bluuebunny/crossref_metadata_2025 huggingface dataset:
Econometric analysis of forest conservation: the Finnish experience
Authors: Linden Mikael , Uusivuori Jussi
Published: July 2002

This paper examines the dynamic effects of forest conservation measures. Both short-term and long-term impacts on timber markets are studied using a Finnish example. Forest conservation is interpreted as an exogenous negative shock in the standing timber growth owned by private forest owners, and its effects are retrieved through an impulse response analysis based on a dynamic demand–supply and timber stock model. The results show that timber markets are influenced by forest conservation. The conservation of forest assets reduces traded quantities and increases timber prices. These effects are of the same relative magnitude, thus leaving the annual timber-selling income of private forest owners unchanged. The results suggest that some of the negative short-run impacts of conservation on commercial timber stock will be compensated in the long run by intensified use of the remaining stock. This may to some extent reduce the ecological usefulness of the conservation policies. While the timber markets converge in the long run towards a new post-conservation state the short-term interruptions suggest that caution should be used in establishing set-asides in privately owned forestland.
Poverty, market imperfections and time preferences: of relevance for environmental policy?
Authors: HOLDEN STEIN T. , SHIFERAW BEKELE , WIK METTE
Published: July 2002

Rates of time preference (RTPs) of rural households in Indonesia, Zambia and Ethiopia have been measured using hypothetical questions about preferences for current versus future consumption. In general, the rates were found to be very high. Factors influencing or correlated with the personal rates of time preference were investigated through regression methods. OLS was the technique used in the estimation. Market imperfections, particularly in credit and insurance markets lead to variation in RTPs. Poverty in assets, or cash liquidity constraints, was leading to or correlated with higher rates of time preference. The poor are, therefore, less likely to invest in environmental conservation. In Zambia, independent estimates of risk preferences were made. More risk-averse people tended to have lower RTPs. The results support the hypothesis that poverty and/or liquidity scarcity lead to high RTPs. Poverty reduction may thus reduce the RTPs of the poor and reduce the 'intertemporal externality' due to high RTPs. The high average RTPs indicate, however, that complementary policies may be needed to ensure sufficient levels of investment in conservation. Another logical implication is that institutionalization of private property rights may not be a sufficient tool to initiate sustainable resource management.
Progress on the environmental Kuznets curve?
Authors: STERN DAVID I.
Published: July 2002

The environmental Kuznets curve (EKC) hypothesis proposes that there is an inverted U-shape relation between environmental degradation and income per capita. This has been taken to imply that economic growth will eventually redress the environmental impacts of the early stages of economic development. The literature on this issue has developed rapidly over the last few years. This paper examines whether progress has been made on both understanding the EKC phenomenon and on addressing the various criticisms raised against some of the empirical studies and their interpretation in the policy literature. Though basic EKC studies continue to be carried out, recent work has focused on the effect of a variety of conditioning variables on the environmental impact-GDP relationship. Some attempts have also been made to examine the history of the relationship in individual countries. The econometric techniques used have improved. However, empirical decompositions of the EKC into proximate or underlying causes are either limited in scope or non-systematic, and explicit testing of the various theoretical models has not yet been attempted.
Cognitive Behaviour Therapy
for Post Traumatic Stress Disorder
Authors: Smith Patrick , Perrin Sean , Yule William
Published: July 2002

It is only relatively recently that Post Traumatic Stress Disorder (PTSD) has
been recognised in children. Controlled treatment outcome studies of
childhood PTSD are scarce, but those that exist indicate that Cognitive
Behaviour Therapy (CBT) is an effective intervention. In this article, we
briefly describe PTSD in children and outline some behavioural and
cognitive models of the disorder. Derived from these models, prolonged
therapeutic exposure and cognitive restructuring as part of a CBT package
are then described. In practice, effective therapy will include more than
exposure-based work, and additional procedures, including work with
parents, are highlighted. While CBT is the treatment of choice of PTSD in
childhood, there is an urgent need for further treatment outcome studies.
On deriving complex polysemy: the grammaticalization of get
Authors: Gronemeyer Claire
Published: July 2002

This paper examines the polysemy in the English verb get, which can denote possession, movement, causation, obligation, and change of state among other senses. The analysis builds on a decomposition of get (based on the Benveniste/Freeze/Kayne analysis of possessive constructions) into [ingressive + ‘be’ + preposition]; this lexical entry allows the current polysemy to be derived from a number of reanalyses within different syntactic contexts. Using diachronic data, I show that possession leads to movement as well as stative uses (possession and obligation), movement develops into the causative and inchoative, from which the passive develops, and the infinitival causative gives rise to permission and ingressive aspect. The appearance of each new meaning-construction is motivated by context-dependent mechanisms of reanalysis which account for language change as the result of the language learner reanalyzing the correspondences between syntactic and semantic elements.
Morphometric and molecular analysis of the Encarsia inaron species-group (Hymenoptera: Aphelinidae), parasitoids of whiteflies (Hemiptera: Aleyrodidae)
Authors: Manzari S. , Polaszek A. , Belshaw R. , Quicke D.L.J.
Published: July 2002

Several series of host-reared specimens of an Encarsia species, initially thought to be the cosmopolitan Encarsia inaron (Walker), were collected in the Azores Islands (Portugal). Subsequent morphometric analysis supported the presence of two species: E. inaron and a new species, described herein as Encarsia estrellae Manzari & Polaszek sp. n.Encarsia estrellae was reared from Aleyrodes singularis Danzig, A. ?singularis, and Bemisia sp. afer-group on several host plants. In addition, the D2 region of the 28S rDNA gene was sequenced in eight individuals belonging to these species, as well as single representatives of two closely related and one distantly related species. Phylogenetic analysis of these DNA sequences, together with 23 additional Encarsia sequences retrieved from the European Molecular Biology Laboratory (EMBL) and GenBank databases, further supported the specific status of E. estrellae, and the placement of E. dichroa (Mercet) in the E. inaron species-group. Additionally, E. inaron is redescribed and some taxonomic problems in the E. inaron species-group are discussed.
Pre-mRNA splicing in the absence of an SR protein RS domain
Authors: Zhu Jun , Krainer Adrian R.
Published: July 2002

SR proteins are essential pre-mRNA splicing factors that act at the earliest stages of splice-site recognition and spliceosome assembly, as well as later in the splicing pathway. SR proteins consist of one or two RNA-recognition motifs and a characteristic arginine/serine-rich C-terminal RS domain. The RS domain, which is extensively phosphorylated, mediates the subcellular localization of individual SR proteins and also functions as a splicing activation module, apparently by engaging in protein–protein interactions. The RS domain of SF2/ASF is dispensable for the concentration-dependent effects of this SR protein on alternative splice-site selection. However, this RS domain is highly conserved phylogenetically, and was shown to be required for constitutive splicing in vitro and for cell viability. Here, we demonstrate that the RS domain of SF2/ASF is, in fact, dispensable for splicing of several substrates, including constitutive and enhancer-dependent pre-mRNAs. The requirement for this RS domain is substrate specific, and correlates with the strength of the splicing signals. When the 3′ splice site is weak, both the SF2/ASF RS domain and U2AF35are required for splicing. These results show the existence of an RS domain-independent function of SR proteins in constitutive and enhancer-dependent splicing, and suggest mechanisms for their role in enhancer function besides U2AF recruitment.
Dynamic association of capping enzymes with transcribing RNA polymerase II
Authors: Schroeder Stephanie C. , Schwer Beate , Shuman Stewart , Bentley David
Published: July 2002

The C-terminal heptad repeat domain (CTD) of RNA polymerase II (pol II) is proposed to target pre-mRNA processing enzymes to nascent pol II transcripts, but this idea has not been directly tested in vivo. In vitro, the yeast mRNA capping enzymes Ceg1 and Abd1 bind specifically to the phosphorylated CTD. Here we show that yeast capping enzymes cross-link in vivo to the 5′ ends of transcribed genes and that this localization requires the CTD. Both the extent of CTD phosphorylation at Ser 5 of the heptad repeat and the binding of capping enzymes decreased as polymerase moved from the 5′ to the 3′ ends of the ACT1, ENO2, TEF1, GAL1, and GAL10 genes. Ceg1 is released early in elongation, but Abd1 can travel with transcribing pol II as far as the 3′ end of a gene. The CTD kinase, Kin28, is required for binding, and the CTD phosphatase, Fcp1, is required for dissociation of capping enzymes from the elongation complex. CTD phosphorylation and dephosphorylation therefore control the association of capping enzymes with pol II as it transcribes a gene.
FIERY1 encoding an inositol polyphosphate 1-phosphatase is a negative regulator of abscisic acid and stress signaling in Arabidopsis
Authors: Xiong Liming , Lee Byeong-ha , Ishitani Manabu , Lee Hojoung , Zhang Changqing , Zhu Jian-Kang
Published: July 2002

The plant hormone abscisic acid (ABA) plays a wide range of important roles in plant growth and development, including embryogenesis, seed dormancy, root and shoot growth, transpiration, and stress tolerance. ABA and various abiotic stresses also activate the expression of numerous plant genes through undefined signaling pathways. To gain insight into ABA and stress signal transduction, we conducted a genetic screen based on ABA- and stress-inducible gene transcription. Here we report the identification of anArabidopsis mutation, fiery1 (fry1), which results in super-induction of ABA- and stress-responsive genes. Seed germination and postembryonic development of fry1 are more sensitive to ABA or stress inhibition. The mutant plants are also compromised in tolerance to freezing, drought, and salt stresses. Map-based cloning revealed that FRY1 encodes an inositol polyphosphate 1-phosphatase, which functions in the catabolism of inositol 1, 4, 5-trisphosphate (IP3). Upon ABA treatment,fry1 mutant plants accumulated more IP3 than did the wild-type plants. These results provide the first genetic evidence indicating that phosphoinositols mediate ABA and stress signal transduction in plants and their turnover is critical for attenuating ABA and stress signaling.
Genomic Organization of the Dog Dystroglycan Gene DAG1 Locus on Chromosome 20q15.1–q15.2
Authors: Leeb Tosso , Neumann Stephan , Deppe Alexandra , Breen Matthew , Brenig Bertram
Published: July 2002

Dystroglycan is a laminin binding protein, which provides a structural link between the subsarcolemmal cytoskeleton and the extracellular matrix. It is also involved in the organization of basement membranes. So far the genomic organization of the dystroglycan gene DAG1 has not been completely investigated. Here we report the cloning and sequencing of 162 kb of dog genomic DNA containing the complete ∼71-kb canine DAG1 gene, which consists of three exons, with the translation start codon located in exon 2. Its 2679-nucleotide ORF encodes a polypeptide of 892 amino acids, which is highly similar to human, rabbit, and bovine orthologs. To further characterize the dog DAG1 gene we determined the transcription start site and several naturally occurring polymorphisms, which partially result in amino acid substitutions of the dystroglycan protein. The dog DAG1 gene was assigned to chromosome 20q15.1–q15.2 by FISH analysis. The analysis of the entire reported sequence revealed that the genes for aminomethyltransferase (AMT), bassoon (BSN), TCTA(T-cell leukemiatranslocation-associated) gene, and an as yet uncharacterized protein are located very close to the DAG1gene. Therefore, this study defines a novel syntenic region among dog chromosome 20q15, human chromosome 3p21, and murine chromosome 9F.

[The sequence data described in this paper have been submitted to the EMBL nucleotide database under accession no. AJ012166.]
A High-throughput AFLP-based Method for Constructing Integrated Genetic and Physical Maps: Progress Toward a Sorghum Genome Map
Authors: Klein Patricia E. , Klein Robert R. , Cartinhour Samuel W. , Ulanch Paul E. , Dong Jianmin , Obert Jacque A. , Morishige Daryl T. , Schlueter Shannon D. , Childs Kevin L. , Ale Melissa , Mullet John E.
Published: July 2002

Sorghum is an important target for plant genomic mapping because of its adaptation to harsh environments, diverse germplasm collection, and value for comparing the genomes of grass species such as corn and rice. The construction of an integrated genetic and physical map of the sorghum genome (750 Mbp) is a primary goal of our sorghum genome project. To help accomplish this task, we have developed a new high-throughput PCR-based method for building BAC contigs and locating BAC clones on the sorghum genetic map. This task involved pooling 24,576 sorghum BAC clones (∼4× genome equivalents) in six different matrices to create 184 pools of BAC DNA. DNA fragments from each pool were amplified using amplified fragment length polymorphism (AFLP) technology, resolved on a LI-COR dual-dye DNA sequencing system, and analyzed using Bionumerics software. On average, each set of AFLP primers amplified 28 single-copy DNA markers that were useful for identifying overlapping BAC clones. Data from 32 different AFLP primer combinations identified ∼2400 BACs and ordered ∼700 BAC contigs. Analysis of a sorghum RIL mapping population using the same primer pairs located ∼200 of the BAC contigs on the sorghum genetic map. Restriction endonuclease fingerprinting of the entire collection of sorghum BAC clones was applied to test and extend the contigs constructed using this PCR-based methodology. Analysis of the fingerprint data allowed for the identification of 3366 contigs each containing an average of 5 BACs. BACs in ∼65% of the contigs aligned by AFLP analysis had sufficient overlap to be confirmed by DNA fingerprint analysis. In addition, 30% of the overlapping BACs aligned by AFLP analysis provided information for merging contigs and singletons that could not be joined using fingerprint data alone. Thus, the combination of fingerprinting and AFLP-based contig assembly and mapping provides a reliable, high-throughput method for building an integrated genetic and physical map of the sorghum genome.

[The sequence data described in this paper have been submitted to the GenBank data library under accession no. AF218263.]
Evaluation of Gene-Finding Programs on Mammalian Sequences
Authors: Rogic Sanja , Mackworth Alan K. , Ouellette Francis B.F.
Published: July 2002

We present an independent comparative analysis of seven recently developed gene-finding programs: FGENES,GeneMark.hmm, Genie, Genscan,HMMgene, Morgan, and MZEF. For evaluation purposes we developed a new, thoroughly filtered, and biologically validated dataset of mammalian genomic sequences that does not overlap with the training sets of the programs analyzed. Our analysis shows that the new generation of programs has substantially better results than the programs analyzed in previous studies. The accuracy of the programs was also examined as a function of various sequence and prediction features, such as G + C content of the sequence, length and type of exons, signal type, and score of the exon prediction. This approach pinpoints the strengths and weaknesses of each individual program as well as those of computational gene-finding in general. The dataset used in this analysis (HMR195) as well as the tables with the complete results are available athttp://www.cs.ubc.ca/∼rogic/evaluation/.
Molecular Genetic Maps in Wild Emmer Wheat, Triticum dicoccoides: Genome-Wide Coverage, Massive Negative Interference, and Putative Quasi-Linkage
Authors: Peng Junhua , Korol Abraham B. , Fahima Tzion , Röder Marion S. , Ronin Yefim I. , Li Youchun C. , Nevo Eviatar
Published: July 2002

The main objectives of the study reported here were to construct a molecular map of wild emmer wheat, Triticum dicoccoides, to characterize the marker-related anatomy of the genome, and to evaluate segregation and recombination patterns upon crossing T. dicoccoides with its domesticated descendant Triticum durum(cultivar Langdon). The total map length exceeded 3000 cM and possibly covered the entire tetraploid genome (AABB). Clusters of molecular markers were observed on most of the 14 chromosomes. AFLP (amplified fragment length polymorphism) markers manifested a random distribution among homologous groups, but not among genomes and chromosomes. Genetic differentiation between T. dicoccoidesand T. durum was attributed mainly to the B genome as revealed by AFLP markers. The segregation-distorted markers were mainly clustered on 4A, 5A, and 5B chromosomes. Homeoalleles, differentially conferring the vigor of gametes, might be responsible for the distortion on 5A and 5B chromosomes. Quasilinkage, deviation from free recombination between markers of nonhomologous chromosomes, was discovered. Massive negative interference was observed in most of the chromosomes (an excess of double crossovers in adjacent intervals relative to the expected rates on the assumption of no interference). The general pattern of distribution of islands of negative interference included near-centromeric location, spanning the centromere, and median/subterminal location.

[An appendix describing the molecular marker loci is available as an online supplement athttp://www.genome.org.]
Gene Duplication and the Structure of Eukaryotic Genomes
Authors: Friedman Robert , Hughes Austin L.
Published: July 2002

A simple method for understanding how gene duplication has contributed to genomic structure was applied to the complete genomes ofCaenorhabditis elegans, Drosophila melanogaster, and yeast Saccharomyces cerevisiae. By this method, the genes belonging to gene families (the paranome) were identified, and the extent of sharing of two or more families between genomic windows was compared with that expected under a null model. The results showed significant evidence of duplication of genomic blocks in both C. elegans and yeast. In C. elegans, the five block duplications identified all occurred intra-chromosomally, and all but one occurred quite recently. In yeast, by contrast, 39 duplicated blocks were identified, and all but one of these was inter-chromosomal. Of these 39 blocks, 28 showed evidence of ancient duplication, possibly as a result of an ancient polyploidization event. By contrast, three blocks showed evidence of very recent duplication, while seven others showed a mixture of ancient and recent duplication events. Thus, duplication of genomic blocks has been an ongoing feature of yeast evolution over the past 200–300 million years.
Biased Distribution of Inverted and Direct Alus in the Human Genome: Implications for Insertion, Exclusion, and Genome Stability
Authors: Stenger Judith E. , Lobachev Kirill S. , Gordenin Dmitry , Darden Thomas A. , Jurka Jerzy , Resnick Michael A.
Published: July 2002

Alu sequences, the most abundant class of large dispersed DNA repeats in human chromosomes, contribute to human genome dynamics. Recently we reported that long inverted repeats, including humanAlus, can be strong initiators of genetic change in yeast. We proposed that the potential for interactions between adjacent, closely related Alus would influence their stability and this would be reflected in their distribution. We have undertaken an extensive computational analysis of all Alus (the database is athttp://dir.niehs.nih.gov/ALU) to better understand their distribution and circumstances under which Alu sequences might affect genome stability. Alus separated by <650 bp were categorized according to orientation, length of regions sharing high sequence identity, distance between highly identical regions, and extent of sequence identity. Nearly 50% of all Alu pairs have long alignable regions (>275 bp), corresponding to nearly full-length Alus, regardless of orientation. There are dramatic differences in the distributions and character of Alupairs with closely spaced, nearly identical regions. For Alupairs that are directly repetitive, ∼30% have highly identical regions separated by <20 bp, but only when the alignments correspond to near full-size or half-size Alus. The opposite is found for the distribution of inverted repeats: Alu pairs with aligned regions separated by <20 bp are rare. Furthermore, closely spaced direct and inverted Alus differ in their truncation patterns, suggesting differences in the mechanisms of insertion. At larger distances, the direct and inverted Alu pairs have similar distributions. We propose that sequence identity, orientation, and distance are important factors determining insertion of adjacentAlus, the frequency and spectrum of Alu-associated changes in the genome, and the contribution of Alu pairs to genome instability. Based on results in model systems and the present analysis, closely spaced inverted Alu pairs with long regions of alignment are likely at-risk motifs (ARMs) for genome instability.
A Dominant Modifier of Transgene Methylation Is Mapped by QTL Analysis to Mouse Chromosome 13
Authors: Valenza-Schaerly Pascale , Pickard Benjamin , Walter Jörn , Jung Martin , Pourcel Lucille , Reik Wolf , Gauguier Dominique , Vergnaud Gilles , Pourcel Christine
Published: July 2002

The single-copy hepatitis B virus transgene in the E36 transgenic mouse strain undergoes methylation changes in a parent-of-origin, tissue, and strain-specific fashion. In a C57BL/6 background, the paternally transmitted transgene is methylated in 30% of cells, whereas it is methylated in more than 80% of cells in (BALB/c x C57BL/6) F1 mice. We established previously that several genetic factors were likely to contribute to the transgene methylation profile, some with demethylating and some with de novo methylating activities. Using quantitative trait loci (QTL) mapping, we have now localized one major modifier locus on chromosome 13 (Mod13), which explains a 30% increase in the methylation level of this transgene with no effect on the flanking endogenous sequences. No other QTL could be identified, except for a demethylating activity of low significance located on chromosome 12. Recombinant inbred mice containing a BALB/c allele of Mod13 were then used to show that the presence of Mod13 is sufficient to induce de novo methylation. A segregation between de novo methylation and repression of transgene expression was uncovered, suggesting that this genetic system is also useful for the identification of factors that interpret methylation patterns in the genome.
Regulation of the α-Galactosidase Activity in Streptococcus pneumoniae: Characterization of the Raffinose Utilization System
Authors: Rosenow Carsten , Maniar Mita , Trias Joaquim
Published: July 2002

A 10.2-kb gene region was identified in the Streptococcus pneumoniae genome sequence that contains eight genes involved in regulation and metabolism of raffinose. The genes rafR andrafS are transcribed as one operon, and their gene products regulate the raffinose-dependent stimulation of a divergently transcribed second promoter (PA) directing the expression of aga, the structural gene for α-galactosidase. Raffinose-mediated transcription from PA results in a 500-fold increase in α-galactosidase activity in the cell. A third promoter within the cluster is responsible for the transcription of the remaining five genes (rafE, rafF, rafG,gtfA, and rafX), whose gene products might be involved in transport and metabolism of raffinose. The presence of additional internal promoters cannot be excluded. The agapromoter PA is negatively regulated by the presence of sucrose in the growth medium. Consistent with catabolite repression (CR), a DNA sequence with high homology to the CRE (cis-active element) was identified upstream of the aga promoter. Sucrose-mediated CR depends on the phosphoenolpyruvate: sucrose phosphotransferase system (PTS) but is unaffected by a mutation in a gene encoding a homolog of the CRE regulatory protein CcpA.
Brain Gene Expression During REM Sleep Depends on Prior Waking Experience
Authors: Ribeiro Sidarta , Goyal Vikas , Mello Claudio V. , Pavlides Constantine
Published: July 2002

In most mammalian species studied, two distinct and successive phases of sleep, slow wave (SW), and rapid eye movement (REM), can be recognized on the basis of their EEG profiles and associated behaviors. Both phases have been implicated in the offline sensorimotor processing of daytime events, but the molecular mechanisms remain elusive. We studied brain expression of the plasticity-associated immediate-early gene (IEG) zif-268 during SW and REM sleep in rats exposed to rich sensorimotor experience in the preceding waking period. Whereas nonexposed controls show generalized zif-268 down-regulation during SW and REM sleep, zif-268 is upregulated during REM sleep in the cerebral cortex and the hippocampus of exposed animals. We suggest that this phenomenon represents a window of increased neuronal plasticity during REM sleep that follows enriched waking experience.
Reversible Heat-Induced Inactivation of Chimeric β-Glucuronidase in Transgenic Plants
Authors: Almoguera Concepción , Rojas Anabel , Jordano Juan
Published: July 2002

We compared the expression patterns in transgenic tobacco (Nicotiana tabacum) of two chimeric genes: a translational fusion to β-glucuronidase (GUS) and a transcriptional fusion, both with the same promoter and 5′-flanking sequences ofHa hsp17.7 G4, a small heat shock protein (sHSP) gene from sunflower (Helianthus annuus). We found that immediately after heat shock, the induced expression from the two fusions in seedlings was similar, considering chimeric mRNA or GUS protein accumulation. Surprisingly, we discovered that the chimeric GUS protein encoded by the translational fusion was mostly inactive in such conditions. We also found that this inactivation was fully reversible. Thus, after returning to control temperature, the GUS activity was fully recovered without substantial changes in GUS protein accumulation. In contrast, we did not find differences in the in vitro heat inactivation of the respective GUS proteins. Insolubilization of the chimeric GUS protein correlated with its inactivation, as indicated by immunoprecipitation analyses. The inclusion in another chimeric gene of the 21 amino-terminal amino acids from a different sHSP lead to a comparable reversible inactivation. That effect not only illustrates unexpected post-translational problems, but may also point to sequences involved in interactions specific to sHSPs and in vivo heat stress conditions.
Constitutive Expression of the β-Ketothiolase Gene in Transgenic Plants. A Major Obstacle for Obtaining Polyhydroxybutyrate-Producing Plants
Authors: Bohmert Karen , Balbo Ilse , Steinbüchel Alexander , Tischendorf Gilbert , Willmitzer Lothar
Published: July 2002

Polyhydroxybutyrate (PHB) is a member of a class of thermoelastic polymers called polyhydroxyalkanoates that serve many bacteria as intracellular storage molecules for carbon and energy. Transgenic plants provide a potential means of producing this polymer cost-effectively. To date, however, few reports of the successful production of this polymer have been published, with the exception of work with transgenic Arabidopsis. Using a variety of chimeric constructs, we have determined that the constitutive, chloroplast-localized expression of one of the genes involved in PHB production—the β-ketothiolase (phbA) gene—is detrimental to the efficient production of transgenic PHB. The alternate use of either inducible or somatically activated promoters allowed the construction of transgenic PHB-producing potato (Solanum tuberosum) and tobacco (Nicotiana tabacum) plants, although the amount of PHB formed was still rather low. Taking advantage of an inducible promoter, the maximal amount of PHB produced in transgenic potato was 0.09 mg g−1 dry weight. In transgenic tobacco using a somatically activated promoter, up to 3.2 mg g−1 dry weight was accumulated. In Arabidopsis, the formation of high levels of PHB had previously been shown to be accompanied by severe negative effects on growth and development of the plant. Phasins are proteins known from PHB-producing bacteria speculated to serve as protectants against the highly hydrophobic surface of the PHB granules in the bacterial intracellular milieu. Co-expression of the phasin gene in parallel with the PHB synthesis genes, however, did not lead to reduced symptom development.
Effects of COR6.6 and COR15am Polypeptides Encoded by COR (Cold-Regulated) Genes of Arabidopsis thaliana on Dehydration-Induced Phase Transitions of Phospholipid Membranes
Authors: Webb M. S. , Gilmour S. J. , Thomashow M. F. , Steponkus P. L.
Published: July 2002

Cold acclimation of Arabidopsis thaliana includes the expression of cold-regulated (COR) genes and the accumulation of COR polypeptides. The hydration characteristics of two COR polypeptides, COR6.6 and COR15am, have been determined and their effects on the dehydration-induced liquid crystalline-to-gel and lamellar-to-hexagonal II phase transitions in phospholipid mixtures have been examined. After dehydration at osmotic pressures between 8 and 150 MPa, the water content of the COR polypeptides was less than that of bovine serum albumin, with COR15am the least hydrated: bovine serum albumin > COR6.6 > COR15am. Neither COR6.6 nor COR15am altered the dehydration-induced gel lamellar -> fluid lamellar phase transition temperature of either dipalmitoylphosphatidylcholine or dioleoylphosphatidylcholine (DOPC). In multilamellar vesicles of dioleoylphosphatidylethanolamine:DOPC (1:1, mol:mol) prepared by either freeze-thaw or reverse-phase evaporation methods, neither COR6.6, COR15am, nor bovine serum albumin altered the incidence of the dehydration-induced formation of the inverted hexagonal phase as a function of osmotic pressure. However, a specific ultrastructural alteration[mdash] the formation of a striated surface morphology in the lamellar domains[mdash]was observed in mixtures of dioleoylphosphatidylethanolamine:DOPC that were dehydrated in the presence of COR15am. Nevertheless, neither COR6.6 nor COR15am appears to participate in a specific protein-phospholipid interaction that alters the dehydration-induced phase behavior of phospholipid vesicles.
Tissue-Specific Expression and Promoter Analysis of the Tobacco ltp1 Gene
Authors: Canevascini S. , Caderas D. , Mandel T. , Fleming A. J. , Dupuis I. , Kuhlemeier C.
Published: July 2002

The Nicotiana tabacum ltp1 gene (Ntltp1) encodes a small basic protein that belongs to a class of putative lipid transfer proteins. These proteins transfer lipids between membranes in vitro, but their in vivo function remains hotly debated. This gene also serves as an important early marker for epidermis differentiation. We report here the analysis of the spatial and developmental activity of the Ntltp1 promoter, and we define a sequence element required for epidermis-specific expression. Transgenic plants were created containing 1346 bp of the Ntltp1 promoter fused upstream of the β-glucuronidase (GUS) gene. In the mature aerial tissues, GUS activity was detected predominantly in the epidermis, whereas in younger aerial tissues, such as the shoot apical meristem and floral meristem, GUS expression was not restricted to the tunica layer. Unexpectedly, GUS activity was also detected in young roots, particularly in the root epidermis. Furthermore, the Ntltp1 promoter displayed a tissue and developmental specific pattern of activity during germination. These results suggest that the Ntltp1 gene is highly expressed in regions of the plant that are vulnerable to pathogen attack and are thus consistent with the proposed function of lipid transfer proteins in plant defense. Deletions of the promoter from its 5[prime] end revealed that the 148 bp preceding the translational start site are sufficient for epidermis-specific expression. Sequence comparison identified an eight-nucleotide palindromic sequence CTAGCTAG in the leader of Ntltp1, which is conserved in a number of other ltp genes. By gel retardation analysis, the presence of specific DNA-protein complexes in this region was demonstrated. The characterization of these factors may lead to the identification of factors that control early events in epidermis differentiation.
Isolation and Characterization of a Diverse Set of Genes from Carrot Somatic Embryos
Authors: Lin X. , Hwang GJH. , Zimmerman J. L.
Published: July 2002

The early events in plant embryogenesis are critical for pattern formation, since it is during this process that the primary apical meristems and the embryo polarity axis are established. However, little is known about the molecular events that are unique to the early stages of embryogenesis. This study of gene expression during plant embryogenesis is focused on identifying molecular markers from carrot (Daucus carota) somatic embryos and characterizing the expression and regulation of these genes through embryo development. A cDNA library, prepared from polysomal mRNA of globular embryos, was screened using a subtracted probe; 49 clones were isolated and preliminarily characterized. Sequence analysis revealed a large set of genes, including many new genes, that are expressed in a variety of patterns during embryogenesis and may be regulated by different molecular mechanisms. To our knowledge, this group of clones represents the largest collection of embryo-enhanced genes isolated thus far, and demonstrates the utility of the subtracted-probe approach to the somatic embryo system. It is anticipated that many of these genes may serve as useful molecular markers for early embryo development.
Specific Localization of the Respiratory Alternative Oxidase in Meristematic and Xylematic Tissues from Developing Soybean Roots and Hypocotyls
Authors: Hilal M. , Castagnaro A. , Moreno H. , Massa E. M.
Published: July 2002

We used tissue printing and specific immunostaining to examine the localization of the alternative oxidase (AOX) protein in correlation with measurements of AOX capacity. Selected root and hypocotyl regions were analyzed during the first 14 d of growth. It is shown that AOX protein is localized in the apical meristem and in developing xylem. The temporal pattern of expression is coincident with the evolution of AOX capacity. Data suggest that AOX expression is linked to xylem differentiation. Since heat is a major product of the alternative pathway, we speculate that thermogenesis is implicated in morphogenesis.
Regulation of K+ Channels in Maize Roots by Water Stress and Abscisic Acid1
Authors: Roberts Stephen K.
Published: July 2002

Root cortical and stelar protoplasts were isolated from maize (Zea mays L.) plants that were either well watered or water stressed, and the patch-clamp technique was used to investigate their plasma membrane K+ channel activity. In the root cortex water stress did not significantly affect inward- or outward-rectifying K+ conductances relative to those observed in well-watered plants. In contrast, water stress significantly reduced the magnitude of the outward-rectifying K+ current in the root stele but had little effect on the inward-rectifying K+ current. Pretreating well-watered plants with abscisic acid also significantly affected K+currents in a way that was consistent with abscisic acid mediating, at least in part, the response of roots to water stress. It is proposed that the K+ channels underlying the K+ currents in the root stelar cells represent pathways that allow K+exchange between the root symplasm and xylem apoplast. It is suggested that the regulation of K+ channel activity in the root in response to water stress could be part of an important adaptation of the plant to survive drying soils.
Molecular and Physiological Responses to Water Deficit in Drought-Tolerant and Drought-Sensitive Lines of Sunflower1
Authors: Cellier Françoise , Conéjéro Geneviève , Breitler Jean-Christophe , Casse Francine
Published: July 2002

To investigate correlations between phenotypic adaptation to water limitation and drought-induced gene expression, we have studied a model system consisting of a drought-tolerant line (R1) and a drought-sensitive line (S1) of sunflowers (Helianthus annuus L.) subjected to progressive drought. R1 tolerance is characterized by the maintenance of shoot cellular turgor. Drought-induced genes (HaElip1, HaDhn1, andHaDhn2) were previously identified in the tolerant line. The accumulation of the corresponding transcripts was compared as a function of soil and leaf water status in R1 and S1 plants during progressive drought. In leaves of R1 plants the accumulation ofHaDhn1 and HaDhn2 transcripts, but notHaElip1 transcripts, was correlated with the drought-adaptive response. Drought-induced abscisic acid (ABA) concentration was not associated with the varietal difference in drought tolerance. Stomata of both lines displayed similar sensitivity to ABA. ABA-induced accumulation of HaDhn2 transcripts was higher in the tolerant than in the sensitive genotype.HaDhn1 transcripts were similarly accumulated in the tolerant and in the sensitive plants in response to ABA, suggesting that additional factors involved in drought regulation ofHaDhn1 expression might exist in tolerant plants.
Antioxidant Defenses in the Peripheral Cell Layers of Legume Root Nodules1
Authors: Dalton David A. , Joyner Shannon L. , Becana Manuel , Iturbe-Ormaetxe Iñaki , Chatfield J. Mark
Published: July 2002

Ascorbate peroxidase (AP) is a key enzyme that scavenges potentially harmful H2O2and thus prevents oxidative damage in plants, especially in N2-fixing legume root nodules. The present study demonstrates that the nodule endodermis of alfalfa (Medicago sativa) root nodules contains elevated levels of AP protein, as well as the corresponding mRNA transcript and substrate (ascorbate). Enhanced AP protein levels were also found in cells immediately peripheral to the infected region of soybean (Glycine max), pea (Pisum sativum), clover (Trifolium pratense), and common bean (Phaseolus vulgaris) nodules. Regeneration of ascorbate was achieved by (homo)glutathione and associated enzymes of the ascorbate-glutathione pathway, which were present at high levels. The presence of high levels of antioxidants suggests that respiratory consumption of O2in the endodermis or nodule parenchyma may be an essential component of the O2-diffusion barrier that regulates the entry of O2 into the central region of nodules and ensures optimal functioning of nitrogenase.
Moderately High Temperatures Inhibit Ribulose-1,5-Bisphosphate Carboxylase/Oxygenase (Rubisco) Activase-Mediated Activation of Rubisco1
Authors: Feller Urs , Crafts-Brandner Steven J. , Salvucci Michael E.
Published: July 2002

We tested the hypothesis that light activation of ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) is inhibited by moderately elevated temperature through an effect on Rubisco activase. When cotton (Gossypium hirsutum L.) or wheat (Triticum aestivum L.) leaf tissue was exposed to increasing temperatures in the light, activation of Rubisco was inhibited above 35 and 30°C, respectively, and the relative inhibition was greater for wheat than for cotton. The temperature-induced inhibition of Rubisco activation was fully reversible at temperatures below 40°C. In contrast to activation state, total Rubisco activity was not affected by temperatures as high as 45°C. Nonphotochemical fluorescence quenching increased at temperatures that inhibited Rubisco activation, consistent with inhibition of Calvin cycle activity. Initial and maximal chlorophyll fluorescence were not significantly altered until temperatures exceeded 40°C. Thus, electron transport, as measured by Chl fluorescence, appeared to be more stable to moderately elevated temperatures than Rubisco activation. Western-blot analysis revealed the formation of high-molecular-weight aggregates of activase at temperatures above 40°C for both wheat and cotton when inhibition of Rubisco activation was irreversible. Physical perturbation of other soluble stromal enzymes, including Rubisco, phosphoribulokinase, and glutamine synthetase, was not detected at the elevated temperatures. Our evidence indicates that moderately elevated temperatures inhibit light activation of Rubisco via a direct effect on Rubisco activase.
Genetic Manipulation of Condensed Tannins in Higher Plants1
Authors: Robbins Mark P. , Bavage Adrian D. , Strudwicke Catherine , Morris Phillip
Published: July 2002

We have produced and analyzed transgenic birdsfoot trefoil (Lotus corniculatus L.) plants harboring antisense dihydroflavonol reductase (AS-DFR) sequences. In initial experiments the effect of introducing three different antisense Antirrhinum majusL. DFR constructs into a single recipient genotype (S50) was assessed. There were no obvious effects on plant biomass, but levels of condensed tannins showed a statistical reduction in leaf, stem, and root tissues of some of the antisense lines. Transformation events were also found, which resulted in increased levels of condensed tannins. In subsequent experiments a detailed study of AS-DFR phenotypes was carried out in genotype S33 using pMAJ2 (an antisense construct comprising the 5′ half of the A. majus cDNA). In this case, reduced tannin levels were found in leaf and stem tissues and in juvenile shoot tissues. Analysis of soluble flavonoids and isoflavonoids in tannin down-regulated shoot tissues indicated few obvious default products. When two S33 AS-DFR lines were outcrossed, there was an underrepresentation of transgene sequences in progeny plants and no examples of inheritance of an antisense phenotype were observed. To our knowledge, this is the first report of the genetic manipulation of condensed tannin biosynthesis in higher plants.
Specific Soybean Lipoxygenases Localize to Discrete Subcellular Compartments and Their mRNAs Are Differentially Regulated by Source-Sink Status1
Authors: Stephenson Lowry C. , Bunker Thomas W. , Dubbs Wesley E. , Grimes Howard D.
Published: July 2002

Members of the lipoxygenase multigene family, found widely in eukaryotes, have been proposed to function in nitrogen partitioning and storage in plants. Lipoxygenase gene responses to source-sink manipulations in mature soybean (Glycine max [L.] Merr.) leaves were examined using gene-specific riboprobes to the five vegetative lipoxygenases (vlxA–vlxE). Steady-state levels of allvlx mRNAs responded strongly to sink limitation, but specific transcripts exhibited differential patterns of response as well. During reproductive sink limitation, vlxA andvlxB messages accumulated to high levels, whereasvlxC and vlxD transcript levels were modest. Immunolocalization using peptide-specific antibodies demonstrated that under control conditions, VLXB was present in the cytosol of the paraveinal mesophyll and with pod removal accumulated additionally in the bundle-sheath and adjacent cells. With sink limitation VLXD accumulated to apparent high levels in the vacuoles of the same cells. Segregation of gene products at the cellular and subcellular levels may thus permit complex patterns of differential regulation within the same cell type. Specific lipoxygenase isoforms may have a role in short-term nitrogen storage (VLXC/D), whereas others may simultaneously function in assimilate partitioning as active enzymes (VLXA/B).
Controlled Cytokinin Production in Transgenic Tobacco Using a Copper-Inducible Promoter
Authors: McKenzie Marian Jane , Mett Vadim , Reynolds Paul Hugh Stewart , Jameson Paula Elizabeth
Published: July 2002

The cytokinin group of plant hormones regulates aspects of plant growth and development, including the release of lateral buds from apical dominance and the delay of senescence. In this work the native promoter of a cytokinin synthase gene (ipt) was removed and replaced with a Cu-controllable promoter. Tobacco (Nicotiana tabacum L. cv tabacum) transformed with this Cu-inducibleipt gene (Cu-ipt) was morphologically identical to controls under noninductive conditions in almost all lines produced. However, three lines grew in an altered state, which is indicative of cytokinin overproduction and was confirmed by a full cytokinin analysis of one of these lines. The in vitro treatment of morphologically normal Cu-ipt transformants with Cu2+ resulted in delayed leaf senescence and an increase in cytokinin concentration in the one line analyzed. In vivo, inductive conditions resulted in a significant release of lateral buds from apical dominance. The morphological changes seen during these experiments may reflect the spatial aspect of control exerted by this gene expression system, namely expression from the root tissue only. These results confirmed that endogenous cytokinin concentrations in tobacco transformants can be temporally and spatially controlled by the induction of ipt gene expression through the Cu-controllable gene-expression system.
Control of Cl− Efflux in Chara corallina by Cytosolic pH, Free Ca2+, and Phosphorylation Indicates a Role of Plasma Membrane Anion Channels in Cytosolic pH Regulation1
Authors: Johannes Eva , Crofts Alan , Sanders Dale
Published: July 2002

Enhanced Cl− efflux during acidosis in plants is thought to play a role in cytosolic pH (pHc) homeostasis by short-circuiting the current produced by the electrogenic H+ pump, thereby facilitating enhanced H+ efflux from the cytosol. Using an intracellular perfusion technique, which enables experimental control of medium composition at the cytosolic surface of the plasma membrane of charophyte algae (Chara corallina), we show that lowered pHc activates Cl− efflux via two mechanisms. The first is a direct effect of pHc on Cl− efflux; the second mechanism comprises a pHc-induced increase in affinity for cytosolic free Ca2+ ([Ca2+]c), which also activates Cl− efflux. Cl− efflux was controlled by phosphorylation/dephosphorylation events, which override the responses to both pHc and [Ca2+]c. Whereas phosphorylation (perfusion with the catalytic subunit of protein kinase A in the presence of ATP) resulted in a complete inhibition of Cl− efflux, dephosphorylation (perfusion with alkaline phosphatase) arrested Cl− efflux at 60% of the maximal level in a manner that was both pHc and [Ca2+]cindependent. These findings imply that plasma membrane anion channels play a central role in pHc regulation in plants, in addition to their established roles in turgor/volume regulation and signal transduction.