bluuebunny/crossref_metadata_2025 huggingface dataset:
The population genomics of rhesus macaques (Macaca mulatta) based on whole-genome sequences
Authors: Xue Cheng , Raveendran Muthuswamy , Harris R. Alan , Fawcett Gloria L. , Liu Xiaoming , White Simon , Dahdouli Mahmoud , Rio Deiros David , Below Jennifer E. , Salerno William , Cox Laura , Fan Guoping , Ferguson Betsy , Horvath Julie , Johnson Zach , Kanthaswamy Sree , Kubisch H. Michael , Liu Dahai , Platt Michael , Smith David G. , Sun Binghua , Vallender Eric J. , Wang Feng , Wiseman Roger W. , Chen Rui , Muzny Donna M. , Gibbs Richard A. , Yu Fuli , Rogers Jeffrey
Published: October 2016

Rhesus macaques (Macaca mulatta) are the most widely used nonhuman primate in biomedical research, have the largest natural geographic distribution of any nonhuman primate, and have been the focus of much evolutionary and behavioral investigation. Consequently, rhesus macaques are one of the most thoroughly studied nonhuman primate species. However, little is known about genome-wide genetic variation in this species. A detailed understanding of extant genomic variation among rhesus macaques has implications for the use of this species as a model for studies of human health and disease, as well as for evolutionary population genomics. Whole-genome sequencing analysis of 133 rhesus macaques revealed more than 43.7 million single-nucleotide variants, including thousands predicted to alter protein sequences, transcript splicing, and transcription factor binding sites. Rhesus macaques exhibit 2.5-fold higher overall nucleotide diversity and slightly elevated putative functional variation compared with humans. This functional variation in macaques provides opportunities for analyses of coding and noncoding variation, and its cellular consequences. Despite modestly higher levels of nonsynonymous variation in the macaques, the estimated distribution of fitness effects and the ratio of nonsynonymous to synonymous variants suggest that purifying selection has had stronger effects in rhesus macaques than in humans. Demographic reconstructions indicate this species has experienced a consistently large but fluctuating population size. Overall, the results presented here provide new insights into the population genomics of nonhuman primates and expand genomic information directly relevant to primate models of human disease.
How does intentionality of encoding affect memory for episodic information?
Authors: Craig Michael , Butterworth Karla , Nilsson Jonna , Hamilton Colin J. , Gallagher Peter , Smulders Tom V.
Published: October 2016

Episodic memory enables the detailed and vivid recall of past events, including target and wider contextual information. In this paper, we investigated whether/how encoding intentionality affects the retention of target and contextual episodic information from a novel experience. Healthy adults performed (1) a What-Where-When (WWW) episodic memory task involving the hiding and delayed recall of a number of items (what) in different locations (where) in temporally distinct sessions (when) and (2) unexpected tests probing memory for wider contextual information from the WWW task. Critically, some participants were informed that memory for WWW information would be subsequently probed (intentional group), while this came as a surprise for others (incidental group). The probing of contextual information came as a surprise for all participants. Participants also performed several measures of episodic and nonepisodic cognition from which common episodic and nonepisodic factors were extracted. Memory for target (WWW) and contextual information was superior in the intentional group compared with the incidental group. Memory for target and contextual information was unrelated to factors of nonepisodic cognition, irrespective of encoding intentionality. In addition, memory for target information was unrelated to factors of episodic cognition. However, memory for wider contextual information was related to some factors of episodic cognition, and these relationships differed between the intentional and incidental groups. Our results lead us to propose the hypothesis that intentional encoding of episodic information increases the coherence of the representation of the context in which the episode took place. This hypothesis remains to be tested.
Measuring discrimination- and reversal learning in mouse models within 4 days and without prior food deprivation
Authors: Remmelink Esther , Smit August B. , Verhage Matthijs , Loos Maarten
Published: October 2016

Many neurological and psychiatric disorders are characterized by deficits in cognitive flexibility. Modeling cognitive flexibility in mice enables the investigation of mechanisms underlying these deficits. The majority of currently available behavioral tests targeting this cognitive domain are reversal learning tasks that require scheduled food restriction, extended training periods and labor-intensive, and stress-inducing animal handling. Here, we describe a novel 4-day (4-d) continuously running task measuring discrimination- and reversal learning in an automated home cage (CognitionWall DL/RL task) that largely eliminates these limitations. In this task, mice can earn unlimited number of food rewards by passing through the correct hole of the three-holed CognitionWall. To assess the validity and sensitivity of this novel task, the performance of C57BL/6J mice, amyloid precursor protein/presenilin1 transgenic (APP/PS1) mice, α-calmodulin kinase-II (αCaMKII) T305D knock-in mice, and mice with an orbitofrontal cortex lesion were examined. We found that C57BL/6J mice reach stable performance levels within the 4 d of the task, while experiencing only slight reductions in weight and no major effects on circadian rhythm. The task detected learning deficits in APP/PS1 transgenic and αCaMKII T305D mutant mice. Additionally, we established that the orbitofrontal cortex underlies reversal learning performance in our task. Because of its short duration and the absence of food deprivation and concurrent weight loss, this novel automated home-cage task substantially improves comprehensive preclinical assessment of cognitive functions in mouse models of psychiatric and neurological disorders and also enables analysis during specific developmental stages.
Intracellular Localization of Phospholipase D in Leaves and Seedling Tissues of Castor Bean
Authors: Xu L. , Paulsen A. Q. , Ryu S. B. , Wang X.
Published: October 2016

The intracellular distribution of phospholipase D (PLD; EC 3.1.4.4) in castor bean (Ricinus communis L.) tissues was investigated by subcellular fractionation and by immuno-electron microscopy. Centrifugal fractionation revealed that most PLD in young leaves was soluble, whereas in mature leaves a majority of PLD was associated with microsomal membranes. Further separation of microsomal membranes by a two-phase partitioning system indicated that PLD was associated with both plasma and intracellular membranes. Sucrose gradient separation of intracellular membranes showed PLD present in the endoplasmic reticulum, a submicrosomal band, and in soluble fractions but not in mitochondria and glyoxysomes of postgermination endosperm. Immunocytochemical studies found high gold labeling in vacuoles in young leaves, suggesting that the high level of soluble PLD in young leaves is due to release of PLD from vacuoles during tissue disruption. In addition to the labeling in vacuoles, gold particles were also found in the cytoplasmic matrices and plasma membrane in leaves and in 2-d postgermination seedlings. Collectively, these results show that PLD in castor bean leaf and seedling tissues is localized in the vacuole and is associated with the endoplasmic reticulum and plasma membrane and that the relative distribution between the soluble and membrane compartments changes during castor bean leaf development.
Induction of Male Sterility in Wheat by Meiotic-Stage Water Deficit Is Preceded by a Decline in Invertase Activity and Changes in Carbohydrate Metabolism in Anthers
Authors: Dorion S. , Lalonde S. , Saini H. S.
Published: October 2016

Water deficit during meiosis in pollen mother cells of wheat (Triticum aestivum L.) induces male sterility, which can reduce grain set by 40 to 50%. In plants stressed during meiosis and then rewatered, division of pollen mother cells proceeds normally but subsequent pollen development is arrested 3 or 4 d later. An inhibition of starch accumulation within the pollen grain suggested that an alteration in carbohydrate metabolism or assimilate supply may be involved in pollen abortion. We measured levels of various carbohydrates and activities of key enzymes of Suc metabolism and starch synthesis at different stages of pollen development in anthers collected from well-watered and water-stressed plants. Compared to controls, soluble sugars increased in anthers stressed during meiosis, then decreased at later poststress stages. Sucrose and myoinositol accounted for part of the sugar accumulation. The activity of soluble acid invertase declined 4-fold during the stress period and never recovered thereafter. Sucrose synthase activity during starch accumulation in pollen was also lower in the anthers of plants stressed at meiosis. Stress had little negative effect on the activities of ADP-glucose pyrophosphorylase or soluble and granule-bound starch synthase during starch accumulation in pollen, although at the earlier stages, ADP-glucose pyrophosphorylase activity in stressed anthers was slightly lower compared to controls. The results suggest that carbohydrate starvation per se and inhibition of the enzymes of starch synthesis probably were not responsible for the stress-induced pollen abortion. Instead, an inability to metabolize incoming sucrose to hexoses may be involved in this developmental lesion.
Little or No Repair of Cyclobutyl Pyrimidine Dimers Is Observed in the Organellar Genomes of the Young Arabidopsis Seedling
Authors: Chen J. J. , Jiang C. Z. , Britt A. B.
Published: October 2016

A Southern-blot-based, site-specific assay for ultraviolet (UV)-induced cyclobutyl pyrimidine dimers (CPDs), employing the CPD-specific enzyme T4 endonuclease V, was used to follow the repair of this lesion in particular DNA sequences in 5- to 6-d-old Arabidopsis thaliana seedlings. CPDs, measured as enzyme-sensitive sites, in nuclear sequences were removed rapidly in the light but were repaired slowly, if at all, in the dark. This result was identical to that obtained in prior analyses of CPDs in total cellular DNA. Assay of representative chloroplast and mitochondrial sequences in the same DNA preparations revealed that, in contrast to nuclear sequences, enzyme-sensitive sites are inefficiently eliminated in both the presence and absence of visible light. These observations suggest that Arabidopsis seedlings possess little or no capacity for the repair of CPDs in the organellar genomes. Given the fact that the UV dose employed only marginally affected the growth of the seedlings, we suggest that Arabidopsis seedlings must possess very efficient mechanism(s) for the tolerance of UV-induced DNA damage.
Attenuation of the Phenotype Caused by the Root-Inducing, Left-Hand, Transferred DNA and Its rolA Gene (Correlations with Changes in Polyamine Metabolism and DNA Methylation)
Authors: Martin-Tanguy J. , Sun L. Y. , Burtin D. , Vernoy R. , Rossin N. , Tepfer D.
Published: October 2016

We present four examples of attenuation of the transformed phenotype caused by the root-inducing, left-hand, transferred DNA from Agrobacterium rhizogenes in tobacco (Nicotiana tabacum). The first was associated with a genetic variable (homozygosity for the T-DNA), and the second was induced at the physiological level by putrescine and tyramine, suggesting that the transformed phenotype depends on defective polyamine metabolism. Physiological attenuation is further illustrated in the third example, in which the inhibition of flowering caused by P35S-rolA, a gene from the root-inducing, left-hand, transferred DNA driven by a strong viral promoter, was attenuated by grafting the transformed shoot onto non-transformed rootstock that had been induced to flower. Infertility in the resulting flowers was corrected by a mixture of putrescine and tyramine, indicating that P35S-rolA inhibited flowering through interference with polyamine conjugation and that tyramine was essential to fertility. A fourth example of attenuation of the transformed phenotype occurred in lateral branches of plants expressing rolA under the control of its native promoter. In these branches, reduction in the accumulation of rolA transcripts was correlated with the methylation of a site 3[prime] to the rolA coding sequence; thus, the transformed plant seems capable of recognizing and repressing a gene that interferes with flowering.
Sucrolytic Enzyme Activities in Cotyledons of the Faba Bean (Developmental Changes and Purification of Alkaline Invertase)
Authors: Ross H. A. , McRae D. , Davies H. V.
Published: October 2016

In terms of maximum extractable catalytic activity, sucrose synthase is the predominant sucrolytic enzyme in developing cotyledons of faba bean (Vicia faba L.). Although acid invertase activity is extremely low, there is significant activity of alkaline invertase, the majority of which is extractable only with high concentrations of NaCl. Calculations of potential activity in vivo indicate that alkaline invertase is the predominant sucrolytic enzyme from 50 days after anthesis onward. However, at almost all stages of cotyledon development analyzed, the maximum extractable catalytic activities of both enzymes is in excess of the actual rate of starch deposition. Two forms of alkaline invertase were identified in developing cotyledons. The major form has been purified to homogeneity, and antibodies have been raised against it. The native protein has a molecular mass of about 238 [plus or minus] 4.5 kD. It is apparently a homotetramer (subunit molecular mass 53.4 [plus or minus] 0.9 kD). The enzyme has a pH optimum of 7.4, an isoelectric point of 5.2, and a Km[sucrose] of 10 mM and is inhibited by Tris (50% inhibition at 5 mM) and fructose (30% inhibition at 10 mM). Bean alkaline invertase is a β-fructofuranosidase with no significant activity against raffinose, stachyose, trehalose, maltose, or lactose.
Mastoparan-Induced Intracellular Ca2+ Fluxes May Regulate Cell-to-Cell Communication in Plants
Authors: Tucker E. B. , Boss W. F.
Published: October 2016

The relationship of Ca2+ and plasmodesmatal closure was examined in staminal hairs of Setcreasea purpurea by microinjecting cells with active mastoparan (Mas-7), inactive mastoparan (Mas-17), active inositol-1,4,5-trisphosphate (IP3), or inactive IP3. Calcium green dextran 10,000 was used to study cellular free Ca2+, and carboxyfluorescein was used to monitor plasmodesmatal closure. When Mas-7 was microinjected into the cytoplasm of cell 1 (the tip cell of a chain of cells), a rapid increase in calcium green dextran-10,000 fluorescence was observed in the cytoplasmic areas on both sides of the plasmodesmata connecting cells 1 and 2 during the same time that the diffusion of carboxyfluorescein through them was blocked. The inhibition of cell-to-cell diffusion was transient, and the closed plasmodesmata reopened within 30 s. The elevated Ca2+ level near plasmodesmata was also transient and returned to base level in about 1.5 min. The transient increase in Ca2+, once initiated in cell 1, repeated with an oscillatory period of 3 min. Elevated Ca2+ and oscillations of Ca2+ were also observed near interconnecting cell walls throughout the chain of cells, indicating that the signal had been transmitted. Previously, we reported that IP3 closed plasmodesmata; now we report that it stimulated Ca2+ and oscillations similar to Mas-7. The effect was specific for similar concentrations of Mas-7 over Mas-17 and active IP3 over inactive IP3. It is important that the Ca2+ channel blocker La3+ eliminated the responses from Mas-7 and IP3, indicating that an influx of Ca2+ was required. These results support the contention that plasmodesmata functioning is regulated via Ca2+ and that IP3 may be an intermediary between the stimulus and Ca2+ elevations.
Solid-State 13C Nuclear Magnetic Resonance Characterization of Cellulose in the Cell Walls of Arabidopsis thaliana Leaves
Authors: Newman R. H. , Davies L. M. , Harris P. J.
Published: October 2016

Solid-state 13C nuclear magnetic resonance was used to characterize the molecular ordering of cellulose in a cell-wall preparation containing mostly primary walls obtained from the leaves of Arabidopsis thaliana. Proton and 13C spin relaxation time constants showed that the cellulose was in a crystalline rather than a paracrystalline state or amorphous state. Cellulose chains were distributed between the interiors (40%) and surfaces (60%) of crystallites, which is consistent with crystallite cross-sectional dimensions of about 3 nm. Digital resolution enhancement revealed signals indicative of triclinic and monoclinic crystalline forms of cellulose mixed in similar proportions. Of the five nuclear spin relaxation processes used, proton rotating-frame relaxation provided the clearest distinction between cellulose and other cell-wall components for purposes of editing solid-state 13C nuclear magnetic resonance spectra.
NaCl-Induced Alterations in Both Cell Structure and Tissue-Specific Plasma Membrane H+ -ATPase Gene Expression
Authors: Niu X. , Damsz B. , Kononowicz A. K. , Bressan R. A. , Hasegawa P. M.
Published: October 2016

NaCl-induced plasma membrane H+-ATPase gene expression, which occurs in roots and fully expanded leaves of the halophyte Atriplex nummularia L. (X. Niu, M.L. Narasimhan, R.A. Salzman, R.A. Bressan, P.M. Hasegawa [1993] Plant Physiol 103: 713–718), has been differentially localized to specific tissues using in situ RNA hybridization techniques. Twenty-four-hour exposure of plants to 400 mM NaCl resulted in substantial accumulation of H+ pump message in the epidermis of the root tip and the endodermis of the root elongation/differentiation zone. In expanded leaves, NaCl induction of plasma membrane H+-ATPase message accumulation was localized to bundle-sheath cells. Ultrastructural analyses indicated that significant cytological adaptations in root cells included plasmolysis that is accompanied by plasma membrane invaginations, formation of Hechtian strands and vesiculation, and vacuolation. These results identify specific tissues that are involved in the regulation of Na+ and Cl- uptake into different organs of the halophyte A. nummularia and provide evidence of the intercellular and interorgan coordination that occurs in the mediation of NaCl adaptation.
Germin Gene Expression Is Induced in Wheat Leaves by Powdery Mildew Infection
Authors: Hurkman W. J. , Tanaka C. K.
Published: October 2016

Germin gene expression is induced in wheat (Triticum aestivum L.) leaves by powdery mildew (Erysiphe graminis f. sp. tritici) infection. Germin is a protein marker for early cereal development and is an oxalate oxidase, an enzyme that catalyzes the conversion of oxalate to CO2 and H2O2. The induction of germin gene expression by powdery mildew infection is consistent with the importance of H2O2 to plant defense and identifies a mechanism for the elevation of H2O2 levels in wheat leaves. Germin mRNA levels increased 2 d after inoculation of seedlings with powdery mildew and continued to increase throughout an 8-d time course. The increase in accumulation of germin mRNA was accompanied by an increase in the germin oligomer, which reached maximal levels by d 6. An increase in oxalate oxidase activity paralleled germin oligomer accumulation. Germin gene expression was induced in a relatively resistant cultivar (Bobwhite) as well as in a susceptible cultivar (Cheyenne), suggesting that the induction of germin gene expression is an indicator of powdery mildew infection rather than cultivar resistance.
Influence of Salicylic Acid on the Induction of Competence for H2O2 Elicitation (Comparison of Ergosterol with Other Elicitors)
Authors: Kauss H. , Jeblick W.
Published: October 2016

Hypocotyls from etiolated cucumber (Cucumis sativus L.) seedlings were gently abraded at their epidermal surface, and cut segments were used to study the rapid and transient elicitation of H2O2 by ergosterol, chitosan, mastoparan, and a polymeric fungal elicitor. Freshly abraded segments were only barely competent for any H2O2 production, but they developed this competence subsequent to abrasion. This process was enhanced by 2,6-dichloroisonicotinic acid and salicylic acid, which induced acquired resistance to fungal penetration in the epidermal cells. Enhancement of competence induction by salicylic acid was also evident for spontaneous H2O2 production and differed in degree for the various elicitors, indicating that mainly the enzyme complex producing H2O2, but also other components of the elicitation system, improved. Ergosterol, chitosan, and fungal elicitor also rendered the segments refractory to a second stimulation by the same compound, whereas mastoparan was inactive in this respect. The four elicitors also differed markedly in their ability to diminish or enhance H2O2 production by a second treatment with a different elicitor, indicating that several sites of the H2O2 elicitation system are subject to short-term regulation.
Growth Maintenance of the Maize Primary Root at Low Water Potentials Involves Increases in Cell-Wall Extension Properties, Expansin Activity, and Wall Susceptibility to Expansins
Authors: Wu Y. , Sharp R. E. , Durachko D. M. , Cosgrove D. J.
Published: October 2016

Previous work on the growth biophysics of maize (Zea mays L.) primary roots suggested that cell walls in the apical 5 mm of the elongation zone increased their yielding ability as an adaptive response to low turgor and water potential ([psi]W). To test this hypothesis more directly, we measured the acid-induced extension of isolated walls from roots grown at high (-0.03 MPa) or low (-1.6 MPa) [psi]W using an extensometer. Acid-induced extension was greatly increased in the apical 5 mm and was largely eliminated in the 5- to 10-mm region of roots grown at low [psi]W. This pattern is consistent with the maintenance of elongation toward the apex and the shortening of the elongation zone in these roots. Wall proteins extracted from the elongation zone possessed expansin activity, which increased substantially in roots grown at low [psi]W. Western blots likewise indicated higher expansin abundance in the roots at low [psi]W. Additionally, the susceptibility of walls to expansin action was higher in the apical 5 mm of roots at low [psi]W than in roots at high [psi]W. The basal region of the elongation zone (5–10 mm) did not extend in response to expansins, indicating that loss of susceptibility to expansins was associated with growth cessation in this region. Our results indicate that both the increase in expansin activity and the increase in cell-wall susceptibility to expansins play a role in enhancing cell-wall yielding and, therefore, in maintaining elongation in the apical region of maize primary roots at low [psi]W.
Cercospora beticola Toxins (X. Inhibition of Plasma Membrane H+-ATPase by Beticolin-1)
Authors: Simon-Plas F. , Gomes E. , Milat M. L. , Pugin A. , Blein J. P.
Published: October 2016

Beticolin-1 is a toxin produced by the fungus Cercospora beticola. The chemical structure of this toxin was previously elucidated. The effects of beticolin-1 on purified corn root plasma membrane H+-ATPase were studied in a solubilized form or were reconstituted into liposome membranes. The ATP hydrolysis activity of the purified solubilized enzyme was inhibited by micromolar concentrations of beticolin-1, and this inhibition was noncompetitive with respect to ATP. When this purified enzyme was inserted into liposome membranes, a competitive inhibition of the H+-ATPase hydrolysis activity by beticolin-1 was observed. The effect of beticolin-1 on the formation of H+-ATPase-phosphorylated intermediate was also studied. With the purified enzyme in its solubilized form, the level of phosphorylated intermediate was not affected by the presence of beticolin-1, whereas micromolar concentrations of the toxin led to a marked inhibition of its formation when the enzyme was reconstituted into liposomes. These data suggest that (a) the plasma membrane H+-ATPase is a direct target for beticolin-1, and (b) the kinetics of inhibition and the effect on the phosphorylated intermediate are linked and both depend on the lipid environment of the enzyme.
Intracellular Levels of Free Linolenic and Linoleic Acids Increase in Tomato Leaves in Response to Wounding
Authors: Conconi A. , Miquel M. , Browse J. A. , Ryan C. A.
Published: October 2016

An intracellular signaling pathway for activating plant defense genes against attacking herbivores and pathogens is mediated by a lipid-based signal transduction cascade. In this pathway, linolenic acid (18:3) is proposed to be liberated from cell membranes and is converted to cyclopentanones that are involved in transcriptional regulation of defense genes, analogously to prostaglandin synthesis and function in animals. Levels of 18:3 and linoleic acid in tomato (Lycopersicon esculentum) leaves increased within 1 h when the leaves were wounded with a hemostat across the main vein to simulate herbivore attacks. The increase correlated with the time course of accumulation of jasmonic acid, a cyclopentanone product of 18:3, that had previously been shown to increase in leaves in response both to wounding and to elicitors of plant defense genes. One hour after wounding, at least a 15-fold excess of 18:3 was found over that required to account for the levels of newly synthesized jasmonic acid. The free fatty acids in both control and wounded leaves accounted for less than 0.25% of the total fatty acids. However, the total lipid contents of the leaves remained relatively unchanged up to 8 h after wounding, indicating that extensive loss of lipids did not occur, although a gradual decrease in polar lipids was observed, mainly in monogalactosyl diacylglycerol of chloroplast lipids. The data support a role for lipid release as a key step in the signaling events that activate defense genes in tomato leaves in response to wounding by attacking herbivores.
Potamogeton pectinatus Is Constitutively Incapable of Synthesizing Ethylene and Lacks 1-Aminocyclopropane-1-Carboxylic Acid Oxidase
Authors: Summers J. E. , Voesenek LACJ. , Blom CWPM. , Lewis M. J. , Jackson M. B.
Published: October 2016

A highly sensitive laser-driven photoacoustic detector responsive to [less than or equal to]2.1 nmol m-3 ethylene (50 parts per trillion [v/v]) was used for ethylene analysis. Dark-grown plants of Potamogeton pectinatus L. growing from small tubers made no ethylene. Exposure of shoots to white light, wounding, submergence in water followed by desubmergence, partial oxygen shortage, indole acetic acid, or carbon dioxide failed to induce ethylene production, although clear effects were observed in Pisum sativum L. Some ethylene was released after applying high concentrations of the ethylene precursor 1-aminocyclopropane-1-carboxylic acid (ACC; 10 mol m-3) to P. pectinatus, but the amount was trivial compared with that released by P. sativum. More endogenous ACC was found in P. pectinatus than in P. sativum. Considerable ACC oxidase activity was present in tissue extracts of P. sativum. However, no ACC oxidase activity was found in P. pectinatus, indicating that this is where ethylene production is arrested.
The Effect of Elevated [CO2] on Growth and Photosynthesis of Two Eucalyptus Species Exposed to High Temperatures and Water Deficits
Authors: Roden J. S. , Ball M. C.
Published: October 2016

Two species of eucalyptus (Eucalyptus macrorhyncha and Eucalyptus rossii) were grown for 8 weeks in either ambient (350 [mu]L L-1) or elevated (700 [mu]L L-1) CO2 concentrations, either well watered or without water additions, and subjected to a daily, 3-h high-temperature (45[deg]C, maximum) and high-light (1250 [mu]mol photons m-2 s-1, maximum) stress period. Water-stressed seedlings of E. macrorhyncha had higher leaf water potentials when grown in elevated [CO2]. Growth analysis indicated that increased [CO2] may allow eucalyptus species to perform better during conditions of low soil moisture. A down-regulation of photosynthetic capacity was observed for seedlings grown in elevated [CO2] when well watered but not when water stressed. Well-watered seedlings grown in elevated [CO2] had lower quantum efficiencies as measured by chlorophyll fluorescence (the ratio of variable to maximal chlorophyll fluorescence [Fv/Fm]) than seedlings grown in ambient [CO2] during the high-temperature stress period. However, no significant differences in Fv/Fm were observed between CO2 treatments when water was withheld. The reductions in dark-adapted Fv/Fm for plants grown in elevated [CO2] were not well correlated with increased xanthophyll cycle photoprotection. However, reductions in the Fv/Fm were correlated with increased levels of nonstructural carbohydrates. The reduction in quantum efficiencies for plants grown in elevated [CO2] is discussed in the context of feedback inhibition of electron transport associated with starch accumulation and variation in sink strength.
Synthesis and Turnover of Cell-Wall Polysaccharides and Starch in Photosynthetic Soybean Suspension Cultures
Authors: Lozovaya V. V. , Zabotina O. A. , Widholm J. M.
Published: October 2016

Soybean (Glycine max [L.] Merr.) suspension cultures grown under photoautotrophic and photomixotrophic (1% sucrose) culture conditions were used in 14CO2 pulse-chase experiments to follow cell-wall polysaccharide and starch biosynthesis and turnover. Following a 30-min pulse with 14CO2, about one-fourth of the 14C of the photoautotrophic cells was incorporated into the cell wall; this increased to about 80% during a 96-h chase in unlabeled CO2. Cells early in the cell culture cycle (3 d) incorporated more 14C per sample and also exhibited greater turnover of the pectin and hemicellulose fractions as shown by loss of 14C during the 96-h chase than did 10- and 16-d cells. When the chase occurred in the dark, less 14C was incorporated into the cell wall because of the cessation of growth and higher respiratory loss. The dark effect was much less pronounced with the photomixotrophic cells. Even though the cell starch levels were much lower than in leaves, high 14C incorporation was found during the pulse, especially in older cells. The label was largely lost during the chase, indicating that starch is involved in the short-term storage of photosynthate. Thus, these easily labeled and manipulated photosynthetic cells demonstrated extensive turnover of the cell-wall pectin and hemicellulose fractions and starch during the normal growth process.
The Chlorophyll Biosynthetic Enzyme Mg-Protoporphyrin IX Monomethyl Ester (Oxidative) Cyclase (Characterization and Partial Purification from Chlamydomonas reinhardtii and Synechocystis sp. PCC 6803)
Authors: Bollivar D. W. , Beale S. I.
Published: October 2016

A universal structural feature of chlorophyll molecules is the isocyclic ring. This ring is formed by the action of the enzyme Mg-protoporphyrin IX monomethyl ester (oxidative) cyclase, which catalyzes a complex reaction in which Mg-protoporphyrin IX monomethyl ester is converted to divinyl protochlorophyllide (also called Mg-2,4-divinylpheoporphyrin a5), with the participation of NADPH and O2. Cyclase activity was demonstrated in lysed Chlamydomonas reinhardtii chloroplasts and extracts of Synechocystis sp. PCC 6803. The yield of the reaction product was increased by the addition of catalase and ascorbate or isoascorbate to the incubation mixture. These compounds may act by preventing degradation of the tetrapyrroles by reactive oxygen species. Cyclase activity from C. reinhardtii was not inhibited by the flavoprotein inhibitor quinacrine or by the hemoprotein inhibitors CO, KCN, or NaN3. In contrast, cyclase activity in extracts of C. reinhardtii and Synechocystis sp. PCC 6803 was inhibited by chelators of Fe, suggesting that nonheme Fe is involved in the reaction. Cyclase in lysed C. reinhardtii chloroplasts was associated with the membranes, and attempts to further fractionate or solubilize the activity were unsuccessful. In contrast, cyclase in Synechocystis sp. PCC 6803 extracts could be separated into soluble and membrane components, both of which were required for reconstitution of activity. The membrane component retained activity after it was solubilized by the detergent n-octyl-β-D-glucopyranoside in the presence of glycerol and Mg2+. The solubilized membrane component was purified further by dye-affinity and ion-exchange chromatography.
NAD(P)H:(Quinone-Acceptor) Oxidoreductase of Tobacco Leaves Is a Flavin Mononucleotide-Containing Flavoenzyme
Authors: Sparla F. , Tedeschi G. , Trost P.
Published: October 2016

The soluble NAD(P)H:(quinone-acceptor) oxidoreductase [NAD(P)H-QR, EC 1.6.99.2] of Nicotiana tabacum L. leaves and roots has been purified. NAD(P)H-QR contains noncovalently bound flavin mononucleotide. Pairs of subunits of 21.4 kD are linked together by disulfide bridges, but the active enzyme is a homotetramer of 94 to 100 kD showing an isoelectric point of 5.1. NAD(P)H-QR is a B-stereospecific dehydrogenase. NADH and NADPH are electron donors of similar efficiency with Kcat:Km ratios (with duroquinone) of 6.2 x 107 and 8.0 x 107 m-1 s-1, respectively. Hydrophilic quinones are good electron acceptors, although ferricyanide and dichlorophenolindophenol are also reduced. The quinones are converted to hydroquinones by an obligatory two-electron transfer. No spectral evidence for a flavin semiquinone was detected following anaerobic photoreduction. Cibacron blue and 7-iodo-acridone-4-carboxylic acid are inhibitory. Tobacco NAD(P)H-QR resembles animal DT-diaphorase in some respects (identical reaction mechanism with a two-electron transfer to quinones, unusually high catalytic capability, and donor and acceptor substrate specificity), but it differs from DT-diaphorase in molecular structure, flavin cofactor, stereospecificity, and sensitivity to inhibitors. As in the case with DT-diaphorase in animals, the main NAD(P)H-QR function in plant cells may be the reduction of quinones to quinols, which prevents the production of semiquinones and oxygen radicals. The enzyme appears to belong to a widespread group of plant and fungal flavoproteins found in different cell compartments that are able to reduce quinones.
Regulation of De-N-Glycosylation Enzymes in Germinating Radish Seeds
Authors: Berger S. , Menudier A. , Julien R. , Karamanos Y.
Published: October 2016

The activities of the de-N-glycosylation enzymes endo-N-acetyl- β-D-glucosaminidase (ENGase; EC 3.2.1.96) and peptide-N4- (N-acetyl-β-D-glucosaminyl) asparagine amidase (PNGase; EC 3.5.1.52) were monitored during germination and postgerminative development in radish (Raphanus sativus L. cv Flamboyant). The ENGase activity was detected only during postgermination, whereas the PNGase activity was present at high levels in both stages. When germination was inhibited with abscisic acid or cycloheximide, PNGase activity was detected at a basic level and ENGase activity was not detected at all. PNGase is present as an active protein in dry seeds and is apparently synthesized during seed formation. Conversely, the absence of ENGase in dry seeds suggests that its activity is dependent on the protein synthesis that occurs during and after germination. Treatment with gibberellic acid confirmed the production of both de-N-glycosylation enzymes after germination, and demonstrated a temporal delay between the production of the two enzymes during this period. Our results suggest that the two de-N-glycosylation enzymes are differentially regulated during plant development.
Dissection of the Blue-Light-Dependent Signal-Transduction Pathway Involved in Gametic Differentiation of Chlamydomonas reinhardtii
Authors: Pan J. M. , Haring M. A. , Beck C. F.
Published: October 2016

Gametogenesis of the green alga Chlamydomonas reinhardtii may be viewed as a two-step process that is controlled by the environmental cues of nitrogen deprivation and blue light. Initiation of gametogenesis is induced by nitrogen deprivation, resulting in mating-incompetent pregametes, when cells are kept in the dark. For the completion of gametic differentiation light is required. Pregametes were treated with pharmacological compounds to influence the light-dependent conversion to mature gametes. Dibutyryl-cyclic 3[prime]5[prime] adenosinemonophosphate, papaverine, and genistein were found to inhibit the progression of gametogenesis in the light. Treatment of pregametes in the dark with either staurosporine or papaverine resulted in their conversion to mature gametes. Apparently, papaverine has different effects in the dark and in the light; the effect of staurosporine suggested that a protein kinase C-like component inhibits the conversion of pregametes to gametes, a block that normally is relieved by illumination. This hypothesis was corroborated by the observation that activators of protein kinase C, N-heptyl-5-chloro-1-naphthalenesulfonamide, N- (6-phenylhexyl)-5-chloro-1-naphthalenesulfonamide, and the phorbolester phorbol-12-myristate 13-acetate inhibited gametogenesis in the light. Genistein and dibutyryl-cyclic 3[prime]5[prime] adenosinemonophosphate were able to inhibit the dark activation caused by staurosporine treatment, suggesting that their targets work downstream from the “protein kinase C-like” kinase. Surprisingly, staurosporine and papaverine worked synergystically on the activation of pregametes in the dark.
Characterization of Novel Sesquiterpenoid Biosynthesis in Tobacco Expressing a Fungal Sesquiterpene Synthase
Authors: Zook M. , Hohn T. , Bonnen A. , Tsuji J. , Hammerschmidt R.
Published: October 2016

The gene encoding trichodiene synthase (Tri5), a sesquiterpene synthase from the fungus Fusarium sporotrichioides, was used to transform tobacco (Nicotiana tabacum). Trichodiene was the sole sesquiterpene synthase product in enzyme reaction mixtures derived from unelicited transformant cell-suspension cultures, and both trichodiene and 5-epi-aristolochene were observed as reaction products following elicitor treatment. Immunoblot analysis of protein extracts revealed the presence of trichodiene synthase only in transformant cell lines producing trichodiene. In vivo labeling with [3H]mevalonate revealed the presence of a novel trichodiene metabolite, 15-hydroxytrichodiene, that accumulated in the transformant cell-suspension cultures. In a trichodiene-producing transformant, the level of 15-hydroxytrichodiene accumulation increased after elicitor treatment. In vivo labeling with [14C]acetate showed that the biosynthetic rate of trichodiene and 15-hydroxytrichodiene also increased after elicitor treatment. Incorporation of radioactivity from [14C]acetate into capsidiol was reduced following elicitor treatment of a trichodiene-producing transformant as compared with wild type. These results demonstrate that sesquiterpenoid accumulation resulting from the constitutive expression of a foreign sesquiterpene synthase is responsive to elicitation and that the farnesyl pyrophosphate present in elicited cells can be utilized by a foreign sesquiterpene synthase to produce high levels of novel sesquiterpenoids.
Photosynthetic Gas Exchange and Discrimination against 13CO2 and C18O16O in Tobacco Plants Modified by an Antisense Construct to Have Low Chloroplastic Carbonic Anhydrase
Authors: Williams T. G. , Flanagan L. B. , Coleman J. R.
Published: October 2016

The physiological role of chloroplastic carbonic anhydrase (CA) was examined by antisense suppression of chloroplastic CA (on average 8% of wild type) in Nicotiana tabacum. Photosynthetic gas-exchange characteristics of low-CA and wild-type plants were measured concurrently with short-term, on-line stable isotope discrimination at varying vapor pressure deficit (VPD) and light intensity. Low-CA and wild-type plants were indistinguishable in the responses of assimilation, transpiration, stomatal conductance, and intercellular CO2 concentration to changing VPD or light intensity. At saturating light intensity, low-CA plants had lower discrimination against 13CO2 than wild-type plants by 1.2 to 1.8[per mille (thousand) sign]. Consequently, tissue of the low-CA plants was higher in 13C than the control plants. It was calculated that low-CA plants had chloroplast CO2 concentrations 13 to 22 [mu]mol mol-1 lower than wild-type plants. Discrimination against C18O16O in low-CA plants was 20% of that of the wild type, confirming a role of chloroplastic CA in the mechanism of discrimination against C18O16O ([delta]C18O16O). As VPD increased, stomatal closure caused a reduction in chloroplastic C02 concentration, and since VPD and chloroplastic CO2 concentration act in opposing directions on [delta]C18O16O, no effect of VPD was seen on [delta]C18O16O.
NaCI Reduces Indole-3-Acetic Acid Levels in the Roots of Tomato Plants Independent of Stress-Induced Abscisic Acid
Authors: Dunlap J. R. , Binzel M. L.
Published: October 2016

Indole-3-acetic acid (IAA) was measured in leaves and roots of tomato (Lycopersicon esculentum) genotypes subjected to salt stress. An abscisic acid (ABA)-deficient mutant of tomato (sitiens), the genetic parent (Rheinlands Ruhm, RR), and a commercial variety (Large Cherry Red, LCR) of tomato were treated with 50 to 300 mM NaCl in nutrient culture. Both LCR and RR had significantly higher levels of IAA in the roots compared with that in sitiens prior to treatment. The initial levels of IAA in the roots of LCR and RR declined by nearly 75% after exposure to NaCl, whereas those in roots from the sitiens mutant remained unchanged. IAA levels in the leaves of all genotypes remained unchanged or increased slightly in response to NaCl. ABA was highest in leaves from the normal genotypes after exposure to NaCl. ABA levels in the roots of sitiens were similar to the levels in the normal genotypes, whereas levels in the leaves were only 10% of the levels found in normal genotypes regardless of the salt treatment. Treatment of LCR and sitiens with exogenous ABA increased the ABA levels in leaves and roots, but there were no measurable changes in endogenous IAA. Therefore, the reduction in IAA appears to result from an ABA-independent effect of NaCl on IAA metabolism in the roots of stressed plants.
Expression of the Agrobacterium rhizogenes rolC Gene in a Deciduous Forest Tree Alters Growth and Development and Leads to Stem Fasciation
Authors: Nilsson O. , Moritz T. , Sundberg B. , Sandberg G. , Olsson O.
Published: October 2016

We have altered the growth and development of a deciduous forest tree by transforming hybrid aspen (Populus tremula x Populus tremuloides) with the Agrobacterium rhizogenes rolC gene expressed under the strong cauliflower mosaic virus 35S promoter. We demonstrate that the genetically manipulated perennial plants, after a period of dormancy, maintain the induced phenotypical changes during the second growing period. Furthermore, mass-spectrometrical quantifications of the free and conjugated forms of indole-3-acetic acid and cytokinins and several gibberellins on one transgenic line correlate the induced developmental alterations such as stem fasciation to changes in plant hormone metabolism. We also show that the presence of the RolC protein increases the levels of the free cytokinins, but not by a process involving hydrolysis of the inactive cytokinin conjugates.
Temporal and Spatial Expression of 1-Aminocyclopropane-1-Carboxylate Oxidase mRNA following Pollination of Immature and Mature Petunia Flowers
Authors: Tang X. , Woodson W. R.
Published: October 2016

Pollination of petunia (Petunia hybrida) flowers induces a rapid increase in ethylene production by styles, which subsequently leads to increased ethylene production by the corolla, inducing senescence. We have investigated the temporal and spatial expression of 1-aminocyclopropane-1-carboxylate (ACC) oxidase transcripts in petunia styles in an attempt to elucidate its role in increased ethylene biosynthesis following pollination. Previously, we reported that the development of petunia flowers was associated with increased ACC oxidase mRNA localized specifically in the stigmatic regions of the style (X. Tang, A.M.T. Gomes, A. Bhatia, W.R. Woodson [1994] Plant Cell 6: 1227–1239). The rapid increase in ethylene production by styles within the 1st h following pollination was correlated with the expression of ACC oxidase mRNAs during development. Pollination of petunia flowers prior to anthesis and the expression of ACC oxidase mRNA led to a substantial increase in ethylene production, but this was delayed by several hours in comparison with flowers at anthesis. This delayed increase in ethylene production by pollinated styles from immature flowers was associated with an increased ACC oxidase transcript abundance. Treatment with the ethylene action inhibitor 2,5-norbornadiene did not affect the early increase in ethylene production or the expression of ACC oxidase mRNAs. No differences in the rate of pollen germination or tube growth were detected when applied to stigmas from immature or mature flowers, indicating that the delay in ethylene production was likely the result of limited ACC oxidase activity. Localization of ACC oxidase mRNAs following pollination by in situ hybridization revealed an abundance of transcripts in transmitting tract tissue within 4 h of pollination of both immature and mature styles, in contrast to their localization in stigmatic cells during development.
Effect of CO2 Concentration on Carbonic Anhydrase and Ribulose-1,5-Bisphosphate Carboxylase/Oxygenase Expression in Pea
Authors: Majeau N. , Coleman J. R.
Published: October 2016

The effect of external CO2 concentration on the expression of carbonic anhydrase (CA) and ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) was examined in pea (Pisum sativum cv Little Marvel) leaves. Enzyme activities and their transcript levels were reduced in plants grown at 1000 [mu]L/L CO2 compared with plants grown in ambient air. Growth at 160 [mu]L/L CO2 also appeared to reduce steady-state transcript levels for rbcS, the gene encoding the small subunit of Rubisco, and for ca, the gene encoding CA; however, rbcS transcripts were reduced to a greater extent at this concentration. Rubisco activity was slightly lower in plants grown at 160 [mu]L/L CO2, and CA activity was significantly higher than that observed in air-grown plants. Transfer of plants from 1000 [mu]L/L to air levels of CO2 resulted in a rapid increase in both ca and rbcS transcript abundance in fully expanded leaves, followed by an increase in enzyme activity. Plants transferred from air to high-CO2 concentrations appeared to modulate transcript abundance and enzyme activity less quickly. Foliar carbohydrate levels were also examined in plants grown continuously at high and ambient CO2, and following changes in growth conditions that rapidly altered ca and rbcS transcript abundance and enzyme activities.