bluuebunny/crossref_metadata_2025_split huggingface dataset:
Gene transfer from mitochondrion to nucleus: novel mechanisms for gene activation from Cox2
Authors: Daley Daniel O. , Adams Keith L. , Clifton Rachel , Qualmann Svenja , Millar A. Harvey , Palmer Jeffrey D. , Pratje Elke , Whelan James 
Published: March 2003

SummaryThe evolutionarily recent transfer of the gene for cytochrome c oxidase subunit 2 (cox2) from the mitochondrion to the nucleus in legumes is shown to have involved novel gene‐activation steps. The acquired mitochondrial targeting presequence is bordered by two introns. Characterization of the import of soybean Cox2 indicates that the presequence is cleaved in a three‐step process which is independent of assembly. The final processing step takes place only in the mitochondria of legume species, and not in several non‐legume plants. The unusually long presequence of 136 amino acids consists of three regions: the first 20 amino acids are required for mitochondrial targeting and can be replaced by another presequence; the central portion of the presequence is required for efficient import of the Cox2 protein into mitochondria; and the last 12 amino acids, derived from the mitochondrially encoded protein, are required for correct maturation of the imported protein. The acquisition of a unique presequence, and the capacity for legume mitochondria to remove this presequence post‐import, are considered to be essential adaptations for targeting of Cox2 to the mitochondrion and therefore activation of the transferred gene in the nucleus.
Changes in secondary metabolism and deposition of an unusual lignin in the ref8 mutant of Arabidopsis
Authors: Franke Rochus , Hemm Matthew R. , Denault Jeff W. , Ruegger Max O. , Humphreys John M. , Chapple Clint 
Published: March 2003

SummaryThe end products of the phenylpropanoid pathway play important roles in plant structure and development, as well as in plant defense mechanisms against biotic and abiotic stresses. From a human perspective, phenylpropanoid pathway‐derived metabolites influence both human health and the potential utility of plants in agricultural contexts. The last known enzyme of the phenylpropanoid pathway that has not been characterized is p‐coumarate 3‐hydroxylase (C3H). By screening for plants that fail to accumulate soluble fluorescent phenylpropanoid secondary metabolites, we have identified a number of Arabidopsis mutants that display a reduced epidermal fluorescence (ref) phenotype. We have now shown that the ref8 mutant is defective in the gene encoding C3H. Phenotypic characterization of the ref8 mutant has revealed that the lack of C3H activity in the mutant leads to diverse changes in phenylpropanoid metabolism. The ref8 mutant accumulates p‐coumarate esters in place of the sinapoylmalate found in wild‐type plants. The mutant also deposits a lignin formed primarily from p‐coumaryl alcohol, a monomer that is at best a minor component in the lignin of other plants. Finally, the mutant displays developmental defects and is subject to fungal attack, suggesting that phenylpropanoid pathway products downstream of REF8 may be required for normal plant development and disease resistance.
The WUSCHEL gene promotes vegetative‐to‐embryonic transition in Arabidopsis
Authors: Zuo Jianru , Niu Qi‐Wen , Frugis Giovanna , Chua Nam‐Hai 
Published: March 2003

SummaryFormation of somatic embryos in plants is known to require high concentrations of auxin or 2,4‐dichlorophenoxyacetic acid (2,4‐D), which presumably acts to trigger a signalling cascade. However, very little is known about the molecular mechanism that mediates the vegetative‐to‐embryogenic transition. We have employed a genetic approach to dissect the signal transduction pathway during somatic embryogenesis. In a functional screen using a chemical‐inducible activation‐tagging system, we identified two alleles of Arabidopsis gene PGA6 whose induced overexpression caused high‐frequency somatic embryo formation in all tissues and organs tested, without any external plant hormones. Upon inducer withdrawal, all these somatic embryos were able to germinate directly, without any further treatment, and to develop into fertile adult plants. PGA6 was found to be identical to WUSCHEL (WUS), a homeodomain protein previously shown to be involved in specifying stem cell fate in shoot and floral meristems. Transgenic plants carrying an estradiol‐inducible XVE‐WUS transgene can phenocopy pga6‐1 and pga6‐2. Our results suggest that WUS/PGA6 also plays a key role during embryogenesis, presumably by promoting the vegetative‐to‐embryogenic transition and/or maintaining the identity of the embryonic stem cells.
DNA binding and dimerization specificity and potential targets for the TCP protein family
Authors: Kosugi Shunichi , Ohashi Yuko 
Published: March 2003

SummaryThe TCP domain is a plant‐specific DNA binding domain found in proteins from a diverse array of species, including the cycloidea (cyc) and teosinte branched1 (tb1) gene products and the PCF1 and PCF2 proteins. To understand the role in transcriptional regulation of proteins with this domain, we have analysed the DNA binding and dimerization specificity of the TCP protein family using rice PCF proteins, and further evaluated potential targets for the TCP protein. The seven PCF members including five newly isolated proteins, were able to be grouped into two classes, I and II, based on sequence similarity in the TCP domain. Random binding site selection experiments and electrophoretic mobility shift assays (EMSAs) revealed the consensus DNA binding sequences of these two classes to be distinct but overlapping; GGNCCCAC for class I and GTGGNCCC for class II. The TB1 protein from maize, which belongs to class II, had the same specificity as the rice class II proteins, suggesting the conservation of binding specificity between TCP domains from different species. The yeast 2‐hybrid assay and EMSA revealed that these proteins tend to form a homodimer or a heterodimer between members of the same class. We searched predicted 5′ flanking sequences of Arabidopsis genes for the consensus binding sequences and found that the consensus sites are distributed in the genome at a considerably lower frequency. We further analysed eight promoters containing the class I consensus TCP sites. The transcriptional activities of six promoters were decreased by a mutation of the TCP binding site, which is consistent with the observation that the class I TCP site can confer transactivation function on a heterologous promoter. These results suggest that the two classes of TCP protein are distinct in DNA binding specificity and transcriptional regulation.
Expression of Arabidopsis SR‐like splicing proteins confers salt tolerance to yeast and transgenic plants
Authors: Forment Javier , Naranjo Miguel Ángel , Roldán Marta , Serrano Ramón , Vicente Oscar 
Published: March 2003

SummarySearching for novel targets of salt toxicity in eukaryotic cells, we have screened an Arabidopsis thaliana cDNA library to isolate genes conferring increased tolerance to salt stress when expressed in the yeast Saccharomyces cerevisiae. Here we show that expression of the ‘alternating arginine‐rich’ (or RS) domains of two different SR‐like, putative splicing proteins from Arabidopsis allows yeast cells to tolerate higher lithium and sodium concentrations. Protection against salt stress appears to require the in vivo phosphorylation of these plant polypeptides, since the yeast SR protein kinase Sky1p, which was able to phosphorylate in vitro at least one of them, also proved to be essential for the observed salt tolerance phenotype. In addition, a clone encoding the U1A protein, a previously characterised Arabidopsis splicing factor, was also isolated in the screening. No significant decrease in the intracellular concentration of lithium was observed in yeast cells incubated in the presence of LiCl upon expression of any of the Arabidopsis proteins, suggesting that their effects are not mediated by the stimulation of ion transport. In support of the general significance of these data, we also show that the expression of the RS domain of one of the SR‐like proteins in transgenic Arabidopsis plants increases their tolerance to LiCl and NaCl. These results point to an important role of pre‐mRNA splicing and SR‐like proteins in the salt tolerance of eukaryotic cells, offering a novel route to improve this important trait in crop plants.
Male gametophyte development in bread wheat (Triticum aestivum L.): molecular, cellular, and biochemical analyses of a sporophytic contribution to pollen wall ontogeny
Authors: Wang Aiming , Xia Qun , Xie Wenshuang , Dumonceaux Tim , Zou Jitao , Datla Raju , Selvaraj Gopalan 
Published: March 2003

SummaryBread wheat (hexaploid AABBDD genome; 16 billion basepairs) is a genetically complex, self‐pollinating plant with bisexual flowers that produce short‐lived pollen. Very little is known about the molecular biology of its gametophyte development despite a longstanding interest in hybrid seeds. We present here a comprehensive characterization of three apparently homeologous genes (TAA1a, TAA1b and TAA1c) and demonstrate their anther‐specific biochemical function. These eight‐exon genes, found at only one copy per haploid complement in this large genome, express specifically within the sporophytic tapetum cells. The presence of TAA1 mRNA and protein was evident only at specific stages of pollen development as the microspore wall thickened during the progression of free microspores into vacuolated‐microspores. This temporal regulation matched the assembly of wall‐impregnated sporopollenin, a phenylpropanoid‐lipid polymer containing very long chain fatty alcohols (VLCFAlc), described in the literature. Our results establish that sporophytic genes contribute to the production of fatty alcohols: Transgenic expression of TAA1 afforded production of long/VLCFAlc in tobacco seeds (18 : 1; 20 : 1; 22 : 1; 24 : 0; 26 : 0) and in Escherichia coli (14 : 0; 16 : 0; 18 : 1), suggesting biochemical versatility of TAA1 with respect to cellular milieu and substrate spectrum. Pollen walls additionally contain fatty alcohols in the form of wax esters and other lipids, and some of these lipids are known to play a role in the highly specific sexual interactions at the pollen–pistil interface. This study provides a handle to study these and to manipulate pollen traits, and, furthermore, to understand the molecular biology of fatty alcohol metabolism in general.
Six active phage‐type RNA polymerase genes in Nicotiana tabacum
Authors: Hedtke Boris , Legen Julia , Weihe Andreas , Herrmann Reinhold G. , Börner Thomas 
Published: March 2003

SummaryIn higher plants, a small nuclear gene family encodes mitochondrial as well as chloroplast RNA polymerases (RNAP) homologous to the bacteriophage T7‐enzyme. The Arabidopsis genome contains three such RpoT genes, while in monocotyledonous plants only two copies have been found. Analysis of Nicotiana tabacum, a natural allotetraploid, identified six different RpoT sequences. The study of the progenitor species of tobacco, N. sylvestris and N. tomentosiformis, uncovered that the sequences represent two orthologous sets each of three RpoT genes (RpoT1, RpoT2 and RpoT3). Interestingly, while the organelles are inherited exclusively from the N. sylvestris maternal parent, all six RpoT genes are expressed in N. tabacum. GFP‐fusions of Nicotiana RpoT1 revealed mitochondrial targeting properties. Constructs containing the amino‐terminus of RpoT2 were imported into mitochondria as well as into plastids. Thus, the dual‐targeting feature, first described for Arabidopsis RpoT;2, appears to be conserved among eudicotyledonous plants. Tobacco RpoT3 is targeted to chloroplasts and the RNA is differentially expressed in plants lacking the plastid‐encoded RNAP. Remarkably, translation of RpoT3 mRNA has to be initiated at a CUG codon to generate a functional plastid transit peptide. Thus, besides AGAMOUS in Arabidopsis, Nicotiana RpoT3 provides a second example for a non‐viral plant mRNA that is exclusively translated from a non‐AUG codon.
The ABC transporter SpTUR2 confers resistance to the antifungal diterpene sclareol
Authors: Van Den Brûle Sybille , Müller Axel , Fleming Andrew J. , Smart Cheryl C. 
Published: March 2003

SummaryPDR5‐like proteins represent one group of the ABC superfamily of transporters. Members of this group are present in plants and, due to the function of PDR5‐related proteins in fungi in the excretion of xenobiotics (including antifungal agents), it has been proposed that they might play a similar role in plants in the response to and detoxification of herbicides and fungicides. However, until now no functional data has been presented showing an altered plant response to any herbicide or fungicide as a result of manipulating the expression of a PDR5‐like gene in plants. In this paper, we show that the plant SpTUR2 PDR5‐like ABC transporter is localised to the plasma membrane and that expression of this protein in Arabidopsis leads to the acquisition of resistance to the diterpenoid antifungal agent sclareol. These data both define a possible endogenous substrate for this transporter and highlight the potential of manipulating plant chemical resistance via modulating the expression of specific PDR5‐like transporters.
Germination and storage reserve mobilization are regulated independently in Arabidopsis
Authors: Pritchard Sarah L. , Charlton Wayne L. , Baker Alison , Graham Ian A. 
Published: March 2003

SummaryThe phytohormone abscisic acid (ABA) inhibits the germination of many seeds, including Arabidopsis, but the mechanism for this is not known. In cereals, ABA inhibits the expression of genes involved in storage reserve mobilization. We have found that in Arabidopsis ABA decreases transcription from the promoters of marker genes for β‐oxidation and the glyoxylate cycle, essential pathways for the conversion of storage lipid (triacylglycerol) into sucrose. Thirty per cent of stored lipid is broken down over 6 days following imbibition of ABA‐treated seed. Sucrose levels in ABA‐treated seeds, rather than decreasing as under normal growth conditions, actually double during the 3 days following imbibition. This sucrose is derived from triacylglycerol as demonstrated by two mutants disrupted in the conversion of triacylglycerol into sucrose, kat2 and icl1, which do not accumulate sucrose in the presence of ABA. We conclude that the ABA block on germination is not a consequence of inhibition of storage lipid mobilization. Two independent programmes appear to operate, one that is blocked by ABA, governing developmental growth resulting in germination; and a second that governs storage lipid mobilization which is largely ABA‐independent.
Does proline accumulation play an active role in stress‐induced growth reduction?
Authors: Maggio Albino , Miyazaki Saori , Veronese Paola , Fujita Tomomichi , Ibeas José I. , Damsz Barbara , Narasimhan Meena L. , Hasegawa Paul M. , Joly Robert J. , Bressan Ray A. 
Published: March 2003

SummaryAn interesting observation, reported for transgenic plants that have been engineered to overproduce osmolytes, is that they often exhibit impaired growth in the absence of stress. As growth reduction and accumulation of osmolytes both typically result from adaptation, we hypothesized that growth reduction may actually result from osmolyte accumulation. To examine this possibility more closely, intracellular proline level was manipulated by expressing mutated derivatives of tomPRO2 (a Δ1‐pyrroline‐5‐carboxylate synthetase, P5CS, from tomato) in Saccharomyces cerevisiae. This was done in the presence and absence of a functional proline oxidase, followed by selection and screening for increased accumulation of proline in the absence of any stress. Here we show, in support of our hypothesis, that the level of proline accumulation and the amount of growth are inversely correlated in cells grown under normal osmotic conditions. In addition, the intracellular concentration of proline also resulted in increases in ploidy level, vacuolation and altered accumulation of several different transcripts related to cell division and gene expression control. Because these cellular modifications are common responses to salt stress in both yeast and plants, we propose that proline and other osmolytes may act as a signaling/regulatory molecule able to activate multiple responses that are part of the adaptation process. As in previous studies with transgenic plants that overaccumulate osmolytes, we observed some increase in relative growth of proline‐overaccumulating cells in mild hyperosmotic stress.
Virus‐induced gene silencing in tomato
Authors: Liu Yule , Schiff Michael , Dinesh‐Kumar S. P. 
Published: March 2003

AbstractWe have previously demonstrated that a tobacco rattle virus (TRV)‐based vector can be used in virus‐induced gene silencing (VIGS) to study gene function in Nicotiana benthamiana. Here we show that recombinant TRV infects tomato plants and induces efficient gene silencing. Using this system, we suppressed the PDS, CTR1 and CTR2 genes in tomato. Suppression of CTR1 led to a constitutive ethylene response phenotype and up‐regulation of an ethylene response gene, CHITINASE B. This phenotype is similar to Arabidopsis ctr1 mutant plants. We have constructed a modified TRV vector based on the GATEWAY recombination system, allowing restriction‐ and ligation‐free cloning. Our results show that tomato expressed sequence tags (ESTs) can easily be cloned into this modified vector using a single set of primers. Using this vector, we have silenced RbcS and an endogenous gene homologous to the tomato EST cLED3L14. In the future, this modified vector system will facilitate large‐scale functional analysis of tomato ESTs.
Functional characterization of a heavy metal binding protein CdI19 from Arabidopsis
Authors: Suzuki Nobuaki , Yamaguchi Yube , Koizumi Nozomu , Sano Hiroshi 
Published: March 2003

SummaryHeavy metals are potentially highly toxic for organisms. Plants possess the ability to minimize damage but the underlying molecular mechanisms have yet to be detailed. Screening Cd‐responsive genes in Arabidopsis, we previously identified a gene encoding a putative metal binding protein CdI19, which, upon introduction into yeast cells, conferred marked toleration of Cd exposure. Here we describe that bacterially expressed CdI19 directly interacts with Cd at its CXXC motif, as revealed by circular dichroism analysis, and that it is exclusively localized at plasma membranes, as revealed by heterologous expression of fusion product with a green fluorescent protein in BY2 cells. Northern blot analyses indicated that CdI19 transcripts were induced not only by Cd, but also by dicationic forms of Hg, Fe and Cu. Histochemical assays using transgenic Arabidopsis expressing the CdI19 promoter::GUS showed CdI19 to be expressed in petiole, hypocotyl, peduncle and vascular bundles in root tissues. Overexpression of the CdI19 cDNA conferred Cd tolerance in transgenic Arabidopsis. These results suggest that CdI19 plays an important role in the maintenance of heavy metal homeostasis and/or in detoxification by endowing plasma membranes with the capacity to serve as an initial barrier against inflow of free heavy metal ions into cells.
Distribution of 1000 sequenced T‐DNA tags in the Arabidopsis genome
Authors: Szabados László , Kovács Izabella , Oberschall Attila , Ábrahám Edit , Kerekes Irén , Zsigmond Laura , Nagy Réka , Alvarado Martha , Krasovskaja Inga , Gál Mónika , Berente Anikó , Rédei George P. , Haim Amit Ben , Koncz Csaba 
Published: March 2003

SummaryInduction of knockout mutations by T‐DNA insertion mutagenesis is widely used in studies of plant gene functions. To assess the efficiency of this genetic approach, we have sequenced PCR amplified junctions of 1000 T‐DNA insertions and analysed their distribution in the Arabidopsis genome. Map positions of 973 tags could be determined unequivocally, indicating that the majority of T‐DNA insertions landed in chromosomal domains of high gene density. Only 4.7% of insertions were found in interspersed, centromeric, telomeric and rDNA repeats, whereas 0.6% of sequenced tags identified chromosomally integrated segments of organellar DNAs. 35.4% of T‐DNAs were localized in intervals flanked by ATG and stop codons of predicted genes, showing a distribution of 62.2% in exons and 37.8% in introns. The frequency of T‐DNA tags in coding and intergenic regions showed a good correlation with the predicted size distribution of these sequences in the genome. However, the frequency of T‐DNA insertions in 3′‐ and 5′‐regulatory regions of genes, corresponding to 300 bp intervals 3′ downstream of stop and 5′ upstream of ATG codons, was 1.7–2.3‐fold higher than in any similar interval elsewhere in the genome. The additive frequency of insertions in 5′‐regulatory regions and coding domains provided an estimate for the mutation rate, suggesting that 47.8% of mapped T‐DNA tags induced knockout mutations in Arabidopsis.
NPP1, a Phytophthora‐associated trigger of plant defense in parsley and Arabidopsis
Authors: Fellbrich Guido , Romanski Annette , Varet Anne , Blume Beatrix , Brunner Frédéric , Engelhardt Stefan , Felix Georg , Kemmerling Birgit , Krzymowska Magdalena , Nürnberger Thorsten 
Published: March 2003

SummaryActivation of non‐cultivar‐specific plant defense against attempted microbial infection is mediated through the recognition of pathogen‐derived elicitors. Previously, we have identified a peptide fragment (Pep‐13) within a 42‐kDa cell wall transglutaminase from various Phytophthora species that triggers a multifacetted defense response in parsley cells. Many of these oomycete species have now been shown to possess another cell wall protein (24 kDa), that evoked the same pattern of responses in parsley as Pep‐13. Unlike Pep‐13, necrosis‐inducing Phytophthora protein 1 (NPP1) purified from P. parasitica also induced hypersensitive cell death‐like lesions in parsley. NPP1 structural homologs were found in oomycetes, fungi, and bacteria, but not in plants. Structure–activity relationship studies revealed the intact protein as well as two cysteine residues to be essential for elicitor activity. NPP1‐mediated activation of pathogen defense in parsley does not employ the Pep‐13 receptor. However, early induced cellular responses implicated in elicitor signal transmission (increased levels of cytoplasmic calcium, production of reactive oxygen species, MAP kinase activation) were stimulated by either elicitor, suggesting the existence of converging signaling pathways in parsley. Infiltration of NPP1 into leaves of Arabidopsis thaliana Col‐0 plants resulted in transcript accumulation of pathogenesis‐related (PR) genes, production of ROS and ethylene, callose apposition, and HR‐like cell death. NPP1‐mediated induction of the PR1 gene is salicylic acid‐dependent, and, unlike the P. syringae pv. tomato DC3000(avrRpm1)‐induced PR1 gene expression, requires both functional NDR1 and PAD4. In summary, Arabidopsis plants infiltrated with NPP1 constitute an experimental system that is amenable to forward genetic approaches aiming at the dissection of signaling pathways implicated in the activation of non‐cultivar‐specific plant defense.