a preparatory accumulation of cryoprotectants during seasonal exposure to low temperature. Accepted: 10 February 1999     

Implications of quantum yield differences on the distributions of C3 and C4 grasses

Summary The implications of a reduced quantum yield (initial slope of the photosynthetic light response curve) in C₄ plants and temperature dependence of quantum yield in C₃ plants on total canopy primary production were investigated using computer simulations. Since reduced quantum yield represents the only known disadvantage of the C₄ photosynthetic pathway, simulations were conducted with grass canopies (high LAI and hence photosynthesis in most leaves will be light-limited) to see if quantum yield is a significant factor in limiting the primary production and thus distributions of C₄ grasses. Simulations were performed for three biogeographical or environmental conditions: the Great Plains region of North America, the Sonoran Desert of North America, and shade habitats. For all three cases, the simulations predicted either spatial or temporal gradients in the abundances of C₄ grasses identical to the abundance patterns of C₄ grasses observed in the field. It is thus concluded that while the C₄ photosynthetic mechanism may be highly advantageous in specific environments, it may be disadvantageous in others.C.I.W.-D.P.B. Publication No. 598     

The quantum yield for CO2 uptake in C3 and C4 grasses

The quantum yield for CO₂ uptake was measured in C₃ and C₄ monocot species from several different grassland habitats. When the quantum yield was measured in the presence of 21% O₂ and 340 cm³ m^-3 CO₂, values were very similar in C₃ monocots, C₃ dicots, and C₄ monocots (0.045–0.056 mole CO₂ · mole^-1 quanta absorbed). In the presence of 2% O₂ and 800 cm³ m^-3 CO₂, enhancements of the quantum yield values occurred for the C₃ plants (both monocots and dicots), but not for C₄ monocots. A dependence of the quantum yield on leaf temperature was observed in the C₃ grass, Agropyron smithii, but not in the C₄ grass, Bouteloua gracilis, in 21% O₂ and 340 cm³ m^-3 CO₂. At leaf temperatures between 22–25°C the quantum yield values were approximately equal in the two species.     

Biochemical and physiological mechanisms related to cold acclimation and enhanced freezing tolerance in poplar plantlets

Temperature is one of the abiotic factors limiting growth and productivity of plants. In the present work, the effect of low non-freezing temperature, as inducer of 'cold acclimation', was studied in poplar. Actively growing plantlets of Populus tremula  ×  Populus tremuloides cv. Muhs 1 were used, and cold treatment consisted in whole plants exposure to 4°C in controlled conditions. Leaves of cold-treated poplars were shown to be acclimated, as an increase of their freezing tolerance was measured using electrolyte leakage. Chlorophyll fluorescence measurements revealed a decrease in photosystem II efficiency while the pigment contents of leaves did not vary. In contrast, after 1 week of cold exposure, an accumulation of pigments was noted in the stems near the apex of the stressed plants as confirmed by chromatographic analyses. Simultaneously, a rapid accumulation of osmoprotectants, i.e. carbohydrates (measured by spectrometry), and of stress indicators (e.g. putrescine) occurred; changes in protein patterns also arose. Indeed, Western blot studies revealed that the expression of three families of stress-related proteins, i.e. dehydrins, stress protein 1 and heat-shock protein 70, was activated or induced by low temperatures. This study complements a previous work on proteomic and individual carbohydrates and provides insight in the ability of poplar plantlets to cold acclimate and to cope with low temperatures by diverse mechanisms (growth cessation, carbohydrate, pigment, polyamine and protein accumulations) related to stress response or involved in acclimation process.     

Improving our understanding of environmental controls on the distribution of C3 and C4 grasses

A number of studies have demonstrated the ecological sorting of C₃ and C₄ grasses along temperature and moisture gradients. However, previous studies of C₃ and C₄ grass biogeography have often inadvertently compared species in different and relatively unrelated lineages, which are associated with different environmental settings and distinct adaptive traits. Such confounded comparisons of C₃ and C₄ grasses may bias our understanding of ecological sorting imposed strictly by photosynthetic pathway. Here, we used MaxEnt species distribution modeling in combination with satellite data to understand the functional diversity of C₃ and C₄ grasses by comparing both large clades and closely related sister taxa. Similar to previous work, we found that C₄ grasses showed a preference for regions with higher temperatures and lower precipitation compared with grasses using the C₃ pathway. However, air temperature differences were smaller (2 °C vs. 4 °C) and precipitation and % tree cover differences were larger (1783 mm vs. 755 mm, 21.3% vs. 7.7%, respectively) when comparing C₃ and C₄ grasses within the same clade vs. comparing all C₄ and all C₃ grasses (i.e., ignoring phylogenetic structure). These results were due to important differences in the environmental preferences of C₃ BEP and PACMAD clades (the two main grass clades). Winter precipitation was found to be more important for understanding the distribution and environmental niche of C₃ PACMADs in comparison with both C₃ BEPs and C₄ taxa, for which temperature was much more important. Results comparing closely related C₃–C₄ sister taxa supported the patterns derived from our modeling of the larger clade groupings. Our findings, which are novel in comparing the distribution and niches of clades, demonstrate that the evolutionary history of taxa is important for understanding the functional diversity of C₃ and C₄ grasses, and should have implications for how grasslands will respond to global change.     

Induction of freezing tolerance in spinach is associated with the synthesis of cold acclimation induced proteins

Spinach (Spinacia oleracea L. cv Bloomsdale) seedlings cultured in vitro were used to study changes in protein synthesis during cold acclimation. Seedlings grown for 3 weeks postsowing on an inorganic-nutrient-agar medium were able to increase their freezing tolerance when grown at 5°C. During cold acclimation at 5°C and deacclimation at 25°C, the kinetics of freezing tolerance induction and loss were similar to that of soil-grown plants. Freezing tolerance increased after 1 day of cold acclimation and reached a maximum within 7 days. Upon deacclimation at 25°C, freezing tolerance declined within 1 day and was largely lost by the 7th day. Leaf proteins of intact plants grown at 5 and 25°C were in vivo radiolabeled, without wounding or injury, to high specific activities with [³⁵S]methionine. Leaf proteins were radiolabeled at 0, 1, 2, 3, 4, 7, and 14 days of cold acclimation and at 1, 3, and 7 days of deacclimation. Up to 500 labeled proteins were separated by two-dimensional gel electrophoresis and visualized by fluorography. A rapid and stable change in the protein synthesis pattern was observed when seedlings were transferred to the low temperature environment. Cold-acclimated leaves contained 22 polypeptides not found in nonacclimated leaves. Exposure to 5°C induced the synthesis of three high molecular weight cold acclimation proteins (CAPs) (M_r of about 160,000, 117,000, and 85,000) and greatly increased the synthesis of a fourth high molecular weight protein (M_r 79,000). These proteins were synthesized during day 1 and throughout the 14 day exposure to 5°C. During deacclimation, the synthesis of CAPs 160, 117, and 85 was greatly reduced by the first day of exposure to 25°C. However, CAP 79 was synthesized throughout the 7 day deacclimation treatment. Thus, the induction at low temperature and termination at warm temperature of the synthesis of CAPs 160, 117, and 85 was highly correlated with the induction and loss of freezing tolerance. Cold acclimation did not result in a general posttranslational modification of leaf proteins. Most of the observed changes in the two-dimensional gel patterns could be attributed to the de novo synthesis of proteins induced by low temperature. In spinach leaf tissue, heat shock altered the pattern of protein synthesis and induced the synthesis of several heat shock proteins (HSPs). One polypeptide synthesized in cold-acclimated leaves had a molecular weight and net charge (M_r 79,000, pI 4.8) similar to that of a HSP (M_r 83,000, pI 4.8). However, heat shock did not increase the freezing tolerance, and cold acclimation did not increase heat tolerance over that of nonacclimated plants, but heat-shocked leaf tissue was more tolerant to high temperatures than nonacclimated or cold-acclimated leaf tissue. When protein extracts from heat-shocked and cold-acclimated leaves were mixed and separated in the same two-dimensional gel, the CAP and HSP were shown to be two separate polypeptides with slightly different isoelectric points and molecular weights.     

Olive (Olea europaea L.) freezing tolerance related to antioxidant enzymes activity during cold acclimation and non acclimation

The changes in the antioxidant enzymes activity, total protein and proline content and their correlations with freezing tolerance (FT) (expressed as LT₅₀) were investigated at 11 different olive cultivars at cold-acclimation (CA, in February) and non-acclimation (NA, in August) stages. Leaf samples were collected from each cultivar and were divided into two groups. The first group was immediately frozen in liquid nitrogen for further biochemical analysis. The second ones was subjected to different freezing temperatures (?5, ?10, ?15 and ?20 °C) for 10 h, in order to determine their FT. The unfrozen control samples were kept at 4 °C. The results showed that Fishomi, Mission and Shengeh were the most freezing tolerant among other cultivars. In contrast, Zard, Manzanilla and Amigdalolia were the most sensitive ones. The cold acclimation enhanced the activities of superoxide dismutase (SOD), peroxidase (POD), ascorbate peroxidase (APX), catalase (CAT), polyphenol oxidase (PPO) and total protein content. However, proline content and phenylalanine ammonia-lyase (PAL) activity did not change or even decreased slightly at CA stage, compare to those samples at NA stage. It was found that LT₅₀ to be closely correlated to POD, CAT, and PPO activity at CA and NA stages. Overall, higher leaf POD, CAT, and PPO activity could be used as important selection criteria in screening tolerant olive cultivars for cold zone climatic.     

Mycorrhizal symbiosis induces plant carbon reallocation differently in C3 and C4 Panicum grasses

Plant and Soil - Although arbuscular mycorrhizal symbiosis is common in many plants with either C3 or C4 photosynthesis, it remains poorly understood whether photosynthesis type has any significant...