bluuebunny/crossref_metadata_2025_split huggingface dataset:
Aspartic Acid 413 Is Important for the Normal Allosteric Functioning of ADP-Glucose Pyrophosphorylase
Authors: Greene T. W. , Woodbury R. L. , Okita T. W. 
Published: July 2002

Abstract
As part of a structure-function analysis of the higher-plant ADP-glucose pyrophosphorylase (AGP), we used a random mutagenesis approach in combination with a novel bacterial complementation system to isolate over 100 mutants that were defective in glycogen production (T.W. Greene, S.E. Chantler, M.L. Khan, G.F. Barry, J. Preiss, T.W. Okita [1996] Proc Natl Acad Sci USA 93: 1509–1513). One mutant of the large subunit M27 was identified by its capacity to only partially complement a mutation in the structural gene for the bacterial AGP (glg C), as determined by its light-staining phenotype when cells were exposed to I2 vapors. Enzyme-linked immunosorbent assay and enzymatic pyrophosphorylysis assays of M27 cell extracts showed that the level of expression and AGP activity was comparable to those of cells that expressed the wild-type recombinant enzyme. Kinetic analysis indicated that the M27 AGP displays normal Michaelis constant values for the substrates glucose-1-phosphate and ATP but requires 6- to 10-fold greater levels of 3-phosphoglycerate (3-PGA) than the wild-type recombinant enzyme for maximum activation. DNA sequence analysis showed that M27 contains a single point mutation that resulted in the replacement of aspartic acid 413 to alanine. Substitution of a lysine residue at this site almost completely abolished activation by 3-PGA. Aspartic acid 413 is adjacent to a lysine residue that was previously identified by chemical modification studies to be important in the binding of 3-PGA (K. Ball, J. Preiss [1994] J Biol Chem 269: 24706–24711). The kinetic properties of M27 corroborate the importance of this region in the allosteric regulation of a higher-plant AGP.
Molecular Cloning of a Ripening-Specific Lipoxygenase and Its Expression during Wild-Type and Mutant Tomato Fruit Development
Authors: Kausch K. D. , Handa A. K. 
Published: July 2002

Abstract
A 94-kD protein that accumulates predominately in tomato (Lycopersicon esculentum) fruit during ripening was purified, and antibodies specific for the purified protein were used to isolate cDNA clones from a red-ripe fruit cDNA library. A sequence analysis of these cDNAs and cross-reactivity of the 94-kD-specific antibodies to the soybean lipoxygenase (LOX) L-1, L-2, and L-3 proteins and soybean LOX L-1-specific antibodies to the 94-kD protein identified it as a member of the LOX gene family. Maximum levels of the 94-kD LOX mRNA and protein are present in breaker to ripe and red-ripe stages, respectively. Expression of 94-kD LOX in different tissues from mature green and red-ripe tomato fruits was found to be greatest in the radial walls of ripe fruit, but immunocytolocalization using tissue printing suggests that the highest accumulation of its protein occurs in locular jelly. None of 94-kD LOX is expressed in nonripening mutant fruits of any age. Never-ripe mutant fruit accumulate the 94-kD LOX mRNA to levels similar to those obtained in wild-type fruit, but fail to accumulate the 94-kD LOX protein. Collectively, the results show that expression of 94-kD LOX is regulated by the ripening process, and ethylene may play a role in its protein accumulation.
Changes in the Level and Activation State of the Plasma Membrane H+-ATPase during Aging of Red Beet Slices
Authors: Papini R. , De Michelis M. I. 
Published: July 2002

Abstract
The effect of aging on the plasma membrane (PM) H+-ATPase of red beet (Beta vulgaris L.) parenchyma discs was analyzed in PM purified by aqueous two-phase partitioning. Aging increased both the activity and the amount of immunodetectable H+-ATPase in the PM. The activity assayed at slightly alkaline pH values increased earlier and more strongly than that assayed at acidic pH values, so that the pH curve of the enzyme from aged beet discs was shifted toward more alkaline values. Aging decreased the stimulation of the PM H+-ATPase activity by controlled trypsin treatments or by lysophosphatidylcholine. After trypsin treatment the pH dependence of H+-ATPase from dormant or aged beet discs became equal. These results indicate that aging not only increases the level of H+-ATPase in the PM, but also determines its activation, most likely by modifying the interaction between the autoinhibitory carboxyl-terminal domain and the catalytic site. When the PM H+-ATPase activity was assayed at a slightly alkaline pH, the tyrosine modifier N-acetylimidazole inhibited the H+-ATPase in the PM from dormant beet discs much less than in the PM from aged discs, suggesting that modification of a tyrosine residue may be involved in the activation of the PM H+-ATPase induced by aging. The results are discussed with regard to aging-induced development of transmembrane transport activities.
The Dark-Adaptation Response of the De-Etiolated Pea Mutant lip1 Is Modulated by External Signals and Endogenous Programs
Authors: Frances S. , Thompson W. F. 
Published: July 2002

Abstract
The lip1 mutant of pea (Pisum sativum L.) exhibits a de-etiolated phenotype. When grown in darkness, lip1 plants have several characteristics normally associated only with light-grown plants. Young wild-type (WT) seedlings accumulate high levels of transcripts from plastid-related genes (such as those encoding chlorophyll a/b-binding proteins, ferredoxin, and the small subunit of Rubisco) only in the light. In contrast, regardless of the light conditions under which the plants are grown, young mutant seedlings accumulate transcript levels equal to or greater than those seen in light-grown WT seedlings of the same age. Under some conditions, light-grown lip1 seedlings failed to respond to dark treatment. The largest response to darkness observed in the mutant occurred when older seedlings were first grown under low-light conditions before transfer to darkness. The mutant's inability to respond to darkness is not due to a gross disturbance in the circadian clock. We conclude that environmental signals (light) and endogenous programs (developmental and circadian) regulate gene expression in both WT and mutant plants. However, mutant seedlings exhibit a developmentally regulated and exaggerated response to light. In addition, the effect of the mutation may be greatest during a brief period early in development.
Characterization of the Common Bean Uricase II and Its Expression in Organs Other than Nodules
Authors: Capote-Mainez N. , Sanchez F. 
Published: July 2002

Abstract
Uricase II is a purine metabolic enzyme highly induced in root nodules during the symbiosis established between legumes and bacteria of the genera Rhizobium and Bradyrhizobium. Here we describe the characterization of bean (Phaseolus vulgaris) nodule uricase II cDNA and show that uricase II is encoded by a single gene in the bean genome. This gene is also expressed in cotyledons, roots, and hypocotyls during bean seedling establishment, and an anti-uricase antibody recognizes the protein in different seedling organs. Uricase II has also been found in Leucaena leucocephala seedlings, suggesting that it participates during seedling establishment in legumes that do not transport ureides. A 50-kD polypeptide that is detected by the anti-uricase antibody is found in cotyledons during seedling development. This higher-molecular-mass form is also detected in developing roots and hypocotyls but not in nodules. In situ hybridization experiments in root seedlings showed uricase II transcripts in the metaxylem parenchyma cells and phloem fibers of the vascular system.
Uridine 5′-Diphosphate-Glucose Dehydrogenase from Soybean Nodules
Authors: Stewart Douglas C. , Copeland Les 
Published: July 2002

Abstract
A highly purified preparation of uridine 5′-diphosphate (UDP)-glucose (Glc) dehydrogenase (DH; EC 1.1.1.22) has been characterized from soybean (Glycinemax L.) nodules. The enzyme had native and subunit molecular masses of approximately 272 and 50 kD, respectively. UDP-Glc DH displayed typical hyperbolic substrate kinetics and hadKm values for UDP-Glc and NAD+of 0.05 and 0.12 mm, respectively. Thymidine 5′-diphosphate-Glc and UDP-galactose could replace UDP-Glc as the sugar nucleotide substrate to some extent, but the enzyme had no activity with NADP+. Soybean nodule UDP-Glc DH was labile in the absence of NAD+ and was inhibited by a heat-stable, low-molecular-mass solute in crude extracts of soybean nodules. UDP-Glc DH was also isolated from developing soybean seeds and shoots of 5-d-old wheat and canola seedlings and was shown to have similar affinities for UDP-Glc and NAD+ as those of the soybean nodule enzyme. UDP-Glc DH from all of these sources was most active in young, rapidly growing tissues.
Cloning of a Tobacco Apoplasmic Invertase Inhibitor1
Authors: Greiner Steffen , Krausgrill Silke , Rausch Thomas 
Published: July 2002

Abstract
Higher plants express several isoforms of vacuolar and cell wall invertases (CWI), some of which are inactivated by inhibitory proteins at certain stages of plant development. We have purified an apoplasmic inhibitor (INH) of tobacco (Nicotiana tabacum) CWI to homogeneity. Based on sequences from tryptic fragments, we have isolated a full-length INH-encoding cDNA clone (Nt-inh1) via a reverse transcriptase-polymerase chain reaction. Southern-blot analysis revealed that INH is encoded by a single- or low-copy gene. Comparison with expressed sequence tag clones from Arabidopsis thaliana and Citrus unshiu indicated the presence of Nt-inh1-related proteins in other plants. The recombinantNt-inh1-encoded protein inhibits CWI from tobacco andChenopodium rubrum suspension-cultured cells and vacuolar invertase from tomato (Lycopersicon esculentum) fruit, whereas yeast invertase is not affected. However, only in the homologous system is the inhibition modulated by the concentration of Suc as previously shown for INH isolated from tobacco cells. Highly specific binding of INH to CWI could be shown by affinity chromatography of a total cell wall protein fraction on immobilized recombinant Nt-inh1 protein. RNA-blot analysis of relative transcript ratios for Nt-inh1 and CWI in different parts of adult tobacco plants revealed that the expression of both proteins is not always coordinate.
Induced β-Carotene Synthesis Driven by Triacylglycerol Deposition in the Unicellular AlgaDunaliella bardawil1
Authors: Rabbani Said , Beyer Peter , Lintig Johannes v. , Hugueney Philippe , Kleinig Hans 
Published: July 2002

AbstractUnder stress conditions such as high light intensity or nutrient starvation, cells of the unicellular alga Dunaliella bardawil overproduce β-carotene, which is accumulated in the plastids in newly formed triacylglycerol droplets. We report here that the formation of these sequestering structures and β-carotene are interdependent. When the synthesis of triacylglycerol is blocked, the overproduction of β-carotene is also inhibited. During overproduction of β-carotene no up-regulation of phytoene synthase or phytoene desaturase is observed on the transcriptional or translational level, whereas at the same time acetyl-CoA carboxylase, the key regulatory enzyme of acyl lipid biosynthesis, is increased, at least in its enzymatic activity. We conclude that under normal conditions the carotenogenic pathway is not maximally active and may be appreciably stimulated in the presence of sequestering structures, creating a plastid-localized sink for the end product of the carotenoid biosynthetic pathway.
In Vitro Reconstitution of Electron Transport from Glucose-6-Phosphate and NADPH to Nitrite1
Authors: Jin Tie , Huppe Heather C. , Turpin David H. 
Published: July 2002

Abstract
An NADPH-dependent NO2−-reducing system was reconstituted in vitro using ferredoxin (Fd) NADP+oxidoreductase (FNR), Fd, and nitrite reductase (NiR) from the green alga Chlamydomonas reinhardtii. NO2− reduction was dependent on all protein components and was operated under either aerobic or anaerobic conditions. NO2− reduction by this in vitro pathway was inhibited up to 63% by 1 mm NADP+. NADP+ did not affect either methyl viologen-NiR or Fd-NiR activity, indicating that inhibition was mediated through FNR. When NADPH was replaced with a glucose-6-phosphate dehydrogenase (G6PDH)-dependent NADPH-generating system, rates of NO2− reduction reached approximately 10 times that of the NADPH-dependent system. G6PDH could be replaced by either 6-phosphogluconate dehydrogenase or isocitrate dehydrogenase, indicating that G6PDH functioned to: (a) regenerate NADPH to support NO2− reduction and (b) consume NADP+, releasing FNR from NADP+ inhibition. These results demonstrate the ability of FNR to facilitate the transfer of reducing power from NADPH to Fd in the direction opposite to that which occurs in photosynthesis. The rate of G6PDH-dependent NO2− reduction observed in vitro is capable of accounting for the observed rates of dark NO3−assimilation by C. reinhardtii.
Mutation of Residue Threonine-2 of the D2 Polypeptide and Its Effect on Photosystem II Function inChlamydomonas reinhardtii1
Authors: Andronis Christos , Kruse Olaf , Deák Zsuzsanna , Vass Imre , Diner Bruce A. , Nixon Peter J. 
Published: July 2002

Abstract
The D2 polypeptide of the photosystem II (PSII) complex in the green algaChlamydomonas reinhardtii is thought to be reversibly phosphorylated. By analogy to higher plants, the phosphorylation site is likely to be at residue threonine-2 (Thr-2). We have investigated the role of D2 phosphorylation by constructing two mutants in which residue Thr-2 has been replaced by either alanine or serine. Both mutants grew photoautotrophically at wild-type rates, and noninvasive biophysical measurements, including the decay of chlorophyll fluorescence, the peak temperature of thermoluminescence bands, and rates of oxygen evolution, indicate little perturbation to electron transfer through the PSII complex. The susceptibility of mutant PSII to photoinactivation as measured by the light-induced loss of PSII activity in whole cells in the presence of the protein-synthesis inhibitors chloramphenicol or lincomycin was similar to that of wild type. These results indicate that phosphorylation at Thr-2 is not required for PSII function or for protection from photoinactivation. In control experiments the phosphorylation of D2 in wild-type C. reinhardtii was examined by 32P labeling in vivo and in vitro. No evidence for the phosphorylation of D2 in the wild type could be obtained. [14C]Acetate-labeling experiments in the presence of an inhibitor of cytoplasmic protein synthesis also failed to identify phosphorylated (D2.1) and nonphosphorylated (D2.2) forms of D2 upon sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Our results suggest that the existence of D2 phosphorylation in C. reinhardtii is still in question.
Plasma Membrane-Associated Actin in Bright Yellow 2 Tobacco Cells
Authors: Collings David A. , Asada Tetsuhiro , Allen Nina S. , Shibaoka Hiroh 
Published: July 2002

Abstract
Plasma membrane ghosts form when plant protoplasts attached to a substrate are lysed to leave a small patch of plasma membrane. We have identified several factors, including the use of a mildly acidic actin stabilization buffer and the inclusion of glutaraldehyde in the fixative, that allow immunofluorescent visualization of extensive cortical actin arrays retained on membrane ghosts made from tobacco (Nicotiana tabacum L.) suspension-cultured cells (line Bright Yellow 2). Normal microtubule arrays were also retained using these conditions. Membrane-associated actin is random; it exhibits only limited coalignment with the microtubules, and microtubule depolymerization in whole cells before wall digestion and ghost formation has little effect on actin retention. Actin and microtubules also exhibit different sensitivities to the pH and K+ and Ca2+ concentrations of the lysis buffer. There is, however, strong evidence for interactions between actin and the microtubules at or near the plasma membrane, because both ghosts and protoplasts prepared from taxol-pretreated cells have microtubules arranged in parallel arrays and an increased amount of actin coaligned with the microtubules. These experiments suggest that the organization of the cortical actin arrays may be dependent on the localization and organization of the microtubules.
Regulation of Ferulate-5-Hydroxylase Expression in Arabidopsis in the Context of Sinapate Ester Biosynthesis1
Authors: Ruegger Max , Meyer Knut , Cusumano Joanne C. , Chapple Clint 
Published: July 2002

Abstract
Sinapic acid is an intermediate in syringyl lignin biosynthesis in angiosperms, and in some taxa serves as a precursor for soluble secondary metabolites. The biosynthesis and accumulation of the sinapate esters sinapoylglucose, sinapoylmalate, and sinapoylcholine are developmentally regulated in Arabidopsis and other members of the Brassicaceae. The FAH1 locus of Arabidopsis encodes the enzyme ferulate-5-hydroxylase (F5H), which catalyzes the rate-limiting step in syringyl lignin biosynthesis and is required for the production of sinapate esters. Here we show that F5H expression parallels sinapate ester accumulation in developing siliques and seedlings, but is not rate limiting for their biosynthesis. RNA gel-blot analysis indicated that the tissue-specific and developmentally regulated expression of F5H mRNA is distinct from that of other phenylpropanoid genes. Efforts to identify constructs capable of complementing the sinapate ester-deficient phenotype of fah1 mutants demonstrated thatF5H expression in leaves is dependent on sequences 3′ of the F5H coding region. In contrast, the positive regulatory function of the downstream region is not required forF5H transcript or sinapoylcholine accumulation in embryos.
Feedback Control and Diurnal Regulation of Gibberellin 20-Oxidase Transcript Levels in Potato1
Authors: Carrera Esther , Jackson Stephen D. , Prat Salomé 
Published: July 2002

Abstract
Tuber formation in potato (Solanum tuberosum) is promoted by short photoperiods and is inhibited by gibberellins (GAs). Endogenous levels of GA1 were shown to decrease in stolons and leaves of potato plants induced to tuberize, which suggests that photoperiodic regulation of GA biosynthesis may play a role in tuber induction. We report the isolation of three potato cDNA clones (StGA20ox1–3) encoding GA 20-oxidase, a key regulatory enzyme in the GA-biosynthetic pathway. Using northern analysis, we detected a differential pattern of tissue-specific expression of the mRNAs corresponding to these clones. StGA20ox mRNAs were also very abundant in leaves of the potatoga1 mutant, which is blocked in the 13-hydroxylation step, and were strongly down-regulated by gibberellic acid, suggesting a feedback regulation of these genes. In plants grown in short-day (inductive) conditions, levels of the StGA20ox transcripts in leaves fluctuated during a 24-h period, with a peak of accumulation observed about 4 h after the lights were turned off. Interruption of the night with a 30-min “night break” of light (noninductive conditions) did not have a marked effect on the levels of accumulation of the three GA 20-oxidase mRNAs during the day, but it induced a second peak of expression of StGA20ox1 and StGA20ox3 transcripts late in the night. This observation, together with the finding that StGA20ox1 mRNA is expressed at high levels in leaves, suggests that night-break induction of this gene might play a role in the control of tuberization by regulating endogenous levels of GAs in response to daylength conditions.
Cyanide-Resistant, ATP-Synthesis-Sustained, and Uncoupling-Protein-Sustained Respiration during Postharvest Ripening of Tomato Fruit1
Authors: Almeida Andrea Miyasaka , Jarmuszkiewicz Wieslawa , Khomsi Hamid , Arruda Paulo , Vercesi Anibal E. , Sluse Francis E. 
Published: July 2002

Abstract
Tomato (Lycopersicon esculentum) mitochondria contain both alternative oxidase (AOX) and uncoupling protein as energy-dissipating systems that can decrease the efficiency of oxidative phosphorylation. We followed the cyanide (CN)-resistant, ATP-synthesis-sustained, and uncoupling-protein-sustained respiration of isolated mitochondria, as well as the immunologically detectable levels of uncoupling protein and AOX, during tomato fruit ripening from the mature green stage to the red stage. The AOX protein level and CN-resistant respiration of isolated mitochondria decreased with ripening from the green to the red stage. The ATP-synthesis-sustained respiration followed the same behavior. In contrast, the level of uncoupling protein and the total uncoupling-protein-sustained respiration of isolated mitochondria decreased from only the yellow stage on. We observed an acute inhibition of the CN-resistant respiration by linoleic acid in the micromolar range. These results suggest that the two energy-dissipating systems could have different roles during the ripening process.
Cloning and Inactivation of Genes Encoding Ferredoxin- and NADH-Dependent Glutamate Synthases in the CyanobacteriumPlectonema boryanum. Imbalances in Nitrogen and Carbon Assimilations Caused by Deficiency of the Ferredoxin-Dependent Enzyme1
Authors: Okuhara Hiroaki , Matsumura Tomohiro , Fujita Yuichi , Hase Toshiharu 
Published: July 2002

Abstract
Glutamate synthase (GOGAT) is a key enzyme in the assimilation of inorganic nitrogen in photosynthetic organisms. We found that, like higher plants, the facultative heterotrophic cyanobacterium Plectonema boryanum had ferredoxin (Fd)- and NADH-dependent GOGATs. The genes glsF, gltB, and gltDwere cloned, and structural analyses and target mutageneses demonstrated that glsF encoded Fd-GOGAT and thatgltB and gltD encoded the two subunits of NADH-GOGAT. All three mutants lacking one of the GOGAT genes were able to grow photosynthetically and heterotrophically. However, the Fd-GOGAT mutant exhibited a phenotype of marked nitrogen deficiency when grown under conditions of saturating illumination and CO2 supply. In these conditions the rate of the ammonia uptake from the culture medium was slower in the Fd-GOGAT mutant than in the wild type or in the NADH-GOGAT mutant, but no significant differences were found in the rate of the CO2 fixation-dependent O2 evolution among these strains. Our results suggest that, although both Fd- and NADH-GOGATs were operative in the cells growing in light, the contribution of Fd-GOGAT, which directly utilizes photoreducing power for the catalytic reaction, is essential for balancing photosynthetic nitrogen and carbon assimilation.
Cold-Induced Freezing Tolerance in Arabidopsis1
Authors: Wanner Leslie A. , Junttila Olavi 
Published: July 2002

Abstract
Changes in the physiology of plant leaves are correlated with enhanced freezing tolerance and include accumulation of compatible solutes, changes in membrane composition and behavior, and altered gene expression. Some of these changes are required for enhanced freezing tolerance, whereas others are merely consequences of low temperature. In this study we demonstrated that a combination of cold and light is required for enhanced freezing tolerance in Arabidopsis leaves, and this combination is associated with the accumulation of soluble sugars and proline. Sugar accumulation was evident within 2 h after a shift to low temperature, which preceded measured changes in freezing tolerance. In contrast, significant freezing tolerance was attained before the accumulation of proline or major changes in the percentage of dry weight were detected. Many mRNAs also rapidly accumulated in response to low temperature. All of the cold-induced mRNAs that we examined accumulated at low temperature even in the absence of light, when there was no enhancement of freezing tolerance. Thus, the accumulation of these mRNAs is insufficient for cold-induced freezing tolerance.