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Fructokinase and hexokinase from pollen grains of bell pepper (Capsicum annuum L.): possible role in pollen germination under conditions of high temperature and CO2 enrichment.

The processes of pollen grain development and germination depend on the uptake and metabolism of pollen sugars. In pepper (Capsicum annuum L.), initial sugar metabolism includes sucrose hydrolysis by invertase and subsequent phosphorylation of glucose and fructose by hexose kinases. The main objective of this study was to investigate changes in fructokinase (EC 2.7.1.4) and hexokinase (EC.2.7.1.1) activities in pepper flowers during their development, and to study the possible roles of these enzymes in determining pollen germination capacity under high temperature and under CO(2) enrichment, previously shown to modify sugar concentrations in pepper pollen (Aloni et al., 2001 Physiologia Plantarum 112: 505-512). Fructokinase (FK) activity was predominant in pepper pollen, and increased during pollen maturation. Pollen hexokinase (HK) activity was low and did not change throughout pollen development. High-temperature treatment (day/night, 32/26 degrees C) of pepper plants reduced the percentage of pollen that germinated compared with that under normal temperatures (26/22 degrees C), and concomitantly reduced the activity of FK in mature pollen. High temperature also reduced FK and HK activity in the anther. Under high ambient CO(2) (800 micro l l(-1)) pollen FK activity was enhanced. The results suggest that pollen and anther FK may play a role in the regulation of pollen germination, possibly by providing fructose-6-phosphate for glycolysis, or through conversion to UDP-glucose (UDPG) to support the biosynthesis of cell wall material for pollen tube growth. High temperature stress and CO(2) enrichment may influence pollen germination capacity by affecting these pathways.

Calcium Chloride↗

A new yeast gene, HTR1, required for growth at high temperature, is needed for recovery from mating pheromone-induced G1 arrest.

A new temperature-sensitive mutant of Saccharomyces cerevisiae was isolated. Arrested cells grown at the nonpermissive temperature were of dumb-bell shape and contained large vacuoles. A DNA fragment was cloned based on its ability to complement this temperature sensitivity. The HTR1 gene encodes a putative protein of 93 kDa without significant homology to any known proteins. The gene was mapped between ade5 and lys5 on the left arm of chromosome VII. The phenotype of the gene disruptant appeared to be strain-specific; disruption of the gene in strain W303 caused the cells to become temperature sensitive. The arrested phenotype here was similar to that of the original ts mutant and cells in G2/M phase predominated at high temperature. Another disruptant in a strain YPH background grew slowly at high temperature due to slow progression through G2/M phase, and morphologically abnormal (elongated) cells accumulated. A single-copy suppressor that alleviated the temperature-sensitive defects in both strains was identified as MCS1/SSD1. The wild-type strains W303 and YPH are known to carry defective MCS1/SSD1 alleles; hence HTR1 may function redundantly with MCS1/SSD1 to suppress the temperature-sensitive phenotypes. In addition, based on a halo bioassay, the disruptant strains appeared to be defective in recovery from, or adaptive response to G1 arrest mediated by mating pheromone, even at the permissive temperature. Thus the gene has at least two functions and is designated HTR1 (required for high temperature growth and recovery from G1 arrest induced by mating pheromone).

Amino Acid Sequence↗

Antigenicity of cell wall mannans of Candida albicans and Candida stellatoidea cultured at high temperatures in BACTEC medium.

The study of the antigenicity of pathogenic Candida albicans and Candida stellatoidea cells grown in BACTEC fungal medium (BFM) is useful for clinical analysis so as accurately to diagnose candidiasis. When C. albicans NIH A-207 was grown in BFM and fetal bovine serum-added BFM at the high temperatures of 36 and 40 degrees C, the cell density increased, with a mixture of yeast cells, pseudohyphae, and hyphae and with full hyphal development in the cultures compared with cultivation (mostly cells in yeast form) at 27 degrees C in both media. The mannans produced when cells were grown at these high temperatures were less reactive by enzyme-linked immunosorbent assay with factor sera 4, 5, and 6 in the commercially available kit 'Candida Check' than were the mannans obtained following growth at 27 degrees C. Based on 1H-nuclear magnetic resonance analysis, the mannans from cells grown at high temperatures had lost a phosphate group and a beta-1,2-linked mannopyranose unit, and had increased the number of non-reducing terminal alpha-1,3-linked mannopyranose units. We obtained similar results for mannans produced by C. albicans J-1012, C. albicans NIH B-792, C. albicans JCM 9061, C. stellatoidea ATCC 20408, and C. stellatoidea ATCC 36232 strains cultivated in BFM at 36 degrees C. These results suggest that both C. albicans and C. stellatoidea cells cultured at high temperatures, irrespective of the medium and shape of the cells, alter their antigenicity and chemical structure of cell wall mannans.

Antigens, Fungal↗

High temperature promotes auxin-mediated hypocotyl elongation in Arabidopsis.

Physiological studies with excised stem segments have implicated the plant hormone indole-3-acetic acid (IAA or auxin) in the regulation of cell elongation. Supporting evidence from intact plants has been somewhat more difficult to obtain, however. Here, we report the identification and characterization of an auxin-mediated cell elongation growth response in Arabidopsis thaliana. When grown in the light at high temperature (29 degreesC), Arabidopsis seedlings exhibit dramatic hypocotyl elongation compared with seedlings grown at 20 degreesC. This temperature-dependent growth response is sharply reduced by mutations in the auxin response or transport pathways and in seedlings containing reduced levels of free IAA. In contrast, mutants deficient in gibberellin and abscisic acid biosynthesis or in ethylene response are unaffected. Furthermore, we detect a corresponding increase in the level of free IAA in seedlings grown at high temperature, suggesting that temperature regulates auxin synthesis or catabolism to mediate this growth response. Consistent with this possibility, high temperature also stimulates other auxin-mediated processes including auxin-inducible gene expression. Based on these results, we propose that growth at high temperature promotes an increase in auxin levels resulting in increased hypocotyl elongation. These results strongly support the contention that endogenous auxin promotes cell elongation in intact plants.

Arabidopsis↗

Identification of a gene, closely linked to dnaK, which is required for high-temperature growth of Escherichia coli.

We have constructed four deletion derivatives of the cloned dnaK gene. Plasmid pDD1, in which the last 10 amino acids of the DnaK protein have been replaced by three different amino acids derived from the pBR322 vector, was as effective as plasmid pKP31, from which it was derived, in restoring the ability of a dnaK null mutant, Escherichia coli BB1553, to plate lambda phage and to grow at high temperatures. The other three mutations, involving much larger deletions of the dnaK gene, did not restore the ability to plate lambda phage or the ability to grow at high temperatures. Plasmid pKUC2, which contains the whole dnaK gene and its promoters, was capable of restoring the ability of E. coli BB1553 to plate lambda phage but, surprisingly, it did not restore the ability to grow at high temperatures, even though it was shown that the DnaK protein was efficiently expressed in these cultures. By transposon mutagenesis and sub-cloning, we have shown the presence of a second gene in plasmid pKP31 which is required for high-temperature growth of E. coli BB1553. This gene, which we call htg A, is presumably also defective in the dnaK null mutant E. coli BB1553. We have also demonstrated that the inability of E. coli K756 to grow above 43.5 degrees C is complemented by sub-clones which contain the htg A gene, but not by plasmid pKUC2.

Amino Acid Sequence↗

Reduction of Q(A) in the dark: Another cause of fluorescence F(o) increases by high temperatures in higher plants.

Increases in the chlorophyll fluorescence F(o) (dark level fluorescence) during heat treatments were studied in various higher plants. Besides the dissociation of light-harvesting chlorophyll a/b protein complexes from the reaction center complex of PS II and inactivation of PS II, dark reduction of Q(A) via plastoquinone (PQ) seemed to be related to the F(o) increase at high temperatures. In potato leaves or green tobacco cultured cells, a part of the F(o) increase was quenched by light, reflecting light-induced oxidation of Q(A) (-) which had been reduced in the dark at high temperatures. Appearance of the F(o) increase due to Q(A) reduction depended on the plant species, and the mechanisms for this are proposed. The reductants seemed to be already present and formed by very brief illumination of the leaves at high temperatures. A ndhB-less mutant of tobacco showed that complex I type NAD(P)H dehydrogenase is not involved in the heat-induced reduction of Q(A). Quite strong inhibition of the Q(A) reduction by diphenyleneiodonium suggests that a flavoenzyme is one of the electron mediator to PQ from the reductant in the stroma. Reversibility of the heat-induced Q(A) reduction suggests that an enzyme(s) involved is activated at high temperatures and mostly returns to an inactive form at room temperature (25 degrees C).

Journal Article↗

Superconducting properties of copper oxide high-temperature superconductors.

The equations for the magnon pairing theory of high-temperature copper-oxide-based superconductors are solved and used to calculate several properties, leading to results for specific heat and critical magnetic fields consistent with experimental results. In addition, the theory suggests an explanation of why there are two sets of transition temperatures (T(c) approximately 90 K and T(c) approximately 55 K) for the Y(1)Ba(2)Cu(3)O(6+x) class of superconductors. It also provides an explanation of why La(2-x)Sr(x)CuO(4) is a superconductor for only a small range of x (and suggests an experiment to independently test the theory). These results provide support for the magnon pairing theory of high-temperature superconductors. On the basis of the theory, some suggestions are made for improving these materials.

Journal Article↗

[Effects of salicylic acid on chlorophyll fluorescence and xanthophyll cycle in cucumber leaves under high temperature and strong light].

In this study, cucumber leaves were under high temperature and strong light stress after two days of spraying with 50 to approximately 400 micromol x L(-1) salicylic acid (SA) solution. The measurements showed that SA pretreatment could inhibit the decrease of intrinsic photochemical efficiency (Fv/ /Fm), quantum yield of PSII(phiPSII), maximum fluorescence value (Fm), and photochemical quenching (qP) by 16.1% to approximately 30.2%, 11.9% to approximately 33.0%, 7.2% to approximately 41.0% and 27.2% to approximately 160.8%, respectively, and promote the increase of non-photochemical quenching (NPQ) by 13.1% to approximately 62.9%, but had little effect on initial fluorescence value (Fo). SA pretreatment could also inhibit the decrease of xanthophyll cycle pool in cucumber leaves, and increase the de-epoxidation extent of xanthophyll cycle (A + Z)/(V + A + Z),which was 29.5% and 24.6% higher than control, respectively. It was suggested that SA could protect the photosynthetic apparatus of cucumber leaves under high temperature and strong light through promoting non-radiative energy dissipation.

Chlorophyll↗

Reversed-phase liquid chromatography on a microspherical carbon column at high temperature.

The chromatographic conditions for the use of a spherical carbon column at high temperatures up to 160 degrees C are described. The column heating system and the eluate cooling system are also described. Under the conditions studied, it was found that the carbon column at elevated temperature was stable and could be applicable to any compounds that are durable under these conditions. The peptide bonds at this high temperature and at this acidity were still stable enough for analysis, but the indole residues showed poor stability.

Amino Acid Sequence↗

Enzyme catalysis in organic solvents with low water content at high temperatures. The adenosinetriphosphatase of submitochondrial particles.

A system composed of toluene, phospholipids, and Triton X-100 in which the ATPase activity of bovine heart submitochondrial particles can be studied at low water concentrations and high temperatures is described. In this system, ATPase activity starts to appear at 0.5% (v/v) water and increases as the concentration of water is increased. At 3.8% water, the enzyme exhibits saturation kinetics with respect to Mg-ATP with a Km similar to that observed in an all-water system (approximately 300 microM), but the Vmax is about 100 times lower (6 nmol min-1 mg-1) than that in water. At concentrations of water between 0.5% and 2%, the enzyme catalyzes ATP hydrolysis at temperatures of up to 91 degrees C. The conditions for achieving catalysis at high temperatures are described. Even though at low water concentrations the enzyme catalyzes ATP hydrolysis at temperatures significantly higher than in totally aqueous media, the optimal temperature for hydrolysis (approximately 58 degrees C) is independent of the water content. The half-life of the enzyme at high temperatures is significantly higher at low water concentrations than in aqueous media. In the system described, the enzyme is located in a compartment distinct from that of the substrate and products of the reaction. Apparently, the enhancement of catalytic rates by water is due to a higher conformational mobility of the protein; the same factor causes a decrease in the thermostability of the enzyme.

Adenosine Triphosphatases↗

Some advantages of high temperature for the separation of pharmaceutical compounds with mass spectrometry detection.

The beneficial effects of high temperature on separation and detection of basic compounds, the detection being performed by MS via ESI, are investigated. The influence of various parameters on both separation and detection performances is studied. These parameters include the mobile phase pH, the temperature, and the type of stationary phase. Experiments are performed under gradient elution conditions. The results obtained with four different supports, silica-, zirconia-, carbon-, and polymer-based columns, are compared by means of different criteria including the elution composition, the peak asymmetry, and the S/N. High temperature liquid chromatography at high pH with volatile buffers suitable for MS detection was shown to be an interesting choice for solutes with basic sites.

Chromatography, Liquid↗

Escherichia coli mutants incapable of supporting replication of F-like plasmids at high temperature: isolation and characterization of mafA and mafB mutants.

Mutants of Escherichia coli K-12 defective in replication of F-like plasmids at a high temperature (42 degrees C) were found among threonine-independent (Thr+) revertants of a threonine-requiring F' stain after localized mutagenesis with N-methyl-N'-nitro-N-nitrosoguanidine. Transduction experiments with phage P1 permitted us to divide these mutations into two classes with respect to man location; some mutations were located between thr and ara at about 0.8 min, very close to maf-1 reported previously (Wada et al., J. Mol. Biol. 108:25-41, 1976 and the others probably were located between leu and azi at about 1.8 min. The former class of mutants designated mafA exhibited the same plasmid specificity as maf-1; replication of plasmids F and ColVB trp, but not R386 or R222, were affected at a high temperature. By contrast, the latter mutants designated mafB were defective in replication of nay of these plasmids at a high temperature. When a culture of mafA mutants carrying an F' plasmid was transferred from 30 to 42 degrees C, the plasmid replication as determined by incorporation of [3H]thymidine into covalently closed circular F DNA was markedly inhibited. Under certain conditions, the temperature shift-up caused severe growth inhibition of the mutant cells. Examination of merodiploids (mafA/FmafA+) for plasmid maintenance suggested that the two mafA mutations tested (mafA23 and mafA36) were both dominant, at least partially, over the wild-type mafA+ allele. These properties of the mafA mutants, manifested at the restrictive temperature, are similar to those previously reported for the maf-1 mutant. Taken together with other evidence it is likely that these mutations affect either the same gene (mafA) or a set of closely linked genes, playing a specific role in autonomous plasmid replication in E. coli.

Chromosome Mapping↗

Influence of sodium ion dynamics on the 23Na quadrupolar interaction in sodalite: a high-temperature 23Na MAS NMR study.

High-temperature 33Na MAS NMR experiments up to 873 K for a number of different sodalites (Na8[AlSiO4]6(NO3)2, Na8[AlSiO4]6(NO2)2, Na8[AlSiO4]6I2, Na7.9[AlSiO4]6(SCN)7.9 x 0.5H2O, Na8[AlGeO4]6(NO3)2, and Na7[AlSiO4]6(H3O2) x 4H2O) were carried out. The spectra of the first five sodalites consist of a quadrupolar MAS pattern with different quadrupolar coupling constants. The quadrupolar interaction for the thiocyanate sodalite, the nitrate aluminosilicate, and germanate sodalite decreases strongly passing a coalescence state on heating, while the quadrupolar interaction of the iodide and nitrite sample shows nearly no change. The basic hydrosodalite shows an asymmetric lineshape at room temperature and, between 350 and 370 K, a second line due to the evaporation of cage-water emerges. The linewidth increases with rising temperature. The temperature dependence of the quadrupolar interaction seems to be a function of the sodalite beta-cage expansion. Two conceivable jump mechanisms are proposed for a tetrahedral two-site jump between occupied and unoccupied tetrahedral sites.

Magnetic Resonance Spectroscopy↗

Folded conformation of an immunostimulating tetrapeptide rigin: high temperature molecular dynamics simulation study.

Employing high temperature quenched molecular dynamics (QMD) stimulations the conformational energy space of an immunostimulating tetrapeptide rigin: H-Gly341-Gln-Pro-Arg344-OH, is explored. Using distance dependent dielectric (epsilon =r(ij)) 31 different low energy starting structures with identical sequence were computed for their conformational preferences. According to the hypothesis of O'Connors et al. [J. Med. Chem. 35 (1992), 2870], 83 low-energy conformers resulted from unrestrained molecular dynamics (MD) simulations, could be classified into two energy minimized families: A and B, comprised of 64 (Pro C(gamma)-endo orientation) and 19 (Pro C(gamma)-exo orientation) structures, respectively. An examination of these families revealed the existence of a remarkably similar folded backbone conformation: torsion angles being phi(i+1) approximately -65 degrees, psi(i+1) approximately -65 degrees, phi(i+2) approximately -65 degrees, psi(i+2) approximately -60 degrees, characterizing a distorted type III beta-turn structure across the central Gln-Pro segment. The folded conformation of rigin is devoid of a classical 1 <-- 4 intra-molecular hydrogen bond nevertheless, the conformation is stabilized by an effective 'salt-bridge', i.e., Gly H(3)N(+)...C(alpha)OO(-) Arg interaction. Surprisingly, in both the families the unusual folded side-chain dispositions of the Gln residue favor the formation of a unique intra-residue 'main-chain to side-chain' H-bond, i.e., N(alpha)-H...N(epsilon) interaction, encompassing a seven-membered ring motif. The conformational attributes may be valuable in de novo construction of structure-based drug candidates having sufficient stimulating activity.

Adjuvants, Immunologic↗

Spatially resolved electronic structure inside and outside the vortex cores of a high-temperature superconductor.

Puzzling aspects of high-transition-temperature (high-Tc) superconductors include the prevalence of magnetism in the normal state and the persistence of superconductivity in high magnetic fields. Superconductivity and magnetism generally are thought to be incompatible, based on what is known about conventional superconductors. Recent results, however, indicate that antiferromagnetism can appear in the superconducting state of a high-Tc superconductor in the presence of an applied magnetic field. Magnetic fields penetrate a superconductor in the form of quantized flux lines, each of which represents a vortex of supercurrents. Superconductivity is suppressed in the core of the vortex and it has been suggested that antiferromagnetism might develop there. Here we report the results of a high-field nuclear-magnetic-resonance (NMR) imaging experiment in which we spatially resolve the electronic structure of near-optimally doped YBa2Cu3O7-delta inside and outside vortex cores. Outside the cores, we find strong antiferromagnetic fluctuations, whereas inside we detect electronic states that are rather different from those found in conventional superconductors.

Journal Article↗

Detection of 16S rDNA sequences representing the novel phylum "Nanoarchaeota": indication for a wide distribution in high temperature biotopes.

We screened samples from high temperature biotopes for 16S rRNA genes of the novel archaeal phylum "Nanoarchaeota". Positive PCR amplifications were obtained from Yellowstone National Park, Uzon Caldera, and an abyssal vent system. These sequences form a cluster with the sequence of "Nanoarchaeum equitans", indicating a wide distribution of this phylum.

Archaea↗