Membrane Lipid Changes During Cold Acclimation of Spring Rape

238

192

The abilities of two races ofArabidopsis thaliana L. (Heyn), Landsberg erecta and Columbia, to cold harden were examined. Landsberg, grown at 22 to 24°C, increased in freezing tolerance from an initial 50% lethal temperature (LT5o) of about -3°C to an LT5o of about -6°C after 24 hours at 4°C; LT5o values of -8 to -10°C were achieved after 8 to 9 days at 4°C. Similar increases in freezing tolerance were obtained with Columbia. In vitro translation of poly(A ) RNA isolated from control and coldtreated Columbia showed that low temperature induced changes in the population of translatable mRNAs. An mRNA encoding a polypeptide of about 160 kilodaltons (isoelectric point about 4.5) increased markedly after 12 to 24 h at 4°C, as did mRNAs encoding four polypeptides of about 47 kilodaltons (isoelectric points ranging from 5-5.5). Incubation of Columbia callus tissue at 4°C also resulted in increased levels of the mRNAs encoding the 160 kilodalton polypeptide and at least two of the 47 kilodalton polypeptides. In vivo labeling experiments using Columbia plants and callus tissue indicated that the 160 kilodalton polypeptide was synthesized in the cold and suggested that at least two of the 47 kilodalton polypeptides were produced. Other differences in polypeptide composition were also observed in the in vivo labeling experiments, some of which may be the result of posttranslational modifications of the 160 and 47 kilodalton polypeptides.Many plants become more resistant to freezing temperatures when first exposed to low nonfreezing temperatures, a process known as cold acclimation or cold hardening (11,23 (25). Plants were maintained in a controlled environment chamber at 22 to 24°C under constant illumination from cool white fluorescent lights. Light intensity was approximately 100 ,uE m 2 s-' PAR at soil level, and the RH was about 40 to 50%. Plants were subirrigated as necessary with distilled water. Cold treatment was at 4°C under constant illumination from cool-white fluorescent lights with light intensity ranging from approximately 20 to 50 ,uE m-2 s-' PAR, and the RH was between 75 and 95%. Experiments were performed before the plants had started to bolt, generally about 19 d after planting.For RNA extraction, rosette leaves and stems were excised at soil level, frozen in liquid N2, pulverized using a mortar and pestle, and stored at -80°C prior to extraction.Callus Culture. Arabidopsis (Columbia) seeds were surfacesterilized and placed on semisolid MSa2 medium (MSal) (17) and containing 3% (w/v)

Section: Discussionsupporting

confidence: 79%

Cold Acclimation in Arabidopsis thaliana

Gilmour¹,

Hajela²,

Thomashow³

1988

238

192

“…Thus, it is suggested that the membranes have to undergo certain changes during cold acclimation in order to withstand freezing stress. There are several reports on membrane changes during cold acclimation (2,3,9,11,16). However, most of these studies focused on changes in chemical and biophysical properties such as composition and fluidity of the lipid component of membranes (3,11,16).…”

mentioning

confidence: 99%

“…There are several reports on membrane changes during cold acclimation (2,3,9,11,16). However, most of these studies focused on changes in chemical and biophysical properties such as composition and fluidity of the lipid component of membranes (3,11,16). Altered pattern of synthesis of membrane proteins has not received much attention (15,16).…”

mentioning

confidence: 99%

Alterations in Membrane Protein-Profile during Cold Treatment of Alfalfa

Mohapatra¹,

Poole²,

Dhindsa³

1988

“…Thus, it follows that plant plasma membranes must undergo certain chemical and biophysical changes during cold acclimation in order to withstand freezing stresses. There are several reports on changes in cellular membranes during cold acclimation (6,7,27). However, these are based on analytical data obtained from crude membrane fractions or total membrane extracts.…”

mentioning

confidence: 99%

Involvement of Plasma Membrane Alterations in Cold Acclimation of Winter Rye Seedlings (Secale cereale L. cv Puma)

Uemura¹,

Yoshida²

1984

142

There is mounting evidence to indicate that membranes, primarily the plasma membrane, is intimately involved in cold acclimation and freezing injury (11,18,19,(24)(25)(26). Thus, it follows that plant plasma membranes must undergo certain chemical and biophysical changes during cold acclimation in order to withstand freezing stresses.There are several reports on changes in cellular membranes during cold acclimation (6,7,27). However, these are based on analytical data obtained from crude membrane fractions or total membrane extracts. Therefore, it is uncertain whether the reported changes are related to the changes in the plasma membranes per se during cold acclimation. Based on these facts, it is important to know what changes in the plasma membranes are responsible for the development offreezing tolerance during cold acclimation. A major problem encountered has been the difficulty ofisolating the plasma membrane in a high degree ofpurity needed for detailed studies. Recently, we have succeeded in preparing plasma membranes from light-grown winter rye seedlings with a high degree of purity using an aqueous two-polymer phase partition system (29).In the present study, Evaluation of Freezing Resistance. Freezing tolerance of the crown was evaluated by the ion-leakage test. One g fresh weight of tissues were cut into 5-mm-long pieces and were frozen in test tubes at -3°C for 2 h with small pieces of ice. Thereafter, the tissues were cooled in steps of 5°C at 2-h intervals and held at the desired temperatures for 2 h. After thawing at OC, the tissues were immersed in 5 ml of distilled H20 and incubated at room temperature for more than 5 h by gently shaking before measuring the conductivity. The tissues frozen directly in the liquid nitrogen and unfrozen were taken as 100% and 0% in the relative conductivity, respectively.Isolation of Plasma Membranes. Crown tissues were homogenized with a Polytron PT 20 at the medium speed for 90 s in a medium consisting of 0.5 M sorbitol, 50 mm MOPS2-KOH (pH 7.6), 10 mm EGTA neutralized with KOH, 2.5 mm potassium metabisulfite, 4 mM SHAM, 1 mm PMSF, 5% (w/v) soluble PVP (mol wt 24,500), and 1% (w/v) defatted BSA. The homogenate was passed through two layers of gauze and then two layers of Miracloth, and then centrifuged at 14,000g for 15 min and 156,000g for 20 min. The 156,000g pellet (14,000-156,000g fraction) was used for plasma membrane isolation using the aqueous two-polymer phase partition technique as reported previously (29). The purity of the isolated plasma membrane was determined by phosphotungstate-chromate staining and by specific aggregation in the presence ofZnCl2 or at pH 4.5 as reported previously (29). The isolated plasma membranes were suspended in 0.5 M sorbitol-3 mm Tris-maleate at pH 7.3 and kept at -70°C until use.Lipid Extraction. Total lipids were extracted from membrane samples according to the method of Bligh and Dyer (1) with the 2 Abbreviations: MOPS, 34N-morpholino)propanesulfonic acid;