bluuebunny/crossref_metadata_2025_split huggingface dataset:
Gene Fusions of Signal Sequences with a Modified β-Glucuronidase Gene Results in Retention of the β-Glucuronidase Protein in the Secretory Pathway/Plasma Membrane
Authors: Yan X. , Gonzales R. A. , Wagner G. J. 
Published: July 2002

Abstract
Signal sequences and endoplasmic reticulum (ER) retention signals are known to play central roles in targeting and translocation in the secretory pathway, but molecular aspects about their involvement are poorly understood. We tested the effectiveness of deduced signal sequences from various genes (hydroxyproline-rich glycoprotein [HRGP] from Phaseolus vulgaris; Serpin from Manduca sexta) to direct a modified β-glucuronidase (GUS) protein into the secretory pathway in transgenic tobacco (Nicotiana tabacum L.). The reporter protein was not secreted to the cell wall/extracellular space as monitored using extracellular fluid analysis (low- or high-ionic-strength conditions) but occurred in membranes with a density of 1.16 to 1.20 g/mL. Membrane-bound GUS equilibrated with the plasma membrane (PM) and the ER on linear sucrose gradients with or without ethylenediaminetetraacetic acid, suggesting that GUS associates with the ER and the PM. Confocal microscopy of fixed cultured cells prepared from GUS control and HRGP signal peptide (SP)-GUS-expressing plants suggested only cytosolic localization in GUS-expressing plants but substantial peripheral localization in HRGP SP-GUS plants, which is consistent with GUS being associated with the PM. Aqueous two-phase partitioning of microsomal membranes from HRGP SP-GUS and Serpin SP-GUS transgenic leaves also indicated that GUS activity was enriched in the ER and the PM. These observations, together with hydrophobic moment plot analysis, suggest that properties of the SP-GUS protein result in its retention in the secretory pathway and PM.
Endogenous Methyl Salicylate in Pathogen-Inoculated Tobacco Plants1
Authors: Seskar Mirjana , Shulaev Vladimir , Raskin Ilya 
Published: July 2002

Abstract
The tobacco (Nicotiana tabacum) cultivar Xanthi-nc (genotype NN) produces high levels of salicylic acid (SA) after inoculation with the tobacco mosaic virus (TMV). Gaseous methyl salicylate (MeSA), a major volatile produced in TMV-inoculated tobacco plants, was recently shown to be an airborne defense signal. Using an assay developed to measure the MeSA present in tissue, we have shown that in TMV-inoculated tobacco plants the level of MeSA increases dramatically, paralleling increases in SA. MeSA accumulation was also observed in upper, noninoculated leaves. In TMV-inoculated tobacco shifted from 32 to 24°C, the MeSA concentration increased from nondetectable levels to 2318 ng/g fresh weight 12 h after the temperature shift, but subsequently decreased with the onset of the hypersensitive response. Similar results were observed in plants inoculated withPseudomonas syringae pathovarphaseolicola, in which MeSA levels were highest just before the hypersensitive response-induced tissue desiccation. Transgenic NahG plants unable to accumulate SA also did not accumulate MeSA after TMV inoculation, and did not show increased resistance to TMV following MeSA treatment. Based on the spatial and temporal kinetics of its accumulation, we conclude that tissue MeSA may play a role similar to that of volatile MeSA in the pathogen-induced defense response.
Stress Induction of Mitochondrial Formate Dehydrogenase in Potato Leaves1
Authors: Hourton-Cabassa Cécile , Ambard-Bretteville Françoise , Moreau François , de Virville Jacques Davy , Rémy René , Colas des Francs-Small Catherine 
Published: July 2002

Abstract
In higher plants formate dehydrogenase (FDH, EC 1.2.1.2.) is a mitochondrial, NAD-dependent enzyme. We previously reported that in potato (Solanum tuberosum L.) FDH expression is high in tubers but low in green leaves. Here we show that in isolated tuber mitochondria FDH is involved in formate-dependent O2 uptake coupled to ATP synthesis. The effects of various environmental and chemical factors on FDH expression in leaves were tested using the mitochondrial serine hydroxymethyltransferase as a control. The abundance of FDH transcripts is strongly increased under various stresses, whereas serine hydroxymethyltransferase transcripts decline. The application of formate to leaves strongly enhances FDH expression, suggesting that it might be the signal for FDH induction. Our experiments using glycolytic products suggest that glycolysis may play an important role in formate synthesis in leaves in the dark and during hypoxia, and in tubers. Of particular interest is the dramatic accumulation of FDH transcripts after spraying methanol on leaves, as this compound is known to increase the yields of C3 plants. In addition, although the steady-state levels of FDH transcript increase very quickly in response to stress, protein accumulation is much slower, but can eventually reach the same levels in leaves as in tubers.
Role of the Ascorbate-Glutathione Cycle of Mitochondria and Peroxisomes in the Senescence of Pea Leaves
Authors: Jiménez Ana , Hernández José A. , Pastori Gabriela , del Rı́o Luis A. , Sevilla Francisca 
Published: July 2002

Abstract
We investigated the relationship between H2O2 metabolism and the senescence process using soluble fractions, mitochondria, and peroxisomes from senescent pea (Pisum sativum L.) leaves. After 11 d of senescence the activities of Mn-superoxide dismutase, dehydroascorbate reductase (DHAR), and glutathione reductase (GR) present in the matrix, and ascorbate peroxidase (APX) and monodehydroascorbate reductase (MDHAR) activities localized in the mitochondrial membrane, were all substantially decreased in mitochondria. The mitochondrial ascorbate and dehydroascorbate pools were reduced, whereas the oxidized glutathione levels were maintained. In senescent leaves the H2O2 content in isolated mitochondria and the NADH- and succinate-dependent production of superoxide (O2 ·−) radicals by submitochondrial particles increased significantly. However, in peroxisomes from senescent leaves both membrane-bound APX and MDHAR activities were reduced. In the matrix the DHAR activity was enhanced and the GR activity remained unchanged. As a result of senescence, the reduced and the oxidized glutathione pools were considerably increased in peroxisomes. A large increase in the glutathione pool and DHAR activity were also found in soluble fractions of senescent pea leaves, together with a decrease in GR, APX, and MDHAR activities. The differential response to senescence of the mitochondrial and peroxisomal ascorbate-glutathione cycle suggests that mitochondria could be affected by oxidative damage earlier than peroxisomes, which may participate in the cellular oxidative mechanism of leaf senescence longer than mitochondria.
Arabidopsis Roots and Shoots Have Different Mechanisms for Hypoxic Stress Tolerance
Authors: Ellis Marc H. , Dennis Elizabeth S. , James Peacock W. 
Published: July 2002

Abstract
Arabidopsis has inducible responses for tolerance of O2 deficiency. Plants previously exposed to 5% O2 were more tolerant than the controls to hypoxic stress (0.1% O2 for 48 h) in both roots and shoots, but hypoxic acclimation did not improve tolerance to anoxia (0% O2). The acclimation of shoots was not dependent on the roots: increased shoot tolerance was observed when the roots of the plants were removed. An adh(alcohol dehydrogenase) null mutant did not show acclimation of the roots but retained the shoot survival response. Abscisic acid treatment also differentiated the root and shoot responses; pretreatment induced root survival in hypoxic stress conditions (0.1% O2) but did not induce any increase in the survival of shoots. Cycloheximide blocked both root and shoot acclimation, indicating that both acclimation mechanisms are dependent on protein synthesis.
Proline Accumulation in Developing Grapevine Fruit Occurs Independently of Changes in the Levels of Δ1-Pyrroline-5-Carboxylate Synthetase mRNA or Protein1
Authors: Stines Anna P. , Naylor Dean J. , Høj Peter B. , van Heeswijck Robyn 
Published: July 2002

AbstractMature fruit of grapevine (Vitis vinifera) contains unusually high levels of free proline (Pro; up to 24 μmol or 2.8 mg/g fresh weight). Pro accumulation does not occur uniformly throughout berry development but only during the last 4 to 6 weeks of ripening when both berry growth and net protein accumulation have ceased. In contrast, the steady-state levels of both the mRNA encoding V. viniferaΔ1-pyrroline-5-carboxylate synthetase (VVP5CS), a key regulatory enzyme in Pro biosynthesis, and its protein product remain relatively uniform throughout fruit development. In addition, the steady-state protein levels of Pro dehydrogenase, the first enzyme in Pro degradation, increased throughout early fruit development but thereafter remained relatively constant. The developmental accumulation of free Pro late in grape berry ripening is thus clearly distinct from the osmotic stress-induced accumulation of Pro in plants. It is not associated with either sustained increases in steady-state levels of P5CS mRNA or protein or a decrease in steady-state levels of Pro dehydrogenase protein, suggesting that other physiological factors are important for its regulation.
Carbon and Amino Acids Reciprocally Modulate the Expression of Glutamine Synthetase in Arabidopsis
Authors: Oliveira Igor C. , Coruzzi Gloria M. 
Published: July 2002

Abstract
In bacteria and yeast, glutamine synthetase (GS) expression is tightly regulated by the metabolic status of the cell, both at the transcriptional and posttranscriptional levels. We discuss the relative contributions of light and metabolic cues on the regulation of members of the GS gene family (chloroplastic GS2 and cytosolic GS1) in Arabidopsis. These studies reveal that the dramatic induction of mRNA for chloroplastic GS2 by light is mediated in part by phytochrome and in part by light-induced changes in sucrose (Suc) levels. In contrast, the modest induction of mRNA for cytosolic GS1 by light is primarily mediated by changes in the levels of carbon metabolites. Suc induction of mRNA for GS2 and GS1 occurs in a time- and dose-dependent manner. Suc-induced changes in GS mRNA levels were also observed at the level of GS enzyme activity. In contrast, amino acids were shown to antagonize the Suc induction of GS, both at the level of mRNA accumulation and that of enzyme activity. For GS2, the gene whose expression was the most dramatically regulated by metabolites, we used a GS2 promoter-β-glucuronidase fusion to demonstrate that transcriptional control is involved in this metabolic regulation. Our results suggest that the metabolic regulation of GS expression in plants is controlled by the relative abundance of carbon skeletons versus amino acids. This would allow nitrogen assimilation into glutamine to proceed (or not) according to the metabolic status and biosynthetic needs of the plant. This type of GS gene regulation is reminiscent of the nitrogen regulatory system in bacteria, and suggests an evolutionary link between metabolic sensing and signaling in bacteria and plants.
A Proline-, Threonine-, and Glycine-Rich Protein Down-Regulated by Drought Is Localized in the Cell Wall of Xylem Elements
Authors: Harrak Hassan , Chamberland Hélène , Plante Michèle , Bellemare Guy , Lafontaine Jean G. , Tabaeizadeh Zohreh 
Published: July 2002

Abstract
A cDNA clone encoding a proline-, threonine-, and glycine-rich protein (PTGRP) was isolated from a wild tomato species (Lycopersicon chilense) (L.X. Yu, H. Chamberland, J.G. Lafontain, Z. Tabaeizadeh [1996] Genome 39: 1185–1193). Northern-blot analysis and in situ hybridization studies revealed that PTGRP is down-regulated by drought stress. The level of the mRNA in leaves and stems of 8-d drought-stressed plants decreased 5- to 10-fold compared with that in regularly watered plants. The mRNA re-accumulated when drought-stressed plants were rewatered. Antibodies raised against a glutathione S-transferase/PTGRP fusion protein were used to elucidate the subcellular localization of the protein by immunogold labeling. In regularly watered L. chilense plants, PTGRP protein was found to be localized in xylem pit membranes and disintegrated primary walls. Examination of sections from drought-stressed plants revealed a significant decrease in the levels of labeling. In these samples, only a few scattered gold particles were detected in the same areas. In the leaf tissues of plants that had been rewatered for 3 d following an 8-d drought stress, the labeling pattern was similar to that of the regularly watered plants. To our knowledge, PTGRP is the first drought-regulated protein that has been precisely localized in the cell wall.
Snow-Mold-Induced Apoplastic Proteins in Winter Rye Leaves Lack Antifreeze Activity
Authors: Hiilovaara-Teijo Mervi , Hannukkala Asko , Griffith Marilyn , Yu Xiao-Ming , Pihakaski-Maunsbach Kaarina 
Published: July 2002

Abstract
During cold acclimation, winter rye (Secale cereale L.) plants secrete antifreeze proteins that are similar to pathogenesis-related (PR) proteins. In this experiment, the secretion of PR proteins was induced at warm temperatures by infection with pink snow mold (Microdochium nivale), a pathogen of overwintering cereals. A comparison of cold-induced and pathogen-induced proteins showed that PR proteins accumulated in the leaf apoplast to a greater level in response to cold. The PR proteins induced by cold and by snow mold were similar when separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and examined by immunoblotting. Both groups of PR proteins contained glucanase-like, chitinase-like, and thaumatin-like proteins, and both groups exhibited similar levels of glucanase and chitinase activities. However, only the PR proteins induced by cold exhibited antifreeze activity. Our findings suggest that the cold-induced PR proteins may be isoforms that function as antifreeze proteins to modify the growth of ice during freezing while also providing resistance to the growth of low-temperature pathogens in advance of infection. Both functions of the cold-induced PR proteins may improve the survival of overwintering cereals.
Mercuric Chloride Effects on Root Water Transport in Aspen Seedlings
Authors: Wan Xianchong , Zwiazek Janusz J. 
Published: July 2002

Abstract
HgCl2 (0.1 mm) reduced pressure-induced water flux and root hydraulic conductivity in the roots of 1-year-old aspen (Populus tremuloides Michx.) seedlings by about 50%. The inhibition was reversed with 50 mm mercaptoethanol. Mercurial treatment reduced the activation energy of water transport in the roots from 10.82 ± 0.700 kcal mol−1 to 6.67 ± 0.193 kcal mol−1 when measured over the 4°C to 25°C temperature range. An increase in rhodamine B concentration in the xylem sap of mercury-treated roots suggested a decrease in the symplastic transport of water. However, the apoplastic pathway in both control and mercury-treated roots constituted only a small fraction of the total root water transport. Electrical conductivity and osmotic potentials of the expressed xylem sap suggested that 0.1 mmHgCl2 and temperature changes over the 4°C to 25°C range did not induce cell membrane leakage. The 0.1 mmHgCl2 solution applied as a root drench severely reduced stomatal conductance in intact plants, and this reduction was partly reversed by 50 mm mercaptoethanol. In excised shoots, 0.1 mm HgCl2 did not affect stomatal conductance, suggesting that the signal that triggered stomatal closure originated in the roots. We suggest that mercury-sensitive processes in aspen roots play a significant role in regulating plant water balance by their effects on root hydraulic conductivity.
Interaction of a Plant 14-3-3 Protein with the Signal Peptide of a Thylakoid-Targeted Chloroplast Precursor Protein and the Presence of 14-3-3 Isoforms in the Chloroplast Stroma
Authors: Sehnke Paul C. , Henry Ralph , Cline Kenneth , Ferl Robert J. 
Published: July 2002

Abstract
The 14-3-3 proteins are acidic, dimeric proteins that have been implicated in many eukaryotic cellular processes because of direct protein association with enzymes and other metabolic and regulatory proteins. 14-3-3 proteins are largely considered to be cytoplasmic, but a search for proteins that specifically interact with a plant 14-3-3 resulted in the isolation of a nuclear-encoded, thylakoid-targeted chloroplast precursor, the full-length Arabidopsis photosystem I N-subunit At pPSI-N (P.C. Sehnke, R.J. Ferl [1995] Plant Physiol 109: 1126). Using precursor truncations in the two-hybrid system, it was determined that the leader sequence is the site of PSI-N that associates with 14-3-3. This suggested the novel possibility that 14-3-3 would be found within chloroplasts. Immuno-electron microscopy of leaf tissue and western analysis of chloroplast fractions with monoclonal anti-14-3-3 antibodies localized 14-3-3 proteins to the chloroplast stroma and the stromal side of thylakoid membranes. Using peptide-generated, isoform-specific antibodies, GF14ν, GF14ε, GF14μ, and GF14υ were shown to be present in the chloroplast stromal extract. These isoforms represent two distinct phylogenetic 14-3-3 groupings. These data suggest a novel interorganellar role for these phylogenetically distinct 14-3-3 proteins.
Regulatory Role of the N Terminus of the Vacuolar Calcium-ATPase in Cauliflower
Authors: Malmström Susanna , Åkerlund Hans-Erik , Askerlund Per 
Published: July 2002

Abstract
The vacuolar calmodulin (CaM)-stimulated Ca2+-ATPase, BCA1p, in cauliflower (Brassica oleracea) has an extended N terminus, which was suggested to contain a CaM-binding domain (S. Malmström, P. Askerlund, M.G. Palmgren [1997] FEBS Lett 400: 324–328). The goal of the present study was to determine the role of the N terminus in regulating BCA1p. Western analysis using three different antisera showed that the N terminus of BCA1p is cleaved off by trypsin and that the N terminus contains the CaM-binding domain. Furthermore, the expressed N terminus binds CaM in a Ca2+-dependent manner. A synthetic peptide corresponding to the CaM-binding domain of BCA1p (Ala-19 to Leu-43) strongly inhibited ATP-dependent Ca2+pumping by BCA1p in cauliflower low-density membranes, indicating that the CaM-binding region of BCA1p also has an autoinhibitory function. The expressed N terminus of BCA1p and a synthetic peptide (Ala-19 to Met-39) were good substrates for phosphorylation by protein kinase C. Sequencing of the phosphorylated fusion protein and peptide suggested serine-16 and/or serine-28 as likely targets for phosphorylation. Phosphorylation of serine-28 had no effect on CaM binding to the alanine-19 to methionine-39 peptide. Our results demonstrate the regulatory importance of the N terminus of BCA1p as a target for CaM binding, trypsin cleavage, and phosphorylation, as well as its importance as an autoinhibitory domain.
Ectoine, the Compatible Solute of Halomonas elongata, Confers Hyperosmotic Tolerance in Cultured Tobacco Cells
Authors: Nakayama Hideki , Yoshida Kazuya , Ono Hisayo , Murooka Yoshikatsu , Shinmyo Atsuhiko 
Published: July 2002

Abstract
1,4,5,6-Tetrahydro-2-methyl-4-pyrimidinecarboxylic acid (ectoine) functions as a compatible osmolyte in the moderate halophile Halomonas elongata OUT30018. Ectoine is biosynthesized by three successive enzyme reactions from aspartic β-semialdehyde. The genes encoding the enzymes involved in the biosynthesis, ectA, ectB, andectC, encoding l-2,4-diaminobutyric acid acetyltransferase, l-2,4-diaminobutyric acid transaminase, and l-ectoine synthase, respectively, have been previously cloned. To investigate the function of ectoine as a compatible solute in plant cells, the three genes were individually placed under the control of the cauliflower mosaic virus 35S promoter and introduced together into cultured tobacco (Nicotiana tabacum L.) cv Bright Yellow 2 (BY2) cells. The transgenic BY2 cells accumulated a small quantity of ectoine (14–79 nmol g−1 fresh weight) and showed increased tolerance to hyperosmotic shock (900 mOsm). Furthermore, the transgenic BY2 cells exhibited a normal growth pattern even under hyperosmotic conditions (up to 530 mOsm), in which the growth of the untransformed BY2 (wild type) cells was obviously delayed. These results suggest that genetically engineered synthesis of ectoine results in the increased hyperosmotic tolerance of cultured tobacco BY2 cells despite the low level of accumulation of the solute.
The Carboxyl-Terminal Extension of the Precursor D1 Protein of Photosystem II Is Required for Optimal Photosynthetic Performance of the Cyanobacterium Synechocystis sp. PCC 6803
Authors: Ivleva Natalia B. , Shestakov Sergey V. , Pakrasi Himadri B. 
Published: July 2002

Abstract
The D1 protein is an integral component of the photosystem II reaction center complex. In the cyanobacteriumSynechocystis sp. PCC 6803, D1 is synthesized with a short 16-amino acids-long carboxyl-terminal extension. Removal of this extension is necessary to form active oxygen-evolving photosystem II centers. Our earlier studies have shown that this extension is cleaved by CtpA, a specific carboxyl-terminal processing protease. The amino acid sequence of the carboxyl-terminal extension is conserved among D1 proteins from different organisms, although at a level lower than that of the mature protein. In the present study we have analyzed a mutant strain of Synechocystis sp. PCC 6803 with a duplicated extension, and a second mutant that lacks the extension, to investigate the effects of these alterations on the function of the D1 protein in vivo. No significant difference in the growth rates, photosynthetic pigment composition, fluorescence induction, and oxygen evolution rates was observed between the mutants and the control strain. However, using long-term mixed culture growth analysis, we detected significant decreases in the fitness of these mutant strains. The presented data demonstrate that the carboxyl-terminal extension of the precursor D1 protein is required for optimal photosynthetic performance.
Developmental Regulation of Indole-3-Acetic Acid Turnover in Scots Pine Seedlings
Authors: Ljung Karin , Östin Anders , Lioussanne Laetitia , Sandberg Göran 
Published: July 2002

Abstract
Indole-3-acetic acid (IAA) homeostasis was investigated during seed germination and early seedling growth in Scots pine (Pinus sylvestris). IAA-ester conjugates were initially hydrolyzed in the seed to yield a peak of free IAA prior to initiation of root elongation. Developmental regulation of IAA synthesis was observed, with tryptophan-dependent synthesis being initiated around 4 d and tryptophan-independent synthesis occurring around 7 d after imbibition. Induction of catabolism to yield 2-oxindole-3-acetic acid and irreversible conjugation to indole-3-acetyl-N-aspartic acid was noticed at the same time as de novo synthesis was first detected. As a part of the homeostatic regulation IAA was further metabolized to two new conjugates: glucopyranosyl-1-N-indole-3-acetyl-N-aspartic acid and glucopyranosyl-1-N-indole-3-acetic acid. The initial supply of IAA thus originates from stored pools of IAA-ester conjugates, mainly localized in the embryo itself rather than in the general nutrient storage tissue, the megagametophyte. We have found that de novo synthesis is first induced when the stored pool of conjugated IAA is used up and additional hormone is needed for elongation growth. It is interesting that when de novo synthesis is induced, a distinct induction of catabolic events occurs, indicating that the seedling needs mechanisms to balance synthesis rates for the homeostatic regulation of the IAA pool.