bluuebunny/crossref_metadata_2025_split huggingface dataset:
Proteomic Approach to Identify Novel Mitochondrial Proteins in Arabidopsis
Authors: Kruft Volker , Eubel Holger , Jänsch Lothar , Werhahn Wolf , Braun Hans-Peter 
Published: November 2008

Abstract
An Arabidopsis mitochondrial proteome project was started for a comprehensive investigation of mitochondrial functions in plants. Mitochondria were prepared from Arabidopsis stems and leaves or from Arabidopsis suspension cell cultures, and the purity of the generated fractions was tested by the resolution of organellar protein complexes applying two-dimensional blue-native/N-[2-hydroxy-1,1-bis(hydroxymethyl)ethyl]glycine (Tricine) sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The Arabidopsis mitochondrial proteome was analyzed by two-dimensional isoelectric focusing/ Tricine sodium dodecyl sulfate-polyacrylamide gel electrophoresis and 650 different proteins in a pI range of pH 3 to 10 were separated on single gels. Solubilization conditions, pH gradients for isoelectric focusing, and gel staining procedures were varied, and the number of separable proteins increased to about 800. Fifty-two protein spots were identified by immunoblotting, direct protein sequencing, and mass spectrometry. The characterized proteins cooperate in various processes, such as respiration, citric acid cycle, amino acid and nucleotide metabolism, protection against O2, mitochondrial assembly, molecular transport, and protein biosynthesis. More than 20% of the identified proteins were not described previously for plant mitochondria, indicating novel mitochondrial functions. The map of the Arabidopsis mitochondrial proteome should be useful for the analysis of knockout mutants concerning nuclear-encoded mitochondrial genes. Considerations of the total complexity of the Arabidopsis mitochondrial proteome are discussed. The data from this investigation will be made available athttp://www.gartenbau.uni-hannover.de/genetik/AMPP.
Sodium Uptake in Arabidopsis Roots Is Regulated by Cyclic Nucleotides
Authors: Maathuis Frans J.M. , Sanders Dale 
Published: November 2008

AbstractSodium uptake from the soil is a major cause of salinity toxicity in plants, yet little is known about the mechanisms that underlie Na+ influx. We have characterized voltage independent channels (VICs) in Arabidopsis roots that are thought to contribute to Na+ entry. VICs showed no selectivity among monovalent cations, and their gating was found to be voltage independent. However, VIC open probability showed sensitivity to cyclic nucleotides. The presence of micromolar concentrations of cAMP or cGMP at the cytoplasmic side of the plasma membrane evoked a rapid decrease in channel open probability. In accord with predictions from electrophysiological data, our results show that short-term unidirectional Na+ influx is also reduced in the presence of cyclic nucleotides. Moreover, addition of membrane permeable cyclic nucleotides during growth assays improved plant salinity tolerance, which corresponded with lower levels of Na+ accumulation in plants. In summary, these data imply that Arabidopsis plants may contain a cyclic nucleotide-based signaling pathway that directly affects Na+ transport via VICs.
DEX1, a Novel Plant Protein, Is Required for Exine Pattern Formation during Pollen Development in Arabidopsis
Authors: Paxson-Sowders Dawn M. , Dodrill Craig H. , Owen Heather A. , Makaroff Christopher A. 
Published: November 2008

AbstractTo identify factors that are required for proper pollen wall formation, we have characterized the T-DNA-tagged, dex1mutation of Arabidopsis, which results in defective pollen wall pattern formation. This study reports the isolation and molecular characterization of DEX1 and morphological and ultrastructural analyses of dex1 plants.DEX1 encodes a novel plant protein that is predicted to be membrane associated and contains several potential calcium-binding domains. Pollen wall development in dex1 plants parallels that of wild-type plants until the early tetrad stage. Index1 plants, primexine deposition is delayed and significantly reduced. The normal rippling of the plasma membrane and production of spacers observed in wild-type plants is also absent in the mutant. Sporopollenin is produced and randomly deposited on the plasma membrane in dex1 plants. However, it does not appear to be anchored to the microspore and forms large aggregates on the developing microspore and the locule walls. Based on the structure of DEX1 and the phenotype of dex1 plants, several potential roles for the protein are proposed.
Sodium Fluxes through Nonselective Cation Channels in the Plasma Membrane of Protoplasts from Arabidopsis Roots
Authors: Demidchik Vadim , Tester Mark 
Published: November 2008

AbstractThe aim of the present work was to characterize Na+currents through nonselective cation channels (NSCCs) in protoplasts derived from root cells of Arabidopsis. The procedure of the protoplast isolation was modified to increase the stability of Arabidopsis root protoplasts in low external Ca2+ by digesting tissue in elevated Ca2+. Experiments in whole-cell and outside-out modes were carried out. We found that Na+ currents in Arabidopsis root protoplasts were mediated by cation channels that were insensitive to externally applied tetraethylammonium+ and verapamil, had no time-dependent activation (permanently opened or completely activated within 1–2 ms), were voltage independent, and were weakly selective for monovalent cations. The selectivity sequence was as follows: K+ (1.49) &gt; NH4 + (1.24) &gt; Rb+ (1.15) ≈ Cs+ (1.10) ≈ Na+ (1.00) &gt; Li+ (0.73) &gt; tetraethylammonium+ (0.47). Arabidopsis root NSCCs were blocked by H+ (pK ≈ 6.0), Ca2+(K1/2 ≈ 0.1 mm), Ba2+, Zn2+, La3+, Gd3+, quinine, and the His modifier diethylpyrocarbonate. They were insensitive to most organic blockers (nifedipine, verapamil, flufenamate, and amiloride) and to the SH-group modifier p-chloromercuriphenyl sulfonic acid. Voltage-insensitive, Ca2+-sensitive single channels were also resolved. Properties of Arabidopsis root NSCCs are discussed and compared with characteristics of similar conductances studied previously in plants and animals. It is suggested that NSCCs present a distinct group of plant ion channels, mediating toxic Na+ influx to the cell and probably having other important roles in physiological processes of plants.
Colocalization of Plastid Division Proteins in the Chloroplast Stromal Compartment Establishes a New Functional Relationship between FtsZ1 and FtsZ2 in Higher Plants
Authors: McAndrew Rosemary S. , Froehlich John E. , Vitha Stanislav , Stokes Kevin D. , Osteryoung Katherine W. 
Published: November 2008

Abstract
Chloroplast division is driven by a macromolecular complex containing components that are positioned on the cytosolic surface of the outer envelope, the stromal surface of the inner envelope, and in the intermembrane space. The only constituents of the division apparatus identified thus far are the tubulin-like proteins FtsZ1 and FtsZ2, which colocalize to rings at the plastid division site. However, the precise positioning of these rings relative to the envelope membranes and to each other has not been previously defined. Using newly isolated cDNAs with open reading frames longer than those reported previously, we demonstrate here that both FtsZ2 proteins in Arabidopsis, like FtsZ1 proteins, contain cleavable transit peptides that target them across the outer envelope membrane. To determine their topological arrangement, protease protection experiments designed to distinguish between stromal and intermembrane space localization were performed on both in vitro imported and endogenous forms of FtsZ1 and FtsZ2. Both proteins were shown to reside in the stromal compartment of the chloroplast, indicating that the FtsZ1- and FtsZ2-containing rings have similar topologies and may physically interact. Consistent with this hypothesis, double immunofluorescence labeling of various plastid division mutants revealed precise colocalization of FtsZ1 and FtsZ2, even when their levels and assembly patterns were perturbed. Overexpression of FtsZ2 in transgenic Arabidopsis inhibited plastid division in a dose-dependent manner, suggesting that the stoichiometry between FtsZ1 and FtsZ2 is an important aspect of their function. These studies raise new questions concerning the functional and evolutionary significance of two distinct but colocalized forms of FtsZ in plants and establish a revised framework within which to understand the molecular architecture of the plastid division apparatus in higher plants.
Diurnal and Circadian Regulation of Putative Potassium Channels in a Leaf Moving Organ
Authors: Moshelion Menachem , Becker Dirk , Czempinski Katrin , Mueller-Roeber Bernd , Attali Bernard , Hedrich Rainer , Moran Nava 
Published: November 2008

Abstract
In a search for potassium channels involved in light- and clock-regulated leaf movements, we cloned four putative K channel genes from the leaf-moving organs, pulvini, of the legume Samanea saman. The S. saman SPOCK1 is homologous to KCO1, an Arabidopsis two-pore-domain K channel, the S. saman SPORK1 is similar to SKOR and GORK, Arabidopsis outward-rectifying Shaker-like K channels, and theS. saman SPICK1 and SPICK2 are homologous to AKT2, a weakly-inward-rectifying Shaker-like Arabidopsis K channel. All four S. saman sequences possess the universal K-channel-specific pore signature, TXXTXGYG, strongly suggesting a role in transmembrane K+ transport. The fourS. saman genes had different expression patterns within four leaf parts: ‘extensor’ and ‘flexor’ (the motor tissues), the leaf blades (mainly mesophyll), and the vascular bundle (‘rachis’). Based on northern blot analysis, their transcript level was correlated with the rhythmic leaf movements: (a) all four genes were regulated diurnally (Spick2, Spork1, andSpock1 in extensor and flexor, Spick1 in extensor and rachis); (b) Spork1 andSpock1 rhythms were inverted upon the inversion of the day-night cycle; and (c) in extensor and/or flexor, the expression ofSpork1, Spick1, and Spick2was also under a circadian control. These findings parallel the circadian rhythm shown to govern the resting membrane K+permeability in extensor and flexor protoplasts and the susceptibility of this permeability to light stimulation (Kim et al., 1993). Thus,Samanea pulvinar motor cells are the first described system combining light and circadian regulation of K channels at the level of transcript and membrane transport.
Dynamic and Steady-State Responses of Inorganic Nitrogen Pools and NH3 Exchange in Leaves ofLolium perenneandBromus erectusto Changes in Root Nitrogen Supply
Authors: Mattsson Marie , Schjoerring Jan K. 
Published: November 2008

AbstractShort- and long-term responses of inorganic N pools and plant-atmosphere NH3 exchange to changes in external N supply were investigated in 11-week-old plants of two grass species,Lolium perenne and Bromus erectus, characteristic of N-rich and N-poor grassland ecosystems, respectively. A switch of root N source from NO3 − to NH4 + caused within 3 h a 3- to 6-fold increase in leaf apoplastic NH4 + concentration and a simultaneous decrease in apoplastic pH of about 0.4 pH units in both species. The concentration of total extractable leaf tissue NH4 + also increased two to three times within 3 h after the switch. Removal of exogenous NH4 + caused the apoplastic NH4 + concentration to decline back to the original level within 24 h, whereas the leaf tissue NH4 +concentration decreased more slowly and did not reach the original level in 48 h. After growing for 5 weeks with a steady-state supply of NO3 − or NH4 +, L. perenne were in all cases larger, contained more N, and utilized the absorbed N more efficiently for growth than B. erectus, whereas the two species behaved oppositely with respect to tissue concentrations of NO3 −, NH4 +, and total N. Ammonia compensation points were higher for B. erectus than for L. perenne and were in both species higher for NH4 +- than for NO3 −-grown plants. Steady-state levels of apoplastic NH4 +, tissue NH4 +, and NH3 emission were significantly correlated. It is concluded that leaf apoplastic NH4 + is a highly dynamic pool, closely reflecting changes in the external N supply. This rapid response may constitute a signaling system coordinating leaf N metabolism with the actual N uptake by the roots and the external N availability.
Differential Substrate Inhibition Couples Kinetically Distinct 4-Coumarate:Coenzyme A Ligases with Spatially Distinct Metabolic Roles in Quaking Aspen
Authors: Harding Scott A. , Leshkevich Jacqueline , Chiang Vincent L. , Tsai Chung-Jui 
Published: November 2008

Abstract4-Coumarate:coenzyme A ligase (4CL) activates hydroxycinnamates for entry into phenylpropanoid branchways that support various metabolic activities, including lignification and flavonoid biosynthesis. However, it is not clear whether and how 4CL proteins with their broad substrate specificities fulfill the specific hydroxycinnamate requirements of the branchways they supply. Two tissue-specific 4CLs, Pt4CL1 and Pt4CL2, have previously been cloned from quaking aspen (Populus tremuloides Michx.), but whether they are catalytically adapted for the distinctive metabolic roles they are thought to support is not apparent from published biochemical data. Therefore, single- and mixed-substrate assays were conducted to determine whether the 4CLs from aspen exhibit clear catalytic identities under certain metabolic circumstances. Recombinant Pt4CL1 and Pt4CL2 exhibited the expected preference forp-coumarate in single-substrate assays, but strong competitive inhibition favored utilization of caffeate andp-coumarate, respectively, in mixed-substrate assays. The Pt4CL1 product, caffeoyl-CoA, predominated in mixed-substrate assays with xylem extract, and this was consistent with the near absence of Pt4CL2 expression in xylem tissue as determined by in situ hybridization. It is interesting that the Pt4CL2 product p-coumaroyl-CoA predominated in assays with developing leaf extract, although in situ hybridization revealed that both genes were coexpressed. The xylem extract and recombinant 4CL1 data allow us to advance a mechanism by which 4CL1 can selectively utilize caffeate for the support of monolignol biosynthesis in maturing xylem and phloem fibers. Loblolly pine (Pinus taeda), in contrast, possesses a single 4CL protein exhibiting broad substrate specificity in mixed-substrate assays. We discuss these 4CL differences in terms of the contrasts in lignification between angiosperm trees and their gymnosperm progenitors.
Molecular Characterization of Two Arabidopsis Ire1 Homologs, Endoplasmic Reticulum-Located Transmembrane Protein Kinases
Authors: Koizumi Nozomu , Martinez Immaculada M. , Kimata Yukio , Kohno Kenji , Sano Hiroshi , Chrispeels Maarten J. 
Published: November 2008

AbstractA major response of eukaryotic cells to the presence of unfolded proteins in the lumen of the endoplasmic reticulum (ER) is to activate genes that encode ER-located molecular chaperones, such as the binding protein. This response, called the unfolded protein response, requires the transduction of a signal from the ER to the nucleus. In yeast (Saccharomyces cerevisiae) and mammalian cells, an ER-located transmembrane receptor protein kinase/ribonuclease called Ire1, with a sensor domain in the lumen of the ER, is the first component of this pathway. Here, we report the cloning and derived amino acid sequences of AtIre1-1 and AtIre1-2, two Arabidopsis homologs of Ire1. The two proteins are located in the perinuclear ER (based on heterologous expression of fusions with green fluorescent protein). The expression patterns of the two genes (using β-glucuronidase fusions) are nearly nonoverlapping. We also demonstrate functional complementation of the sensor domains of the two proteins in yeast and show that the Ire1-2 protein is capable of autotransphosphorylation. These and other findings are discussed in relation to the involvement of these genes in unfolded protein response signaling in plants.
Extrinsic Photosystem II Carbonic Anhydrase in Maize Mesophyll Chloroplasts
Authors: Lu Yih-Kuang , Stemler Alan J. 
Published: November 2008

Abstract
One form of carbonic anhydrase (CA) has been observed in maize (Zea mays) thylakoids and photosystem II (PSII)-enriched membranes. Here, we show that an antibody produced against a thylakoid lumen-targeted CA found in Chlamydomonas reinhardtiireacts with a single 33-kD polypeptide in maize thylakoids. With immunoblot analysis, we found that this single polypeptide could be identified only in mesophyll thylakoids and derived PSII membranes, but not in bundle sheath thylakoids. Likewise, a CA activity assay confirmed a large amount of activity in mesophyll, but not in bundle sheath membranes. Immunoblot analysis and CA activity assay showed that the maximum CA can be obtained in the supernatant of the PSII-enriched membranes washed with 1 m CaCl2, the same procedure used to remove all extrinsic lumenal proteins from PSII. Because this CA reacts with an antibody to lumen-directed CA inC. reinhardtii, and because it can be removed with 1m CaCl2 wash, we refer to it tentatively as extrinsic CA. This is to distinguish it from another form of CA activity tightly bound to PSII membranes that remains after CaCl2 wash, which has been described previously. The function of extrinsic CA is not clear. It is unlikely to have the same function as the cytoplasmic CA, which has been proposed to increase the HCO3 − concentration for phosphoenolpyruvate carboxylase and the C4 pathway. We suggest that because the extrinsic CA is associated only with thylakoids doing linear electron flow, it could function to produce the CO2 or HCO3 − needed for PSII activity.
Elemental Sulfur and Thiol Accumulation in Tomato and Defense against a Fungal Vascular Pathogen
Authors: Williams Jane S. , Hall Sharon A. , Hawkesford Malcolm J. , Beale Michael H. , Cooper Richard M. 
Published: November 2008

AbstractThe occurrence of fungicidal, elemental S is well documented in certain specialized prokaryotes, but has rarely been detected in eukaryotes. Elemental S was first identified in this laboratory as a novel phytoalexin in the xylem of resistant genotypes of Theobroma cacao, after infection by the vascular, fungal pathogen Verticillium dahliae. In the current work, this phenomenon is demonstrated in a resistant line of tomato,Lycopersicon esculentum, in response to V. dahliae. A novel gas chromatography-mass spectroscopy method using isotope dilution analysis with 34S internal standard was developed to identify unambiguously and quantify 32S in samples of excised xylem. Accumulation of S in vascular tissue was more rapid and much greater in the disease-resistant than in the disease-susceptible line. Levels of S detected in the resistant variety (approximately 10 μg g−1 fresh weight excised xylem) were fungitoxic to V. dahliae (spore germination was inhibited &gt;90% at approximately 3 μg mL−1). Scanning electron microscopy-energy dispersive x-ray microanalysis confirmed accumulation of S in vascular but not in pith cells and in greater amounts and frequency in the Verticillium spp.-resistant genotype. More intensive localizations of S were occasionally detected in xylem parenchyma cells, vessel walls, vascular gels, and tyloses, structures in potential contact with and linked with defense toV. dahliae. Transient increases in concentrations of sulfate, glutathione, and Cys of vascular tissues from resistant but not susceptible lines after infection may indicate a perturbation of S metabolism induced by elemental S formation; this is discussed in terms of possible S biogenesis.
The N-Terminal Region of Arabidopsis Cystathionine γ-Synthase Plays an Important Regulatory Role in Methionine Metabolism
Authors: Hacham Yael , Avraham Tal , Amir Rachel 
Published: November 2008

Abstract
Cystathionine γ-synthase (CGS) is a key enzyme of Met biosynthesis in bacteria and plants. Aligning the amino acid sequences revealed that the plant enzyme has an extended N-terminal region that is not found in the bacterial enzyme. However, this region is not essential for the catalytic activity of this enzyme, as deduced from the complementation test of an Escherichia coli CGS mutant. To determine the function of this N-terminal region, we overexpressed full-length Arabidopsis CGS and its truncated version that lacks the N-terminal region in transgenic tobacco (Nicotiana tabacum) plants. Transgenic plants expressing both types of CGS had a significant higher level of Met,S-methyl-Met, and Met content in their proteins. However, although plants expressing full-length CGS showed the same phenotype and developmental pattern as wild-type plants, those expressing the truncated CGS showed a severely abnormal phenotype. These abnormal plants also emitted high levels of Met catabolic products, dimethyl sulfide and carbon disulfide. The level of ethylene, the Met-derived hormone, was 40 times higher than in wild-type plants. Since the alien CGS was expressed at comparable levels in both types of transgenic plants, we further suggest that post-translational modification(s) occurs in this N-terminal region, which regulate CGS and/or Met metabolism. More specifically, since the absence of the N-terminal region leads to an impaired Met metabolism, the results further suggest that this region plays a role in protecting plants from a high level of Met catabolic products such as ethylene.
Signal Transduction in Maize and Arabidopsis Mesophyll Protoplasts
Authors: Sheen Jen 
Published: November 2008

AbstractPlant protoplasts show physiological perceptions and responses to hormones, metabolites, environmental cues, and pathogen-derived elicitors, similar to cell-autonomous responses in intact tissues and plants. The development of defined protoplast transient expression systems for high-throughput screening and systematic characterization of gene functions has greatly contributed to elucidating plant signal transduction pathways, in combination with genetic, genomic, and transgenic approaches.