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Acclimation of shoot and needle morphology and photosynthesis of two Picea species to differences in soil nutrient availability.

To investigate morphological acclimation to differences in nutrient availability, we compared shoot and needle morphology of Picea glehnii (Friedr. Schmidt) M. T. Mast. and Picea jezoensis (Siebold & Zucc.) Carrière trees growing on nutrient-poor volcanic ash and nutrient-rich, brown forest soil. Trees of both species were shorter and had more open canopies when growing on volcanic ash than when growing on brown forest soil. Nutrient-poor conditions limited height growth less in P. glehnii than in P. jezoensis. In both species, trees growing on volcanic ash had shorter annual increments in the previous year and more needles per shoot length and, hence, a smaller shoot silhouette area (SSA) relative to needle dry mass (NDM) than trees growing on brown forest soil. Soil type had less effect on shoot projected needle area (PNA). Total needle area (TNA) of P. glehnii shoots was similar between soil types, whereas TNA of P. jezoensis was lower in trees growing on volcanic ash than in trees growing on brown forest soil. For both species, low SSA in response to nutrient-poor conditions resulted in low shoot SSA/PNA ratios, indicating high within-shoot self-shading. Shoot SSA/TNA of P. glehnii was lower in trees growing on volcanic ash than in trees growing on brown forest soil, indicating that needles were sun-acclimated. In contrast, shoot SSA/TNA of P. jezoensis was higher in trees growing on volcanic ash than in trees growing on brown forest soil. The contrasting response of TNA to low nutrient availability was associated with species-specific differences in needle morphology. Needles of P. glehnii growing on volcanic ash were slightly shorter, wider, thicker and heavier than those of trees growing on brown forest soil, indicating morphological acclimation to high irradiance. Needles of P. jezoensis growing on volcanic ash were shorter than those of trees growing on brown forest soil, but did not show morphological acclimation to high irradiance in width, thickness or mass. For both species, nutrient-poor conditions decreased maximum photosynthetic rate (Amax) per NDM. However, when expressed per PNA, the decrease in Amax was reduced, and when expressed per SSA, Amax was higher in trees growing on volcanic ash than in trees growing on brown forest soil. On volcanic ash, Amax per NDM was lower for P. glehnii than for P. jezoensis. However, morphological changes at the shoot and needle levels reversed this trend when Amax was expressed per SSA or per PNA. The species-specific differences in morphological response to differences in soil nutrient availability suggest that P. glehnii is more tolerant of nutrient-poor conditions, whereas P. jezoensis is better at exploiting nutrient-rich soils.

Acclimatization↗

Photosynthetic acclimation to light changes in tropical monsoon forest woody species differing in adult stature.

We studied morphological and physiological leaf and whole-plant features of seedlings of six late-successional woody species common in the Xishuangbanna lowland rain forest in southwest China. Study species differed in adult stature and shade tolerance and included the shrubs Lasianthus attenuatus Jack and Lasianthus hookeri C.B. Clarke ex Hook. f.; the sub-canopy species Barringtonia macrostachya (Jack) Kurz and Linociera insignis C.B. Clarke; the canopy tree Pometia tomentosa (Blume) Teijsm. & Binn.; and the emergent species Shorea chinensis (Wang Hsie) H. Zhu. After 1 year of growth in low light (4.5% full sun), seedlings were transferred to high light (24.5% full sun) to investigate acclimation responses of existing leaves to forest gap opening and to determine whether seedling capacity for acclimation is a limiting factor in its natural regeneration. Leaves of the shrub species are shade-adapted, as indicated by their low photosynthetic capacity, efficiency in using sunflecks, low stomatal density, low Chl a/b ratio and high spongy/palisade mesophyll ratio. The shrub species utilized sunflecks efficiently because they had a short photosynthetic induction time and low induction loss. In all species, transfer of seedlings to high light resulted in a substantial initial reduction in the dark-adapted quantum yield of photosystem II (variable chlorophyll fluorescence/maximum chlorophyll fluorescence; Fv/Fm) at midday. Predawn Fv/Fm of the taller species did not change greatly, but predawn Fv/Fm of the shrub species decreased significantly without complete recovery within 25 days of transfer to high light, indicating chronic photoinhibition and damage to the previously shade-adapted leaves. Maximum net photosynthetic rate and dark respiration of the four taller species increased considerably after transfer to high light, but not in the shrub species. Similar trends were observed for the number of newly formed leaves and relative height growth rate. We conclude that the shrubs L. hookeri and L. attenuatus have limited potential for developmental and physiological acclimation to high light, which explains their absence from forest gaps. Compared with the shrub species, the taller tree species, which are more likely to experience high light during their life span, showed a greater potential for light acclimation. Physiological differences among the four tree species were not consistent with differences in adult stature.

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Excitation energy partitioning and quenching during cold acclimation in Scots pine.

We studied the influence of two irradiances on cold acclimation and recovery of photosynthesis in Scots pine (Pinus sylvestris L.) seedlings to assess mechanisms for quenching the excess energy captured by the photosynthetic apparatus. A shift in temperature from 20 to 5 degrees C caused a greater decrease in photosynthetic activity, measured by chlorophyll fluorescence and oxygen evolution, in plants exposed to moderate light (350 micromol m(-2) s(-1)) than in shaded plants (50 micromol m(-2) s(-1)). In response to the temperature shift, maximal photochemical efficiency of photosystem II (PSII), measured as the ratio of variable to maximal chlorophyll fluorescence (Fv/Fm) of dark-adapted samples, decreased to 70% in exposed seedlings, whereas shaded seedlings maintained Fv/Fm close to initial values. After a further temperature decrease to -5 degrees C, only 8% of initial Fv/Fm remained in exposed plants, whereas shaded plants retained 40% of initial Fv/Fm. Seven days after transfer from -5 to 20 degrees C, recovery of photochemical efficiency was more complete in the shaded plants than in the exposed plants (87 and 65% of the initial Fv/Fm value, respectively). In response to cold stress, the estimated functional absorption cross section per remaining PSII reaction center increased at both irradiances, but the increase was more pronounced in exposed seedlings. Estimates of energy partitioning in the needles showed a much higher dissipative component in the exposed seedlings at low temperatures, pointing to stronger development of non-photochemical quenching at moderate irradiances. The de-epoxidation state of the xanthophyll cycle pigments increased in exposed seedlings at 5 degrees C, contributing to the quenching capacity, whereas significant de-epoxidation in the shaded plants was observed only when temperatures decreased to -5 degrees C. Thermoluminescence (TL) measurements of PSII revealed that charge recombinations between the second oxidation state of Mn-cluster S2 and the semireduced secondary electron acceptor quinone Q(B)- (S2Q(B)-) were shifted to lower temperatures in cold-acclimated seedlings compared with control seedlings and this effect depended on irradiance. Concomitant with this, cold-acclimated seedlings demonstrated a significant shift in the S2 recombination with primary acceptor Q(A)- (S2Q(A)-) characteristic TL emission peak to higher temperatures, thus narrowing the redox potential gap between S2Q(B)- and S2Q(A)-, which might result in increased probability for non-radiative radical pair recombination between the PSII reaction center chlorophyll a (P680+) and Q(A)- (P680+)Q(A)-) (reaction center quenching) in cold-acclimated seedlings. In Scots pine seedlings, mechanisms of quenching excess light energy in winter therefore involve light-dependent regulation of reaction center content and both reaction center-based and antenna-based quenching of excess light energy, enabling them to withstand high excitation pressure under northern winter conditions.

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Spatial Distribution of Photosynthesis during Drought in Field-Grown and Acclimated and Nonacclimated Growth Chamber-Grown Cotton.

Inhomogeneous photosynthetic activity has been reported to occur in drought-stressed leaves. In addition, it has been suggested that these water stress-induced nonuniformities in photosynthesis are caused by "patchy" stomatal closure and that the phenomenon may have created the illusion of a nonstomatal component to the inhibition of photosynthesis. Because these earlier studies were performed with nonacclimated growth chamber-grown plants, we sought to determine whether such "patches" existed in drought-treated, field-grown plants or in chamber-grown plants that had been acclimated to low leaf water potentials (psi(leaf)). Cotton (Gossypium hirsutum L.) was grown in the field and subjected to drought by withholding irrigation and rain from 24 d after planting. The distribution of photosynthesis, which may reflect the stomatal aperture distribution in a heterobaric species such as cotton, was assayed by autoradiography after briefly exposing attached leaves of field-grown plants to (14)CO(2). A homogeneous distribution of radioactive photosynthate was evident even at the lowest psi(leaf) of -1.34 MPa. "Patchiness" could, however, be induced by uprooting the plant and allowing the shoot to air dry for 6 to 8 min. In parallel studies, growth chamber-grown plants were acclimated to drought by withholding irrigation for three 5-d drought cycles interspersed with irrigation. This drought acclimation lowered the psi(leaf) value at which control rates of photosynthesis could be sustained by approximately 0.7 MPa and was accompanied by a similar decline in the psi(leaf) at which patchiness first appeared. Photosynthetic inhomogeneities in chamber-grown plants that were visible during moderate water stress and ambient levels of CO(2) could be largely removed with elevated CO(2) levels (3000 muL L(-1)), suggesting that they were stomatal in nature. However, advanced dehydration (less than approximately 2.0 MPa) resulted in "patches" that could not be so removed and were probably caused by nonstomatal factors. The demonstration that patches do not exist in drought-treated, field-grown cotton and that the presence of patches in chamber-grown plants can be altered by treatments that cause an acclimation of photosynthesis leads us to conclude that spatial heterogeneities in photosynthesis probably do not occur frequently under natural drought conditions.

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cDNA sequence, expression, and transcript stability of a cold acclimation-specific gene, cas18, of alfalfa (Medicago falcata) cells.

The nucleotide sequence of a full-length cDNA, the deduced amino acid sequence, and the regulation of expression of a cold acclimation-specific gene, cas18, in cell suspension cultures of a freezing-tolerant cultivar of alfalfa (Medicago falcata cv Anik) have been determined. The deduced polypeptide, CAS18, is relatively small (17.6 kD), is highly hydrophilic, is rich in glycine and threonine, and contains two distinctive repeat elements. It exhibits homology with members of the LEA/RAB/dehydrin family of proteins, which accumulate in response to abscisic acid (ABA) or water stress. It is intriguing that cas18 is induced by neither ABA nor water stress. The cas18 cDNA hybridizes to three transcripts of 1.6, 1.4, and 1.0 kb, and the cDNA characterized here corresponds to the 1.0-kb transcript. The expression of this gene is about 30-fold greater in cold-acclimated cells than in nonacclimated cells. Although the accumulation of transcripts during cold acclimation is relatively slow, their disappearance during deacclimation is dramatically rapid, becoming undetectable in less than 5 h. Studies of nuclear run-on transcription show that cold acclimation enhances the transcription of this gene nearly 9-fold. The stability of cas18-detectable transcripts during deacclimation is considerably increased if transcription is inhibited with cordycepin. It therefore appears that low temperature regulates the expression of cas18 at both the transcriptional and posttranscriptional levels.

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Recovery from Photoinhibition in Peas (Pisum sativum L.) Acclimated to Varying Growth Irradiances (Role of D1 Protein Turnover).

D1 protein turnover and restoration of the photochemical efficiency of photosystem II (PSII) after photoinhibition of pea leaves (Pisum sativum L. cv Greenfeast) acclimated to different light intensities were investigated. All peas acclimated to different light intensities were able to recover from photoinhibition, at least partially, at light intensities far above their growth light irradiance. However, the capacity of pea leaves to recover from photoinhibition under increasing high irradiances was strictly dependent on the light acclimation of the leaves; i.e. the higher the irradiance during growth, the better the capacity of pea leaves to recover from photoinhibition at moderate and high light. In our experimental conditions, mainly D1 protein turnover-dependent recovery was monitored, since in the presence of an inhibitor of chloroplast-encoded protein synthesis, lincomycin, only negligible recovery took place. In darkness, neither the restoration of PSII photochemical efficiency nor any notable degradation of damaged D1 protein took place. In low light, however, good recovery of PSII occurred in all peas acclimated to different light intensities and was accompanied by fast degradation of the D1 protein. The rate of degradation of the D1 protein was estimated to be 3 to 4 times faster in photoinhibited leaves than in nonphotoinhibited leaves under the recovery conditions of 50 [mu]mol of photons m-2 s-1. In moderate light of 400 [mu]mol of photons m-2 s-1, the photoinhibited low-light peas were not able to increase further the rate of D1 protein degradation above that observed in nonphotoinhibited leaves, nor was the restoration of PSII function possible. On the other hand, photoinhibited high-light leaves were able to increase the rate of D1 protein degradation above that of nonphotoinhibited leaves even in moderate and high light, ensuring at least partial restoration of PSII function. We conclude that the capacity of photoinhibited leaves to restore PSII function at different irradiances was directly related to the capacity of the leaves to degrade damaged D1 protein under the recovery conditions.

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Structural organization of the spinach endoplasmic reticulum-luminal 70-kilodalton heat-shock cognate gene and expression of 70-kilodalton heat-shock genes during cold acclimation.

The 70-kD heat-shock proteins (HSP70s) are encoded by a multigene family in eukaryotes. In plants, the 70-kD heat-shock cognate (HSC70) proteins are located in organellar and cytosolic compartments of cells in most tissues. Previous work has indicated that HSC70 proteins of spinach (Spinacia oleracea) are actively synthesized during cold-acclimating conditions. We have isolated, sequenced, and characterized cDNA and genomic clones for the endoplasmic reticulum (ER) luminal HSC70 protein (immunoglobulin heavy chain-binding protein; BiP) of spinach. The spinach ER-luminal HSC70 is a constitutively expressed gene consisting of eight exons. Spinach BiP mRNA appears to be up-regulated during cold acclimation but is not expressed during water stress or heat shock. In contrast to the differential regulation of mRNA, the ER-luminal HSC70 protein levels remain constant in response to various environmental stresses. Two other members of the spinach 70-kD heat-shock (HS70) multigene family also show differential expression in response to a variety of environmental stresses. A constitutively expressed cytosolic HSC70 protein in spinach appears also to be up-regulated in response to both cold-acclimating and heat-shock treatments. Spinach also contains a cold-shock-induced HS70 gene that is not expressed during heat shock or water stress. Since HSP70s are considered to be involved with the chaperoning and folding of proteins, the data further support the concept that they may be important for maintaining cellular homeostasis and proper protein biogenesis during cold acclimation of spinach.

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Improved Cytoplasmic pH Regulation, Increased Lactate Efflux, and Reduced Cytoplasmic Lactate Levels Are Biochemical Traits Expressed in Root Tips of Whole Maize Seedlings Acclimated to a Low-Oxygen Environment.

We tested the hypothesis (J.-H. Xia and P.H. Saglio [1992] Plant Physiol 100: 40-46) that the enhanced ability of maize (Zea mays) root tips to survive anoxia, elicited by a 4-h exposure to 3% O2 ("acclimation"), is due to less cytoplasmic acidosis early in anoxia. Cytoplasmic pH and fermentation reactions were monitored in excised and intact (attached) maize root tips by simultaneous in vivo 13C- and 31P-NMR spectroscopy. We demonstrate that both excised and intact acclimated root tips have significantly higher cytoplasmic pH values under anoxia. This reduction in cytoplasmic acidosis is greater in intact root tips. Remarkably, cytoplasmic pH does not change when root tips are transferred from 3% O2 to anoxia. The earlier observation of considerable lactate efflux and lowered intracellular lactate in excised, acclimated root tips (ibid.) was extended to intact seedlings. The predominant fermentation end product retained in the cells of acclimated root tips is alanine. We discuss the relationship between cytoplasmic pH and levels of intracellular lactate and alanine in sugar-replete roots, and the role of cytoplasmic pH in determining survival under anoxia.

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Parallel changes in H2O2 and catalase during thermotolerance induced by salicylic acid or heat acclimation in mustard seedlings

Spraying mustard (Sinapis alba L.) seedlings with salicylic acid (SA) solutions between 10 and 500 &mgr;m significantly improved their tolerance to a subsequent heat shock at 55 degreesC for 1.5 h. The effects of SA were concentration dependent, with higher concentrations failing to induce thermotolerance. The time course of thermotolerance induced by 100 &mgr;m SA was similar to that obtained with seedlings acclimated at 45 degreesC for 1 h. We examined the hypothesis that induced thermotolerance involved H2O2. Heat shock at 55 degreesC caused a significant increase in endogenous H2O2 and reduced catalase activity. A peak in H2O2 content was observed within 5 min of either SA treatment or transfer to the 45 degreesC acclimation temperature. Between 2 and 3 h after SA treatment or heat acclimation, both H2O2 and catalase activity significantly decreased below control levels. The lowered H2O2 content and catalase activity occurred in the period of maximum thermoprotection. It is suggested that thermoprotection obtained either by spraying SA or by heat acclimation may be achieved by a common signal transduction pathway involving an early increase in H2O2.

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Does a low nitrogen supply necessarily lead to acclimation of photosynthesis to elevated CO2?

Long-term exposure of plants to elevated partial pressures of CO2 (pCO2) often depresses photosynthetic capacity. The mechanistic basis for this photosynthetic acclimation may involve accumulation of carbohydrate and may be promoted by nutrient limitation. However, our current knowledge is inadequate for making reliable predictions concerning the onset and extent of acclimation. Many studies have sought to investigate the effects of N supply but the methodologies used generally do not allow separation of the direct effects of limited N availability from those caused by a N dilution effect due to accelerated growth at elevated pCO2. To dissociate these interactions, wheat (Triticum aestivum L.) was grown hydroponically and N was added in direct proportion to plant growth. Photosynthesis did not acclimate to elevated pCO2 even when growth was restricted by a low-N relative addition rate. Ribulose-1, 5-bisphosphate carboxylase/oxygenase activity and quantity were maintained, there was no evidence for triose phosphate limitation of photosynthesis, and tissue N content remained within the range recorded for healthy wheat plants. In contrast, wheat grown in sand culture with N supplied at a fixed concentration suffered photosynthetic acclimation at elevated pCO2 in a low-N treatment. This was accompanied by a significant reduction in the quantity of active ribulose-1, 5-bisphosphate carboxylase/oxygenase and leaf N content.

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Overexpression of the Arabidopsis CBF3 transcriptional activator mimics multiple biochemical changes associated with cold acclimation.

We further investigated the role of the Arabidopsis CBF regulatory genes in cold acclimation, the process whereby certain plants increase in freezing tolerance upon exposure to low temperature. The CBF genes, which are rapidly induced in response to low temperature, encode transcriptional activators that control the expression of genes containing the C-repeat/dehydration responsive element DNA regulatory element in their promoters. Constitutive expression of either CBF1 or CBF3 (also known as DREB1b and DREB1a, respectively) in transgenic Arabidopsis plants has been shown to induce the expression of target COR (cold-regulated) genes and to enhance freezing tolerance in nonacclimated plants. Here we demonstrate that overexpression of CBF3 in Arabidopsis also increases the freezing tolerance of cold-acclimated plants. Moreover, we show that it results in multiple biochemical changes associated with cold acclimation: CBF3-expressing plants had elevated levels of proline (Pro) and total soluble sugars, including sucrose, raffinose, glucose, and fructose. Plants overexpressing CBF3 also had elevated P5CS transcript levels suggesting that the increase in Pro levels resulted, at least in part, from increased expression of the key Pro biosynthetic enzyme Delta(1)-pyrroline-5-carboxylate synthase. These results lead us to propose that CBF3 integrates the activation of multiple components of the cold acclimation response.

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Cold Acclimation of Hedera helix: Evidence for a Two Phase Process.

The light-enhanced production and accumulation of sugars is only one step in the process of cold acclimation in Hedera helix L. var. Thorndale (English ivy). Applications of 2,4-dinitrophenol to plants with different portions exposed to light and dark indicated that the mere presence or accumulation of the light-generated promoters did not invoke an increase in hardiness. Kinetics of cold acclimation during alternating periods of light and dark also indicate that the light stimulation of cold acclimation is only a partial component of the total process. Incubation on 50 mm solutions of sucrose can replace the light requirement. A second phase which can proceed in the dark is thought to result in the production of proteins which, due to an altered composition or configuration, have a greater capacity to bind sugars. This is evidenced by the fact that protein from cold acclimated tissue exhibited a higher sugar-binding capacity than protein from nonacclimated tissue. Furthermore, the two phases can proceed independently of each other, but only upon complementation of the products of the two phases is an increase in cold hardiness manifested.

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Phytochrome control of growth cessation and initiation of cold acclimation in selected woody plants.

Short day enhancement of cold acclimation in twigs of Cornus (red-osier dogwood), Weigela, and Pyracantha (firethorn) was studied using dark interruptions with red or with red-far red radiation. Hardiness was estimated by freezing stem tissues to preselected temperatures and evaluating injury electrolytically. Dark-period interruptions with red radiation suppressed cold acclimation in Cornus and Weigela. When red light was followed by far red light, suppression was relieved. No radiation control of acclimation was found with Pyracantha. The short day enhancement of cold acclimation in Cornus and Weigela appears to be phytochrome-mediated.

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Effects of Red and Far Red Light on the Initiation of Cold Acclimation in Cornus stolonifera Michx.

Red and far red light distinctly influence the initial phytochrome-mediated phase of cold acclimation in red-osier dogwood (Cornus stolonifera). Under controlled conditions, short days and end-of-day far red light exposure after long days promote growth cessation, cold acclimation, and subsequent cold hardening of dogwood stems in response to low temperature. Nuclear magnetic resonance absorption spectra of the water in internode stem sections imply that the short day-induced phase of cold acclimation involves a change in tissue hydration, at least in part, due to a substantial reduction in bulk phase water as a result of senescence and loss of water from the pith. Seasonal responses to light and an attempt to induce early acclimation under natural conditions with end-of-day far red light are discussed.

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Plasma Membrane Alterations in Callus Tissues of Tuber-bearing Solanum Species during Cold Acclimation.

Plasma membrane alterations in two tuber-bearing potato species during a 20-day cold acclimation period were investigated. Leaf-callus tissues of the frost-resistant Solanum acaule Hawkes ;Oka 3878' and the frost-susceptible, commonly grown Solanum tuberosum ;Red Pontiac,' were used. The former is a species that can be hardened after subjecting to the low temperature, and the latter does not harden. Samples for the electron microscopy were prepared from callus cultures after hardening at 2 C in the dark for 0, 5, 10, 15, and 20 days. After 20 days acclimation, S. acaule increased in frost hardiness from -6 to - 9 C (killing temperature), whereas frost hardiness of S. tuberosum remained unchanged (killed at -3 C). Actually, after 15 days acclimation, a -9 C frost hardiness level in S. acaule callus cultures had been achieved.Membrane protein particle aggregation was monitored using freeze-fracture electron microscopy. Protein particles were aggregated in S. acaule up to 10 days after the initiation of acclimation treatment and then redistributed almost to the level of control after 15 days. No such changes were observed for S. tuberosum under similar experimental conditions. The change in protein particle aggregation pattern in S. acaule is interpreted as indicating the presence of an adaptive fluidity control mechanism in that species.

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Involvement of abscisic Acid in potato cold acclimation.

Upon exposure to 2 degrees C day/night (D/N), leaves of Solanum commersonii (Sc) began acclimating on the 4th day from a -5 degrees C (killing temperature) hardy level to -12 degrees C by the 15th day. Leaves of S. tuberosum L. (St) cv ;Red Pontiac' typically failed to acclimate and were always killed at -3 degrees C. Leaves of control (20/15 degrees C, D/N) and treated plants (2 degrees C, D/N) of St showed similar levels of free abscisic acid (ABA) during a 15-day sampling period. In treated Sc plants, however, free ABA contents increased 3-fold on the 4th day and then declined to their initial level thereafter. The increase was not observed in leaves of Sc control plants.Treated St plants showed a slightly higher content of leaf soluble protein than controls. In Sc, leaves of controls maintained relatively constant soluble proteins, but leaves of treated plants showed a distinct increase. This significant increase was initiated on the 4th day, peaked on the 5th day, and remained at a high level throughout the 15-day sampling period.Exogenously applied ABA induced frost hardiness in leaves of Sc plants whether plants were grown under a 20 degrees C or 2 degrees C temperature regime. When cycloheximide was added to the medium of stem-cultured plants at the beginning of 2 degrees C acclimation, or at the beginning of the ABA treatment in the 20 degrees C regime, it completely inhibited the development of frost hardiness. However, when cycloheximide was added to plants on the 5th day during 2 degrees C acclimation, the induction of frost hardiness was not inhibited. The role of ABA in triggering protein synthesis needed to induce frost hardiness is discussed.

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Regulation of RNA Synthesis by DNA-Dependent RNA Polymerases and RNases during Cold Acclimation in Winter and Spring Wheat.

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.

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Changes in Protein Patterns and Translatable Messenger RNA Populations during Cold Acclimation of Alfalfa.

Changes in the rate and pattern of protein synthesis and in translatable mRNA population during cold acclimation of alfalfa (Medicago falcata cv Anik) seedlings have been examined. There appears to be a positive correlation between the increase in ability to synthesize proteins at 4 degrees C and the increase in freezing resistance (survival at -10 degrees C). Results obtained with three different approaches using sodium dodecyl sulfate-polyacrylamide gel electrophoresis pattern visualized by (a) staining, (b) immunoblotting and autoradiography, and (c) fluorography of in vivo labeled proteins, show that at least eight polypeptides are newly synthesized during cold acclimation. Results of analysis of in vitro translation products of mRNA from nonacclimated and acclimated seedlings show the appearance of new translatable mRNAs. It is concluded that changes in gene expression occur during cold acclimation, most probably at the transcriptional level.

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