The Influence of Season and Phenology on Freezing Tolerance in Silene acaulis L., a Subarctic and Arctic Cushion Plant of Circumpolar Distribution

Changes in the freezing tolerance for Silene acaulis L., a subarcticand arctic species of circumpolar distribution, were examinedto understand the extent of cold hardening and dehardening thatoccurs seasonally and with changes in plant phenology. Shootsof whole plants collected on a mountain ridge near Tromsø,Norway (69° N, 700 m above sea level) were frozen undercontrolled conditions at cooling rates of 3 to 4°C h^-1.The extent of freezing-induced injury was examined both by chlorophyllfluorescence and by visual inspection with a microscope. A freezingtolerance level of -30°C was observed in mid-winter, basedon a 50% lethal point for freezing injury. Loss of cold hardinesswas substantial in mid-summer, with freezing tolerance of -8·5to -9°C observed in mid-July. Plants still covered by snowin mid-July had a freezing tolerance of -12·5 to -13°C.The maintenance of a basic level of freezing tolerance throughoutthe summer may be adaptive in the northern latitude-regionsbecause of the occurrence of episodic frosts during the growingseason.Copyright 1993, 1999 Academic Press Silene acaulis L., Caryophyllaceae, freezing tolerance, chlorophyll fluorescence, cushion plant     

Freezing Avoidance Mechanisms by Supercooling in Some Rhododendron flower Buds with Reference to Water Relations

Excised florets of some hardy Rhododendron species did not toleratefreezing at –5°C when ice-inoculated due to intracellularfreezing. Florets in intact December buds, however, could besupercooled to about –30°C. When flower buds of R.japonicum were slowly cooled with daily decrements of 5°Cto temperatures ranging from 0 to –20°C, the exothermtemperatures of the florets drastically decreased. This wasaccompanied by a decrease in water content of florets and peduncleand an increase in that of scales. The water in florets andthe peduncle is thought to migrate to scales and other tissuesduring the early stages of freezing; the dehydrated floret hasa lower freezing point which enhances its supercooling abilityand the dehydrated peduncle helps to maintain the supercooledstate of the florets. This hypothesis would explain the dependenceon the cooling rate of supercooling in Rhododendron flower buds.Water migration within flower buds was observed in other hardyRhododendron species with some variation in ice formation siteand the quantity of migrated water. The exotherm temperatureof excised florets was inversely proportional to their watercontent. Dehydration of flower buds by wind at 0°C alsoenhanced their supercooling ability. Mechanisms of freezingavoidance by supercooling in Rhododendron flower buds and therelationship of supercooling to freezing tolerance are discussed. ¹ Contribution No. 2254 from the Institute of Low TemperatureScience ² This is a revised form of the master's thesis of the seniorauthor (M.I.) which is cited in the present and previous papers(Sakai 1979a, b, etc.). (Received August 11, 1980; Accepted June 1, 1981)     

Supercooling ability of Rhododendron flower buds in relation to cooling rate and cold hardiness

The freezing process and supercooling ability in flower budsof 11 native Rhododendron species were examined with referenceto the cooling rate and cold hardiness by differential thermalanalysis. The freezing patterns of the excised whole buds variedwith the season: in autumn, buds froze as whole units, whilein winter, freezing was initiated in the scales and propagatedto each floret. The supercooling ability of florets was enhancedduring winter. The freezing patterns in winter buds were stronglyinfluenced by the cooling rate (1 to 30°C/hr). Althoughthe first exotherm in scales occurred at –5 to –10°Gand was rate-independent, the occurrence of several floret exothermsshifted considerably to lower subzero temperatures at slowerrates. The most reliable cooling rate for testing maximum supercoolingability was l°C/hr. The exotherm in florets of hardier speciesoccurred at –20 to –25°C and at –7 to–20°C for less hardy ones, and were well correlatedwith their killing temperatures. Water relations within budtissues in response to freezing are briefly discussed. (Received June 26, 1980; )     

Supercooling ability of Rhododendron flower buds in relation to cooling rate and cold hardiness

The freezing process and supercooling ability in flower budsof 11 native Rhododendron species were examined with referenceto the cooling rate and cold hardiness by differential thermalanalysis. The freezing patterns of the excised whole buds variedwith the season: in autumn, buds froze as whole units, whilein winter, freezing was initiated in the scales and propagatedto each floret. The supercooling ability of florets was enhancedduring winter. The freezing patterns in winter buds were stronglyinfluenced by the cooling rate (1 to 30°C/hr). Althoughthe first exotherm in scales occurred at –5 to –10°Gand was rate-independent, the occurrence of several floret exothermsshifted considerably to lower subzero temperatures at slowerrates. The most reliable cooling rate for testing maximum supercoolingability was l°C/hr. The exotherm in florets of hardier speciesoccurred at –20 to –25°C and at –7 to–20°C for less hardy ones, and were well correlatedwith their killing temperatures. Water relations within budtissues in response to freezing are briefly discussed. (Received June 26, 1980; )     

ABA and Low Temperature Induce Freezing Tolerance via Distinct Regulatory Pathways in Wheat

The role of ABA in the induction of freezing tolerance was investigatedin two wheat (T. aestivum L.) cultivars, Glenlea (spring var)and Fredrick (winter var). Exogenous application of ABA (5x10^–5M for 5 days at 24°C) increased the freezing tolerance ofintact plants by only 3°C (LT₅₀) in both cultivars. Maximalfreezing tolerance (LT₅₀ of –9°C for Glenlea and –17°Cfor Fredrick) could only be obtained with a low temperaturetreatment (6/2°C; day/night) for 40 days. These resultsshow that exogenously applied ABA cannot substitute for lowtemperature requirementto induce freezing tolerance in intactwheat plants. Furthermore, there was no increase in the endogenousABA level of wheat plants during low temperature acclimation,suggesting the absence of an essential role for ABA in the developmentof freezing tolerance in intact plants. On the other hand, ABAapplication (5x10^–5 M for 5 days at 24°C) to embryogenicwheat calli resulted in an increase of freezing tolerance similarto that achieved by low temperature. However, as in intact plants,there was no increase in the endogenous ABA level during lowtemperature acclimation of calli. These results indicate thatthe induction of freezing tolerance by low temperature is notassociated with an increase in ABA content. Using an antibodyspecific to a protein family associated with the developmentof freezing tolerance, we demonstrated that the induction offreezing tolerance by ABA in embryogenic wheat calli was correlatedwith the accumulation of a new 32 kDa protein. This proteinis specifically induced by ABA but shares a common antigenicitywith those induced by low temperature. These results suggestthat ABA induces freezing tolerance in wheat calli via a regulatorymechanism different from that of low temperature. (Received June 15, 1993; Accepted September 16, 1993)     

Betaine Improves Freezing Tolerance in Wheat

The accumulation of the osmolyte betaine was found to be correlatedwith the development of freezing tolerance (FT) of two wheatcultivars where it increases by about three fold during thecold acclimation period. Exogenous betaine application resultedin a large increase in total osmolality mostly due to betaineaccumulation. Plants that accumulated betaine are more tolerantto freezing stress since a four day exposure to 250 mM betaineresulted in a LT₅₀ of –8°C (in spring wheat Glenlea)and –9°C (in winter wheat Fredrick) compared to –3°C(Glenlea) and –4°C (Fredrick) for control non-exposedplants. Betaine treatment (250 mM) during cold acclimation increasedFT in an additive manner since the LT₅₀ reached –14°C(Glenlea) and –22°C (Fredrick) compared to –8°C(Glenlea) and –16°C (Fredrick) for plants that arecold acclimated in the absence of betaine. These results showthat betaine treatment can improve FT by more than 5°C inboth non-acclimated and cold-acclimated plants. The betainetreatment resulted in the induction of a subset of low temperatureresponsive genes, such as the wcor410, and wcor413, that arealso induced by salinity or drought stresses. In addition tothese genetic responses, betaine treatment was also able toimprove the tolerance to photoin-hibition of PSII and the steady-stateyield of electron transport over PSII in a manner that mimickedcold-acclimated plants. These data also suggest that betaineimproves FT by eliciting some of the genetic and physiologicalresponses associated with cold acclimation. (Received April 23, 1998; Accepted September 4, 1998)     

Changes in Tissue Freezing in Senecio vulgaris infected by Rust (Puccinia lagenophorae)

Freezing of healthy and rust (Puccinia lagenophorae) infectedleaves of Senecio vulgaris was compared calorimetrically bythermal analysis. In fully expanded leaves the threshold freezingtemperature was in the range –6.8 to –8.4 °Cin controls but –3.0 to –5.1 °C in leaves withsporulating rust sori. Comparable values in expanding leaveswere –5.0 to –8.9 °C and –3.9 to –6.7°C for healthy and rusted tissues, respectively. The bulktissue freezing point was between –1.0 and –4.0°C in both fully expanded and expanding healthy leaves,and was increased by infection by between +0.2 and 2.5 °C.Whereas healthy leaves supercooled by 3.1–5.8 °C,rusted leaves supercooled by only 1.8–4.9 °C Supercoolingof control leaves was reduced by dusting with aeciospores, particularlywhen leaves were wounded to simulate the rupture of the surfacecaused by sporulation, but wounding alone had no significanteffect. Supercooling of distilled water was also significantlyreduced by aeciospores, suspended at a concentration of 10⁵spores ml^–1. It is concluded that rust-induced changes in leaf freezing inS. vulgaris grown in controlled environments were due to anincrease in the number of sites for ice nucleation, caused bythe presence of the aeciospores, and increased penetration ofice into internal tissues, resulting from damage to the cuticleand epidermis. Although data for frost resistance obtained inthe growth-room are similar to previous field observations,the role of the above mechanisms under field conditions remainsunproven. Senecio vulgaris (groundsel), Puccinia lagenophorae (rust), low temperature, freezing resistance     

Cold tolerance and dehydration in Enchytraeidae from Svalbard

When cooled in contact with moisture, eight species of arctic Enchytraeidae from Svalbard were killed by freezing within minutes or hours at −3 and −5 °C; an exception was Enchytraeus kincaidi which survived for up to 2 days. When the temperature approached 0 °C the enchytraeids apparently tried to escape from the moist soil. The supercooling capacity of the enchytraeids was relatively low, with mean supercooling points of −5 to −8 °C. In contrast, specimens of several species were extracted from soil cores that had been frozen in their intact state at −15 °C for up to 71 days. Compared to freezing in a moist environment, higher survival rates were obtained during cooling at freezing temperatures in dry soil. Survival was recorded in species kept at −3 °C for up to 35 days, and in some species kept at −6 °C for up to 17 days. Slow warming greatly increased survival rates at −6 °C . The results strongly suggest that arctic enchytraeids avoid freezing by dehydration at subzero temperatures. In agreement with this, weight losses of up to ca. 42% of fresh weight were recorded in Mesenchytraeus spp. and of up to 55% in Enchytraeus kincaidi at water vapour pressures above ice at −3 to −6 °C. All specimens survived dehydration under these conditions. Accepted: 12 December 1997     

Freezing tolerance, protein composition, and abscisic acid localization and content of pea epicotyl, shoot, and root tissue 3

The freezing tolerance of many plants, such as pea (Pisum sativum),is increased by exposure to low temperature or abscisic acidtreatment, although the physiological basis of this phenomenonis poorly understood. The freezing tolerance of pea shoot tips,root tips, and epicotyl tissue was tested after cold acclimationat 2C, dehydration/rehydration, applications of 10^–4M abscisic acid (ABA), and deacclimation at 25C. Tests wereconducted using the cultivar ‘Alaska’, an ABA-deficientmutant ‘wil’, and its ‘wildtype’. Freezinginjury was determined graphically as the temperature that caused50% injury (T₅₀) from electrical conductivity. Endogenous ABAwas measured using an indirect enzyme-linked immunosorbant assay,and novel proteins were detected using 2-dimensional polyacrylamidegel electrophoresis. The maximum decrease in T₅₀ for root tissuewas 1C for all genotypes, regardless of treatment. For ‘Alaska’shoot tips and epicotyl tissue, exogenous ABA increased thefreezing tolerance by –1.5 to –4.0C, while coldtreatment increased the freezing tolerance by –7.5 to–14.8C. Cold treatment increased the freezing toleranceof shoot tips by –9 and –15C for ‘wil’and ‘wild-type’, respectively. Cold acclimationincreased endogenous ABA concentrations in ‘Alaska’shoot tips and epicotyls 3- to 4-fold. Immunogold labeling increasednoticeably in the nucleus and cytoplasm of the epicotyl after7 d at 2C and was greatest after 30 d at the time of maximumfreezing tolerance and soluble ABA concentration. Cold treatmentinduced the production of seven, three, and two proteins inshoot, epicotyl, and root tissue of ‘Alaska’, respectively.In ‘Alaska’ shoot tissue, five out of seven novelproteins accumulated in response to both ABA and cold treatment.However, only a 24 kDa protein was produced in ‘wil’and ‘wild-type’ shoot and epicotyl tissues aftercold treatment. Abscisic acid and cold treatment additivelyincreased the freezing tolerance of pea epicotyl and shoot tissuesthrough apparently independent mechanisms that both resultedin the production of a 24 kDa protein. Key words: Pisum sativum, cold acclimation, immuno-localization     

Impedance Spectroscopy in Frost Hardiness Evaluation of Rhododendron Leaves

Impedance spectroscopy was used in studying frost hardinessof leaves of two diploid rhododendron cultivars, RhododendronL. ‘PJM’ and R. ‘Cunningham's White’,and their tetraploid derivatives, R. ‘Northern Starburst’(NSB) and CW4. After the growing season and initial hardeningin a greenhouse, plants were subjected to an acclimation regimein a phytotron: 3 consecutive weeks at +5, +1 and -2°C each.Hardiness was studied with controlled freezing tests beforeeach decrease in temperature and at the end of the experiment,based on data of extracellular resistance r_eand relaxation time of the frost-exposed leaves. The correlation of the two estimateswas 0.92. Generally, the diploid clones had better frost hardinessthan the tetraploid clones. At the end of the experiment, frosthardiness of the diploid ‘PJM’ was -28.7°C andthat of the tetraploid NSB -20.6°C. Leaves of the diploid‘Cunningham's White’ and of the tetraploid CW4 hardenedto -32.0°C and -20.9°C, respectively. Frost hardinessestimated by impedance spectroscopy correlated well with earlierresults based on visual scoring (r = 0.81–0.86) and electrolyteleakage tests (r = 0.84–0.90), but results from impedancespectroscopy indicated weaker hardiness than the other tests.The difference between the results from impedance spectroscopyand the other tests was smaller and more coherent within the‘Cunningham's White’ clones than within ‘PJM’and NSB. Changes in extracellular and intracellular resistanceof non-frozen leaves during the acclimation correlated withthe changes in frost hardiness of ‘Cunningham's White’clones, but not with those of ‘PJM’ and NSB, whichbelong to another subspecies.Copyright 2000 Annals of BotanyCompany Cold resistance, evergreen, frost hardiness, impedance spectroscopy, polyploid, Rhododendron, tetraploid     

Seasonal variations in freezing curves of stem sections of Cornus stolonifera MICHX.I

Comparisons of freezing curves have been used to determine theviability of plant parts exposed to stress. To gain understandingof the natural seasonal variations in freezing curves, uniformtwig sections of red-osier dogwood (Cornus stolonifera MICHX.)were collected throughout the year from a single clone and subjectedto controlled freezing while the tissue temperature was recorded.The supercooling of samples ranges from –2 to –7,but the variation was random and unpredictable. There was noapparent relationship between supercooling and the season ofthe year or the hardiness of the tissue. The freezing pointdepression, as estimated by the temperature of the first freezingplateau, was always between –0.25 and –1.0 andbore no relationship to hardiness or season. The freezing curveswere basically of three types: Summer and winter curves withtwo distinct freezing points; Early autumn curves with 3distinctfreezing points and spring curves with one prominent first freezingpoint which tended to mask the second freezing point. ¹Scientific Journal Series paper No. 6628, Minnesota AgriculturalExperiment Station. This research was supported in part by agrant from the Louis W. and MAUD HILL Family Foundation. ²Present Address: Horticulture Department, University of Wisconsin,Madison, Wisconsin, U.S.A.     

Freezing Tolerance in Hydrated Lactuca sativa (L) Seed: A Model to Explain Observed Variation Between Seed Lots

Low temperature tolerance was investigated in the imbibed seedof 15 seed lots compnsmg seven cultivars of Lactuca sativa L.During rapid cooling (20 °C h^–1) some seeds of allseed lots survived to –16 °C but none to –20°C. The majority of seed lots retained over 50 per centviability above –14 °C due to isolation of the embryofrom external ice by the endosperm, and subsequent embryo super-cooling.Certain seed lots, including all three seed lots of cv. TomThumb, showed high mortality at temperatures above –10°C. Correlation of mortality with the formation of externalice suggested that the endosperm is not an effective nucleationbarrier in these seed lots. Survival to –20 °C was increased at slower coolingrates (6 to 1 °C h^–1) due to freeze desiccation ofthe embryo, but seed lots varied considerably in their toleranceof specific cooling rates. A model to explain this variationwas developed incorporatmg (1) seed lot super-cooling limittemperature, (2) the rate at which freeze dehydration of thesupercooled embryo took place, (3) the moisture content at whichnucleation (at –20 °C) was no longer certain and (4)the.initial equilibrium moisture content of the fully imbibedseed. Factors (1), (2) and (3) were found to be relatively constant,but low (or artificially reduced) seed moisture content wasclosely correlated with high survival at natural cooling rates.Seed size fractions of similar moisture content from a singlecultivar showed that more small seeds survive cooling at 3 °Ch^–1 to –20 °C than larger seed. Seed with pierced endosperms or ineffective nucleation barrierswere capable of surviving to at least –10 °C if cooledslowly (1 °C h^–1) but were killed by rapid (20 °Ch^–1) cooling. Lactuca sativa L, lettuce, seed germination freezing tolerance, super-cooling     

Heat Tolerance of Cold Acclimated Puma Winter Rye Seedlings and the Effect of a Heat Shock on Freezing Tolerance

An increase in tolerance to one form of abiotic stress oftenresults in an increase in tolerance to another stress. The heattolerance of Puma rye (Secale cereale) was determined for seedlingseither not cold hardened or hardened under either controlledenvironmental or natural conditions. The heat tolerance wasdetermined either as a function of time at 42°C or the abilityto tolerate a maximum temperature. The seedlings were eithernot heat preconditioned or heat preconditioned before the heatstress. In all cases cold hardened seedlings were more heattolerant than non or partially cold hardened seedlings. Heatpreconditioning had no effect on the heat tolerance of naturallycold hardened seedlings. In contrast, seedlings cold hardenedin a controlled environment chamber, then heat preconditioned,were more heat tolerant than non preconditioned seedlings. Aheat shock of 36°C for 2 h increased the freezing toleranceof non hardened seedlings from –2.5°C to –4.5°C.Analysis of heat shock protein 70 (HSP70) gene expression indicatedthat the HSP70 gene was not induced by cold acclimation andtherefore not directly involved in the increased thermo toleranceobserved. A number of heat stable proteins, simple sugars andlong chain carbohydrate polymers accumulated during the coldacclimation process and may have a role in increasing heat toleranceas well as freezing tolerance. These data suggest cold hardeningincreases heat tolerance, however, a heat shock to non acclimatedseedlings only marginally increased the freezing tolerance ofPuma rye seedlings. ³Present address: Agriculture and Agri-Food Canada, 107 SciencePlace, Saskatoon SK S7N 0X2, Canada.     

Freeze Desiccation: a Second Mechanism for the Survival of Hydrated Lettuce (Lactuca sativa L.) Seed at Sub-zero Temperatures

Differential Thermal Analysis of hydrated lettuce cv. GreatLakes achenes using a rapid cooling rate (20 °C h^–1)produced two exotherms per achene. Both exotherms representedthe freezing of supercooled water. The high temperature exothermoccurred at –93 °C and was produced by freezing ofwater inside the pericarp but exterior to the endosperm. Thetemperature at which it occurred could be altered by the additionof nucleating agents. The low temperature exotherm produced by freezing of the embryooccurred at –162 °C and marked the death of the seed.Its temperature was not changed by the addition of nucleatingagents but its occurrence required the structural integrityof the endosperm. At low cooling rates (1 and 2 °C h^–1)low temperature exotherms were not recorded and samples removedat –25 °C had high viability. Slow cooling causeda redistribution of water within the seed whereby ice formingoutside the endosperm caused desiccation of the embryo and preventedits freezing. A mechanism is proposed, in terms of established supercoolingand nucleation theory, to explain the observed results and thevalue of freeze tolerance to the species in its natural habitatis discussed. Cooling rate, differential thermal analysis, freezing avoidance, Lactuca sativa L., lettuce, seed, supercooling, water migration     

Effect of Temperature, Light, Nutrients and Dehardening on Abscisic Acid Induced Cold Hardiness in Bromus inermis Leyss Suspension Cultured Cells

The effects of culture conditions on abscisic acid (ABA)-inducedfreezing tolerance were determined in smooth bromegrass Bromusinermis Leyss cv. Manchar) cell suspension cultures. Bromegrasscultures initiated with 2 g fr wt of cells achieved maximumfreezing tolerances (greater than –32?C) at 25 to 30?Cin the presence of 75 to 100 µM ABA. High levels of freezingtolerance induced by ABA were correlated with high growth ratesat 25 and 30?C. In control cells, incubation at 10?C inducedoptimum levels of hardiness with minimal growth. Prolonged exposure(6 weeks) of cells to 3?C, with or without ABA, increased freezingtolerance only by several degrees. Exogenous ABA concentrationsgreater than 100 µM were not inhibitory to growth. Repeatedexposure to ABA, however, retarded growth and made the cellstolerant to temperatures below –40?C. Removal of ABA fromthe medium resulted in dehardening of the cells both at 25 and3?C. Nitrogen had a marginal effect on ABA-induced hardeningat 25?C, but inhibited age-dependent hardening of control cellcultures. Light had no effect on the freezing tolerance of culturedcells. Addition of 10% sucrose, 30 min prior to freezing, tobromegrass cells treated with ABA for 4 days increased freezingtolerance more than 15?C. These observations are discussed inrelation to the contrasting behaviour of the low temperatureand photoperiod dependent cold acclimation of plants (Received July 14, 1989; Accepted October 23, 1989)     

Freezing tolerance and avoidance in high-elevation Hawaiian plants

Freezing resistance mechanisms were studied in five endemic Hawaiian species growing at high elevations on Haleakala volcano, Hawaii, where nocturnal subzero (°C) air temperatures frequently occur. Extracellular freezing occurred at around -5°C in leaves of Argyroxiphium sandwicense and Sophora chrysophylla, but these leaves can tolerate extracellular ice accumulation to -15°C and -12°C, respectively. Mucilage, which apparently acted as an ice nucleator, comprised 9 to 11% of the dry weight of leaf tissue in these two species. Leaves of Vaccinium reticulatum and Styphelia tameiameiae were also found to tolerate substantial extracellular freezing. Dubautia menziesii, on the other hand, exhibited the characteristics of permanent supercooling; a very rapid decline in liquid water content associated with simultaneous intracellular and extracellular freezing. However, in those species that tolerate extracellular freezing, the decline in liquid water content during freezing is relatively slow. Osmotic potential was lower at pre-dawn than at midday in four of the species studied. Nocturnal production of osmotically active solutes may have helped to prevent intracellular freeze dehydration as well as to provide non-colligative protection of cell membranes. Styphelia tameiameiae supercooled to -9·3°C and tolerated tissue freezing to below -15°C, a unique combination of physiological characteristics related to freezing. Tolerance of extracellular ice formation after considerable supercooling may have resulted from low tissue water content and a high degree of intracellular water binding in this species, as determined by nuclear magnetic resonance studies. The climate at high elevations in Hawaii is relatively unpredictable in terms of the duration of subzero temperatures and the lowest subzero temperature reached during the night. It appears that plants growing in this tropical alpine habitat have been under selective pressures for the evolution of freezing tolerance mechanisms.     

Supercooling along an Altitudinal Gradient in Espeletia schultzii, a Caulescent Giant Rosette Species

Rada, F., Goldstein, G., Azocar, A. and Torres, F. 1987. Supercoolingalong an altitudinal gradient in Espeletia schultzii, a caulescentgiant rosette species.—J. exp. Bot. 38: 491–497. Tropical high Andes plants may be exposed to sub-zero temperaturesany night of the year. These plants have to rely on mechanismswhich protect them from these environmental conditions but atthe same time allow their growth and development. Supercoolinghas been found to be the principal avoidance mechanism in leavesof the caulescent giant rosette genus Espeletia in the Andes.We report here the differences in supercooling capacity andcold injury in several Espeletia schultzii populations growingalong an altitudinal gradient. The relationships between supercooling,water potential and leaf anatomy were also investigated. Thesupercooling capacity increased and injury temperature decreasedfrom lower to higher elevation populations. These changes maybe explained in terms of physiological, morphological and anatomicalcharacteristics of the leaves. Key words: Espeletia schultzii, supercooling, freezing avoidance mechanisms     

Changes in Sugar Content during Cold Acclimation and Deacclimation of Cabbage Seedlings

The relationship between freezing tolerance and sugar contentin cabbage seedlings was investigated. Seedlings exposed tonon-freezing low temperature (5 °C) acquired freezing tolerancedown to -6 °C. The degree of freezing tolerance increasedwith duration of exposure to low temperature (up to 10 d). Sucrose,glucose, fructose and myo -inositol were detected as solublesugars in cabbage leaves, and all soluble sugars, except formyo -inositol, and starch increased gradually during cold acclimationsuch that their levels were positively correlated with the degreeof freezing tolerance. The induced freezing tolerance was attributednot to ontogenetic changes but to cold acclimation. However,the induced freezing tolerance was lost after only 1 d of deacclimationat control temperatures, and this change was associated witha large reduction in sugar content. These results reveal that the sugar content of cabbage leavesis positively correlated with freezing tolerance. Brassica oleracea L.; cabbage; cold acclimation; deacclimation; freezing tolerance; sugars     

Developmental control of xylem hydraulic resistances and vulnerability to embolism in Fraxinus excelsior L.: impacts on water relations

The freezing tolerance of many plants, such as pea (Pisum sativum),is increased by exposure to low temperature or abscisic acidtreatment, although the physiological basis of this phenomenonis poorly understood. The freezing tolerance of pea shoot tips,root tips, and epicotyl tissue was tested after cold acclimationat 2C, dehydration/rehydration, applications of 10^–4M abscisic acid (ABA), and deacclimation at 25C. Tests wereconducted using the cultivar ‘Alaska’, an ABA-deficientmutant ‘wil’, and its ‘wildtype’. Freezinginjury was determined graphically as the temperature that caused50% injury (T₅₀) from electrical conductivity. Endogenous ABAwas measured using an indirect enzyme-linked immunosorbant assay,and novel proteins were detected using 2-dimensional polyacrylamidegel electrophoresis. The maximum decrease in T₅₀ for root tissuewas 1C for all genotypes, regardless of treatment. For ‘Alaska’shoot tips and epicotyl tissue, exogenous ABA increased thefreezing tolerance by –1.5 to –4.0C, while coldtreatment increased the freezing tolerance by –7.5 to–14.8C. Cold treatment increased the freezing toleranceof shoot tips by –9 and –15C for ‘wil’and ‘wild-type’, respectively. Cold acclimationincreased endogenous ABA concentrations in ‘Alaska’shoot tips and epicotyls 3- to 4-fold. Immunogold labeling increasednoticeably in the nucleus and cytoplasm of the epicotyl after7 d at 2C and was greatest after 30 d at the time of maximumfreezing tolerance and soluble ABA concentration. Cold treatmentinduced the production of seven, three, and two proteins inshoot, epicotyl, and root tissue of ‘Alaska’, respectively.In ‘Alaska’ shoot tissue, five out of seven novelproteins accumulated in response to both ABA and cold treatment.However, only a 24 kDa protein was produced in ‘wil’and ‘wild-type’ shoot and epicotyl tissues aftercold treatment. Abscisic acid and cold treatment additivelyincreased the freezing tolerance of pea epicotyl and shoot tissuesthrough apparently independent mechanisms that both resultedin the production of a 24 kDa protein. Key words: Pisum sativum, cold acclimation, immuno-localization     

Cold Acclimation, Freezing Resistance and Protein Synthesis in Alfalfa (Medicago sativaL. cv. Saranac)

Mohapatra, S. S., Poole, R. J. and Dhindsa, R. S. 1987. Coldacclimation, freezing resistance and protein synthesis in alfalfa(Medicago sativa L. cv. Saranac).—J. exp. Bot. 38: 1697–1703. Changes in freezing resistance (percent survival at —10°C), pattern of protein synthesis and translatable mRNApopulation during cold acclimation of alfalfa (Medicago sativaL. cv. Saranac) have been examined. Two days of cold acclimationat 4 °C increased freezing resistance from about 6% to 40%,protein content by 200% and total RNA content by 100%. Acclimationfor longer periods did not cause further increases in freezingresistance, protein content or RNA content. Examination of proteinchanges by sodium dodecyl sulphate-polyacrylamide gel electrophoresis(SDS-PAGE) coupled with protein staining, and by fluorographyof in vivo labelled proteins separated by SDS-PAGE, showed thatseveral proteins are increasingly or newly synthesized duringcold acclimation. Analysis of in vitro translation productsby SDS-PAGE and fluorography shows changes in the populationof translatable mRNAs. It is concluded that in this varietyof alfalfa cold acclimation for only 2 d is sufficient to confermaximum freezing resistance, and that changes in proteins duringcold acclimation are regulated most probably at the transcnptionallevel. Key words: Freezing resistance, protein synthesis, cold acclimation, SDS-PAGE, Medicago sativa L.