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Low temperature treatment at the young microspore stage induces protein changes in rice anthers.

Male reproductive development in rice is very sensitive to various forms of environmental stresses including low temperature. A few days of cold treatment (<20 degrees C) at the young microspore stage induce severe pollen sterility and thus large grain yield reductions. To investigate this phenomenon, anther proteins at the early stages of microspore development, with or without cold treatment at 12 degrees C, were extracted, separated by two-dimensional gel electrophoresis, and compared. The cold-sensitive cultivar Doongara and the relatively cold-tolerant cultivar HSC55 were used. The abundance of 37 anther proteins was changed more than 2-fold after 1, 2, and 4 days of cold treatment in cv. Doongara. Among them, one protein was newly induced, 32 protein spots were up-regulated, and four protein spots were down-regulated. Of these 37 protein spots, we identified two anther-specific proteins (putative lipid transfer protein and Osg6B) and a calreticulin that were down-regulated and a cystine synthase, a beta-6 subunit of the 20 S proteasome, an H protein of the glycine cleavage system, cytochrome c oxidase subunit VB, an osmotin protein homologue, a putative 6-phosphogluconolactonase, a putative adenylate kinase, a putative cysteine proteinase inhibitor, ribosomal protein S12E, a caffeoyl-CoA O-methyltransferase, and a monodehydroascorbate reductase that were up-regulated. Identification of these proteins is available upon request. Accumulation of these proteins did not vary greatly after cold treatment in panicles of cv. Doongara or in the anthers of the cv. HSC55. The newly induced protein named Oryza sativa cold-induced anther protein (OsCIA) was identified as an unknown protein. The OsCIA protein was detected in panicles, leaves, and seedling tissues under normal growth conditions. Quantitative real time RT-PCR analysis of OsCIA mRNA expression showed no significant change between low temperature-treated and untreated plants. A possible regulatory role for the newly induced protein is proposed.

Amino Acid Sequence↗

Muscle glycogen availability and temperature regulation in humans.

The effects of intramuscular glycogen availability on human temperature regulation were studied in eight seminude subjects immersed in 18 degrees C water for 90 min or until rectal temperature (Tre) decreased to 35.5 degrees C. Each subject was immersed three times over a 3-wk period. Each immersion followed 2.5 days of a specific dietary and/or exercise regimen designed to elicit low (L), normal (N), or high (H) glycogen levels in large skeletal muscle groups. Muscle glycogen concentration was determined in biopsies taken from the vastus lateralis muscle before and after each immersion. Intramuscular glycogen concentration before the immersion was significantly different among the L, N, and H trials (P less than 0.01), averaging 247 +/- 15, 406 +/- 23, and 548 +/- 42 (SE) mmol glucose units.kg dry muscle-1, respectively. The calculated metabolic heat production during the first 30 min of immersion was significantly lower during L compared with N or H (P less than 0.05). The rate at which Tre decreased was more rapid during the L immersion than either N or H (P less than 0.05), and the time during the immersion at which Tre first began to decrease also appeared sooner during L than N or H. The results suggest that low skeletal muscle glycogen levels are associated with more rapid body cooling during water immersion in humans. Higher than normal muscle glycogen levels, however, do not increase cold tolerance.

Adult↗

Biotechnological applications of plant freezing associated proteins.

Plants use a wide array of proteins to protect themselves against low temperature and freezing conditions. The identification of these freezing tolerance associated proteins and the elucidation of their cryoprotective functions will have important applications in several fields. Genes encoding structural proteins, osmolyte producing enzymes, oxidative stress scavenging enzymes, lipid desaturases and gene regulators have been used to produce transgenic plants. These studies have revealed the potential capacity of different genes to protect against temperature related stresses. In some cases, transgenic plants with significant cold tolerance have been produced. Furthermore, the biochemical characterization of the cold induced antifreeze proteins and dehydrins reveals many applications in the food and the medical industries. These proteins are being considered as food additives to improve the quality and shelf-life of frozen foods, as cryoprotective agents for organ and cell cryopreservation, and as chemical adjuvant in cancer cryosurgery.

Adaptation, Biological↗

General and local cold responses in humans after 2 weeks at high altitude.

To investigate the effects of a short-term high altitude residence (2 weeks between 4150 m and 6885 m in the Andes) on the general and local cold responses after descent, 11 subjects were submitted both to a whole body standard cold air test (SCAT, dry bulb temperature = 1 degree C, 2 h, nude, at rest) and to a local cold water test of the right upper limb (CWT, 5 degrees C, 5 min) both before and after the expedition. Compared to before the expedition, a lower systolic blood pressure was observed after the high altitude residence [130.00 (SEM 3.35) mm Hg vs 140.40 (SEM 4.74) mm Hg at the end of CWT, P < 0.05] whereas no significant change either in diastolic blood pressure or in heart rate was found. All skin temperatures of the right upper limb were lowered (P < 0.05). During SCAT, body temperatures were unchanged (rectal and mean skin temperature, Tsk) but metabolic heat production was slightly but significantly diminished [110 (SEM 4) W.m-2 vs 125 (SEM 3) W.m-2, P < 0.05] and heat debt increased [11.37 (SEM 1.11) kJ.kg-1 vs 9.30 (SEM 2.30) kJ.kg-1, P < 0.05]. Moreover, the time of onset of continuous shivering (d) was shortened [8.20 (SEM 1.90) min vs 17.30 (SEM 3.60) min, P < 0.05] and the level of Tsk observed at (d) was higher [25.70 (SEM 0.80) degrees C vs 23.57 (SEM 0.78) degrees C, P < 0.05] suggesting an increase in the sensitivity of the thermoregulatory system despite the slight decreased shivering activity observed. It was concluded that general and local cold tolerance were modified by a short-term residence at altitude and that the changes observed were not in accordance with general or (and) local cold adaptation. In contrast, high altitude sojourn could be a risk factor for frostbite of the extremities.

Altitude↗

Effect of dietary linoleic acid and essential fatty acid deficiency on resting metabolism, nonshivering thermogenesis and brown adipose tissue in the rat.

Rats were fed a diet either deficient (0.05%) in essential fatty acids (EFA), or providing 4% (control) and 10% (surplus) of the total energy intake in the form of linoleic acid. All diets were isoenergetic and provided 13.9% of the energy as fat. The rats were kept at 29 or 5 degrees C. Growth and food intake of rats fed linoleic acid surplus at either temperature for 10 wk were not different from that of controls; basal metabolism, norepinephrine-induced nonshivering thermogenesis (NST) and thermogenic variables in the brown adipose tissue (amount of mitochondria and mitochondrial uncoupling protein) also were not different. The effects of EFA deficiency were drastically enhanced in the cold: After 10 wk of consuming a diet low in EFA at 5 degrees C, the body weight of rats was 75% of that of controls (87% at 29 degrees C); the food intake was 135% of controls at 5 degrees C (120% at 29 degrees C). The resting respiration in deficient rats was 125% of controls at 5 degrees C (110% at 29 degrees C); body temperatures as low as 35.1 degrees C were measured in deficient rats after 3 wk at 5 degrees C; the cold tolerance of the rats was significantly diminished (30% died within 3 wk at 5 degrees C), thus emphasizing the essential role of dietary EFA during cold stress. Norepinephrine-induced NST and the thermogenic parameters in brown fat were not influenced by EFA deficiency.(ABSTRACT TRUNCATED AT 250 WORDS)

Adipose Tissue, Brown↗

Transposon mutagenesis reveals novel loci affecting tolerance to salt stress and growth at low temperature.

Using transposon mutagenesis in the haploid Saccharomyces cerevisiae strain W303-1A we have identified genes required for growth in high salt medium, survival of a hypo-osmotic shock and growth at 15 degrees C. Screening 25,000 transposon insertions revealed a total of 61 insertions that caused salt-sensitivity; and those insertions affected 31 genes. Only 12 of those genes were previously known to be required for salt-tolerance. Among the 61 insertions, three caused general osmo-sensitivity. We identified one single insertion mutant in the already-known gene, FPS1, required for survival of hypo-osmotic shock. A total of 31 insertions caused failure to grow at low temperature. Those identified ten different genes, three of which had previously been reported to affect cold-tolerance. Four genes were identified in both the salt and the cold-sensitivity screen. We found several unusual insertion mutations: (1) insertions in or close to essential genes, (2) insertion in an intergenic region and (3) insertions causing stress-sensitivity in W303-1A, while the deletion mutant in BY4741 did not show such a phenotype. Surprisingly, our mutant set and that reported in the large-scale transposon insertion project (TRIPLES, http://ygacmed.yale.edu/triples/triples.htm) only marginally overlap. We discuss some of the features of transposon mutagenesis in light of the availability of the complete set of yeast deletion mutants and we discuss the possible roles of the genes we identified.

Base Sequence↗

Glycogen and nonspecific adaptation to cold.

Exposure to moderate cold for a few weeks causes adaptation through the development of nonshivering thermogenesis primarily in the brown adipose tissue. Exposure to severe cold by repeated short exposures also causes adaptation but by mechanisms that seem to be different. These latter results were confirmed in mice. It was also found that this type of adaptation is nonspecific because it can be produced by other stresses such as swimming or fasting. Simultaneous determinations of glycogen in the liver and soleus and tibialis muscles indicated a possible role for this substrate in cold resistance. Repeated cold exposure (8 times at -15 degrees C for 10 min), swimming for 3 h, or fasting for 48 h--all reduced the glycogen stores when measured immediately after the stress. However, the levels of glycogen were significantly increased above the initial values (P less than 0.01) when the determinations were made 24 h later. Cold tolerance measured by resistance to hypothermia at -5 degrees C was improved only when the test was done 24 h after the stress had taken place. Thus, cold resistance, as described in this study, is nonspecific and our results suggest that glycogen stores could serve as a rate-limiting substrate.

Acclimatization↗

[Cold-induced urticaria].

Cold urticaria is characterized by the development of urticaria, usually superficial and/or angioedematous reaction after cold contact. It was found predominantly in young women. The diagnosis is based on the history and ice cube test. Patients with a negative ice cube test may have represented systemic cold urticaria (atypical acquired cold urticaria) induced by general body cooling. The pathogenesis is poorly understood. Cold urticaria can be classified into acquired and familial disorders, with an autosomal dominant inheritance. Idiopathic cold urticaria is most common type but the research of a cryopathy is necessary. Therapy is often difficult. It is essential that the patient be warned of the dangers of swimming in cold water because systemic hypotension can occur. H1 antihistamines can be used for treatment of cold urticaria but the clinical responses are highly variable. The combination with an H2 antagonists is more effective. Doxepin may be useful in the treatment. Leukotriene receptor antagonists may be a novel, promising drug entity. In patients who do not respond to previous treatments, induction of cold tolerance may be tried.

Acetates↗

Cold shock response in mammalian cells.

Compared to bacteria and plants, the cold shock response has attracted little attention in mammals except in some areas such as adaptive thermogenesis, cold tolerance, storage of cells and organs, and recently, treatment of brain damage and protein production. At the cellular level, some responses of mammalian cells are similar to microorganisms; cold stress changes the lipid composition of cellular membranes, and suppresses the rate of protein synthesis and cell proliferation. Although previous studies have mostly dealt with temperatures below 20 degrees C, mild hypothermia (32 degrees C) can change the cell's response to subsequent stresses as exemplified by APG-1, a member of the HSP110 family. Furthermore, 32 degrees C induces expression of CIRP (cold-inducible RNA-binding protein), the first cold shock protein identified in mammalian cells, without recovery at 37 degrees C. Remniscent of HSP, CIRP is also expressed at 37 degrees C and developmentary regulated, possibly working as an RNA chaperone. Mammalian cells are metabolically active at 32 degrees C, and cells may survive and respond to stresses with different strategies from those at 37 degrees C. Cellular and molecular biology of mammalian cells at 32 degrees C is a new area expected to have considerable implications for medical sciences and possibly biotechnology.

Adaptation, Physiological↗

Ectopic expression of a cold-inducible transcription factor, CBF1/DREB1b, in transgenic rice (Oryza sativa L.).

The gene encoding C-repeat/dehydration-responsive element binding factor 1 (CBF1/DREB1b) of Arabidopsis was introduced into rice (Oryza sativa L.) under the control of the maize ubiquitin promoter. Its incorporation and expression in transgenic rice plants were confirmed by DNA and RNA gel-blot analyses. Cold tolerance in the transgenics was not significantly different from that of the wild-type plants, as determined by ion leakage, chlorophyll fluorescence, and survival rates. However, the cold-responsive genes lip5, lip9, and OsDhn1 were up-regulated in the transgenic plants, suggesting that the cold signal transduction pathway involving CBF1 is partially conserved in this cold-labile plant.

Arabidopsis Proteins↗

VSP accumulation and cold-inducible gene expression during autumn hardening and overwintering of alfalfa.

This study describes the time-course of the accumulation of total soluble proteins (TSPs) and vegetative storage proteins (VSPs) and of the transcripts of cold-inducible (CI) and VSP-encoding genes in taproots of two alfalfa cultivars (AC Caribou and Europe) during their acclimation to natural autumn hardening and overwintering conditions in eastern Canada. The impact of a defoliation in September on these winter hardening-related changes was also assessed. Both concentrations and pools of VSPs increased significantly between early September and mid-October and remained unchanged thereafter, concomitantly with the disappearance of VSP-encoding transcripts. Other soluble protein constituents continued to increase later in the autumn and early winter and accounted for nearly 60% of taproot TSP pools in winter. As a result, VSP abundance relative to TSPs decreased markedly during the winter. The increase in the levels of CI transcripts was induced by lowering temperatures, and distinct patterns suggest differences in the regulation of their accumulation. RNA analyses revealed that the accumulation of VSP transcripts during the autumn precedes the accumulation of CI transcripts. Autumn defoliation interrupted the accumulation of both TSPs and VSPs during autumn hardening and repressed the transcript levels of two CI genes differentially between cultivars. The well-documented impact of autumn defoliation on the vigour of spring regrowth and long-term persistence of alfalfa could be related to its negative impact on the accumulation of VSPs and TSPs and on the expression of genes encoding CI proteins potentially involved in cold tolerance and pathogen resistance.

Acclimatization↗

Some endocrine responses to sauna, shower and ice water immersion.

Nine winter swimmer men were exposed to: (A) sauna and ice water immersion; (B) sauna and 15 degrees C shower; (C) sauna and room temperature; (D) head-out ice-water immersion and room temperature. The exposures were repeated and ended with a recumbent recovery. The initial, post-exposure and post-recovery concentrations of plasma ACTH, serum cortisol, serum melatonin, plasma norepinephrine and plasma epinephrine were determined. ACTH and cortisol indicated a slightly increased post-exposure level. Melatonin concentration did not change. Post-exposure norepinephrine levels increased (P less than 0.05) from the initial. Post-exposure epinephrine indicated a tendency to elevated levels with a nearly doubled (P less than 0.05) concentration in experiment A. The tendency toward enhanced ACTH and cortisol secretion and sympathetic activity shown by increased catecholamine secretion suggest that the winter swimming practice may raise the pain threshold and develop a potential for improved cold tolerance, possibly by nonshivering thermogenesis.

Adult↗

Identification of quantitative trait loci and associated candidate genes for low-temperature tolerance in cold-hardy winter wheat.

Low-temperature (LT) tolerance is an important economic trait in winter wheat (Triticum aestivum L.) that determines the plants' ability to cope with below freezing temperatures. Essential elements of the LT tolerance mechanism are associated with the winter growth habit controlled by the vernalization loci (Vrn-1) on the group 5 chromosomes. To identify genomic regions, which in addition to vrn-1 determine the level of LT tolerance in hexaploid wheat, two doubled haploid (DH) mapping populations were produced using parents with winter growth habit (vrn-A1, vrn-B1, and vrn-D1) but showing different LT tolerance levels. A total of 107 DH lines were analyzed by genetic mapping to produce a consensus map of 2,873 cM. The LT tolerance levels for the Norstar (LT(50)=-20.7 degrees C) x Winter Manitou (LT(50)=-14.3 degrees C) mapping population ranged from -12.0 to -22.0 degrees C. Single marker analysis and interval mapping of phenotyped lines revealed a major quantitative trait locus (QTL) on chromosome 5A and a weaker QTL on chromosome 1D. The 5A QTL located 46 cM proximal to the vrn-A1 locus explained 40% of the LT tolerance variance. Two C-repeat Binding Factor (CBF) genes expressed during cold acclimation in Norstar were located at the peak of the 5A QTL.

Cold Temperature↗

Natural resistance to liver cold ischemia-reperfusion injury associated with the hibernation phenotype.

The success of liver grafts is currently limited by the length of time organs are cold preserved before transplant. Novel insights to improve viability of cold-stored organs may emerge from studies with animals that naturally experience low body temperatures (T(b)) for extended periods. In this study, we tested whether livers from hibernating ground squirrels tolerate cold ischemia-warm reperfusion (cold I/R) for longer times and with better quality than livers from rats or summer squirrels. Hibernators were used when torpid (T(b) < 10 degrees C) or aroused (T(b) = 37 degrees C). Livers were stored at 4 degrees C in University of Wisconsin solution for 0-72 h and then reperfused with 37 degrees C buffer in vitro. Lactate dehydrogenase (LDH) release after 60 min was increased 37-fold in rat livers after 72 h cold I/R but only 10-fold in summer livers and approximately three- to sixfold in torpid and aroused hibernator livers, despite twofold higher total LDH content in livers from hibernators compared with rats or summer squirrels. Reperfusion for up to 240 min had the least effect on LDH release in livers from hibernators and the greatest effect in rats. Compared with rats or summer squirrels, livers from hibernators after 72 h cold I/R showed better maintenance of mitochondrial respiration, bile production, and sinusoidal lining cell viability, as well as lower vascular resistance and Kupffer cell phagocytosis. These results demonstrate that the hibernation phenotype in ground squirrels confers superior resistance to liver cold I/R injury compared with rats and summer squirrels. Because hibernation-induced protection is not dependent on animals being in the torpid state, the mechanisms responsible for this effect may provide new strategies for liver preservation in humans.

Animals↗

Arterial abnormalities of the hand in athletes.

Vascular lesions of the hand may be seen in athletes exposed to repetitive blunt trauma. Thirteen athletes seen from 1983 to 1988 experienced symptoms related to hand ischemia. Nine were professional baseball catchers. The majority of patients complained of chronic symptoms, including cold hypersensitivity (four patients), finger numbness (one), finger coolness (three), and finger blanching (three). Two patients had acute symptoms with sudden posttraumatic hand ischemia with finger and palmar pain. Diagnosis was established by history and physical examination and confirmed by noninvasive testing. Testing included use of Doppler examination and cold tolerance examination with thermistors. Two athletes required angiographic evaluation because of severe ischemia and continuous pain. All patients in this group were managed nonoperatively. Those with chronic complaints were counseled regarding cold avoidance and instructed to increase their glove padding. The two patients with acute symptoms required vasodilator (papaverine chloride) infusion, followed by intravenous heparin and dextran. All baseball catchers returned to their sport with dissipation of symptoms.

Arteries↗

Mammalian hibernation: lessons for organ preparation?

The adaptations to low environmental temperatures exhibited in mammalian hibernation are many and varied, and involve molecular and cellular mechanisms as well as the systematic physiology of the whole organism. Natural torpidity is characterised by a profound reduction in body temperature and other functions lasting from a few hours to several weeks. Controlled reduction of heart rate, respiration and oxygen consumption is followed by the fall in body temperature. However, thermoregulation persists such that a decrease in ambient temperature below dangerous levels typically triggers arousal, and shivering and non-shivering thermogenesis from brown fat provide the heat to restore body temperature to normal levels. Many of the cellular mechanisms for survival are similar to those brought into play during medium-term storage of organs destined for transplantation. For example maintenance of ionic regulation and membrane fluxes is fundamental to cell survival and function at low body temperatures. Differences between hibernating and non-hibernating species are marked by differences in Na+/K+ transport and Ca++pumps. These in turn are probably associated with alterations in the lipoproteins of the plasma membrane and inner mitochondrial membrane. We have accordingly conducted a series of pilot studies in captured Richardson's ground squirrels kept in laboratory conditions as a model for hypothermic organ preservation. Tissue function was compared during the summer (non-hibernating season) with that in the winter when the animals could be: (i) in deep hibernation in a cold chamber at 4 degree C; (ii) maintained in an ambient temperature of 4 degree C but active and awake; or (iii) active at an ambient temperature of 22 degree C. The studies involved: whole animal monitoring of standard physiological parameters; whole organ (kidney) storage and transplantation for viability assessment; storage and functional assessment on an ex vivo test circuit with capacity for perfusion at normothermic and hypothermic temperatures; measurement of thyroid function; measurements of total nucleotides (ATP, ADP and AMP)and ratios by standard techniques after freeze-clamping of organs; similar nucleotide and pH measurements using31P-NMR as a non-invasive whole animal technique; and measurement of O2 uptake and gluconeogenesis using isolated renal tubules and isolated hepatocytes. Marked differences in cold tolerance were demonstrated between organs taken from hibernating versus non-hibernating individuals. In particular kidneys transplanted from animals in deep hibernation were capable of withstanding up to 72 hours of cold storage as compared with up to 24 hours in non-hibernating squirrels or in comparable sized rats. Adaptations which might provide valuable clues in our attempts to better preserve human organs for transplantation are explored in some depth in this report.

Journal Article↗

Human cold air habituation is independent of thyroxine and thyrotropin.

Thyroxine (T4) is required in species possessing brown adipose tissue (BAT) for the maintenance of cold tolerance and adaptation. In humans, who possess negligible quantities of BAT, the importance of T4 has not been demonstrated. We studied the effects of decreased serum T4 and thyrotropin (TSH) on human cold habituation after repeated cold air exposures. Eight men (T3+) received a single daily dose of triiodothyronine (T3; 30 micrograms/day), and another eight men (T3-) received a placebo. All 16 normal thyroid men underwent a standardized cold air test (SCAT) under basal conditions in January and again in March after eighty 30-min 4.4 degrees C air exposures (10/wk). Measurements of basal metabolic rate (BMR), O2 consumption (VO2), mean arterial pressure (MAP), plasma norepinephrine (NE), serum TSH, free and total T4, and free and total T3 were repeated before and after 8 wk of exposure. TSH, free T4, and total T4 were 50% lower for T3+ than for T3- subjects. Total and free T3 were not different between groups. BMR was unchanged after habituation, whereas the cold-stimulated VO2, MAP, and NE were significantly reduced for all subjects in March. The relationship between VO2 and NE (r2 = 0.44, P less than 0.001) during the initial SCAT was unchanged with habituation. We suggest that human cold habituation is independent of major changes in circulating T4 and TSH.

Acclimatization↗

Effects of salicylic acid and cold treatments on protein levels and on the activities of antioxidant enzymes in the apoplast of winter wheat leaves.

The effects of salicylic acid (SA) and cold on apoplastic protein levels and activities of apoplastic catalase (CAT), peroxidase (POX) and polyphenol oxidase (PPO) were investigated in winter wheat (Triticum aestivum cv. Dogu-88) leaves. The plants were grown with and without 10 microM SA treatment at both control (20/18 degrees C for 30 and 45-day) and cold (10/5 degrees C for 30-day and 5/3 degrees C for 45-day) acclimatisations. Molecular masses of the apoplastic polypeptides were shown ranging in size from 20 to 66 kDa on SDS-PAGE. Accumulation and pattern of the polypeptides were changed by both SA and cold. It is observed that CAT, POX and PPO activities at 45-day control leaves were higher than at 30-day. When the activities with SA and cold treatments are compared to their controls, CAT activities were decreased while POX and PPO activities were increased by both the treatments. When the activities with cold+SA treatment are compared to their cold treatments, CAT and POX activities were decreased while PPO activity was increased by SA. It is concluded that exogenous SA can be involved in cold tolerance by regulating apoplastic proteins and antioxidant enzyme activities.

Antioxidants↗