bluuebunny/crossref_metadata_2025 huggingface dataset:
Identifying the source of species invasions: sampling intensity vs. genetic diversity
Authors: MUIRHEAD JIM R. , GRAY DEREK K. , KELLY DAVID W. , ELLIS SANDRA M. , HEATH DANIEL D. , MACISAAC HUGH J.
Published: February 2008

Population geneticists and community ecologists have long recognized the importance of sampling design for uncovering patterns of diversity within and among populations and in communities. Invasion ecologists increasingly have utilized phylogeographical patterns of mitochondrial or chloroplast DNA sequence variation to link introduced populations with putative source populations. However, many studies have ignored lessons from population genetics and community ecology and are vulnerable to sampling errors owing to insufficient field collections. A review of published invasion studies that utilized mitochondrial or chloroplast DNA markers reveals that insufficient sampling could strongly influence results and interpretations. Sixty per cent of studies sampled an average of less than six individuals per source population, vs. only 45% for introduced populations. Typically, far fewer introduced than source populations were surveyed, although they were sampled more intensively. Simulations based on published data forming a comprehensive mtDNA haplotype data set highlight and quantify the impact of the number of individuals surveyed per source population and number of putative source populations surveyed for accurate assignment of introduced individuals. Errors associated with sampling a low number of individuals are most acute when rare source haplotypes are dominant or fixed in the introduced population. Accuracy of assignment of introduced individuals is also directly related to the number of source populations surveyed and to the degree of genetic differentiation among them (FST). Incorrect interpretations resulting from sampling errors can be avoided if sampling design is considered before field collections are made.
Intracellular mobility of plasmid DNA is limited by the ParA family of partitioning systems
Authors: Derman Alan I. , Lim‐Fong Grace , Pogliano Joe
Published: February 2008

The highly conserved ParA family of partitioning systems is responsible for positioning DNA and protein complexes in bacteria. In Escherichia coli, plasmids that rely upon these systems are positioned at mid‐cell and are repositioned at the quarter‐cell positions after replication. How they remain fixed at these positions throughout the cell cycle is unknown. We use fluorescence recovery after photobleaching and time‐lapse microscopy to measure plasmid mobility in living E. coli cells. We find that a minimalized version of plasmid RK2 that lacks its Par system is highly mobile, that the intact RK2 plasmid is relatively immobile, and that the addition of a Par system to the minimalized RK2 plasmid limits its mobility to that of the intact RK2. Mobility is thus the default state, and Par systems are required not only to position plasmids, but also to hold them at these positions. The intervention of Par systems is required continuously throughout the cell cycle to restrict plasmid movement that would, if unrestricted, subvert the segregation process. Our results reveal an important function for Par systems in plasmid DNA segregation that is likely to be conserved in bacteria.
A polymerase III‐like reinitiation mechanism is operating in regulation of histone expression in archaea
Authors: Spitalny Patrizia , Thomm Michael
Published: February 2008

An archaeal histone gene from the hyperthermophile Pyrococcus furiosus containing four consecutive putative oligo‐dT terminator sequences was used as a model system to investigate termination signals and the mechanism of termination in vitro. The archaeal RNA polymerase terminated with high efficiency at the first terminator at 90°C when it contained five to six T residues, at 80°C readthrough was significantly increased. A putative hairpin structure upstream of the first terminator had no effect on termination efficiency. Template competition experiments starting with RNA polymerase molecules engaged in ternary complexes revealed recycling of RNA polymerase from the terminator to the promoter of the same template. This facilitated reinitiation was dependent upon the presence of a terminator sequence suggesting that pausing at the terminator is required for recycling as in the RNA polymerase III system. Replacement of the sequences immediately downstream of the oligo‐dT terminator by an AT‐rich segment improved termination efficiency. Both AT‐rich and GC‐rich downstream sequences seemed to impair the facilitated reinitiation pathway. Our data suggest that recycling is dependent on a subtle interplay of pausing of RNA polymerase at the terminator and RNA polymerase translocation beyond the oligo‐dT termination signal that is dramatically affected by downstream sequences.
Double illegitimate recombination events integrate DNA segments through two different mechanisms during natural transformation of Acinetobacter baylyi
Authors: Hülter Nils , Wackernagel Wilfried
Published: February 2008

Acquisition of foreign DNA by horizontal gene transfer is seen as a major source of genetic diversity in prokaryotes. However, strongly divergent DNA is not genomically integrated by homologous recombination and would depend on illegitimate recombination (IR) events which are rare. We show that, by two mechanisms, during natural transformation of Acinetobacter baylyi two IR events can integrate DNA segments. One mechanism is double illegitimate recombination (DIR) acting in the absence of any homology (frequency: 7 × 10−13 per cell). It occurs about 1010‐fold less frequent than homologous transformation. The other mechanism is homology‐facilitated double illegitimate recombination (HFDIR) being about 440‐fold more frequent (3 × 10−10 per cell) than DIR. HFDIR depends on a homologous sequence located between the IR sites and on recA+. In HFDIR two IR events act on the same donor DNA molecule as shown by the joint inheritance of molecular DNA tags. While the IR events in HFDIR occurred at microhomologies, in DIR microhomologies were not used. The HFDIR phenomenon indicates that a temporal recA‐dependent association of donor DNA at a homology in recipient DNA may facilitate two IR events on the 5′ and 3′ heterologous parts of the transforming DNA molecule.
Regulatory interplay between pap operons in uropathogenic Escherichia coli
Authors: Totsika Makrina , Beatson Scott A. , Holden Nicola , Gally David L.
Published: February 2008

Pathogenic bacteria have a large repertoire of surface organelles involved in adherence, motility and protein export, but how individual bacteria co‐ordinate surface organelle expression to prevent interference and excessive immune stimulation is unclear. Phase variation is a mechanism by which expression of surface factors is limited to a fraction of the bacterial population; however, the presence of multiple homologous surface structures controlled by related mechanisms and regulators antagonizes the independent expression achieved by phase variation. To investigate whether other mechanisms have evolved to sort out the bacterial cell surface, we examined regulatory cross‐talk between multiple phase‐variable pyelonephritis‐associated pili (pap) operons in Escherichia coli isolates associated with urinary tract infections. Allelic variation identified in the regulatory regions and regulators acts synergistically to limit coexpression of homologous fimbrial operons. In particular, there is evidence that papI is under positive selection and PapI variants displayed differences in their capacity to activate related pap operons. Alleles of the high‐affinity binding site for PapB were shown to contain a variable number of (T/A)3 repeats occurring every 9 bp that altered the sensitivity of pap operon activation. Taken together with other examples of surface organelle cross‐talk, we illustrate how this regulation could promote sequential expression.
Requirements for translation re‐initiation in Escherichia coli: roles of initiator tRNA and initiation factors IF2 and IF3
Authors: Yoo Jae‐Ho , RajBhandary Uttam L.
Published: February 2008

Despite its importance in post‐transcriptional regulation of polycistronic operons in Escherichia coli, little is known about the mechanism of translation re‐initiation, which occurs when the same ribosome used to translate an upstream open reading frame (ORF) also translates a downstream ORF. To investigate translation re‐initiation in Escherichia coli, we constructed a di‐cistronic reporter in which a firefly luciferase gene was linked to a chloramphenicol acetyltransferase gene using a segment of the translationally coupled geneV–geneVII intercistronic region from M13 phage. With this reporter and mutant initiator tRNAs, we show that two of the unique properties of E. coli initiator tRNA – formylation of the amino acid attached to the tRNA and binding of the tRNA to the ribosomal P‐site – are as important for re‐initiation as for de novo initiation. Overexpression of IF2 or increasing the affinity of mutant initiator tRNA for IF2 enhanced re‐initiation efficiency, suggesting that IF2 is required for efficient re‐initiation. In contrast, overexpression of IF3 led to a marked decrease in re‐initiation efficiency, suggesting that a 30S ribosome and not a 70S ribosome is used for translation re‐initiation. Strikingly, overexpression of IF3 also blocked E. coli from acting as a host for propagation of M13 phage.
The Gα subunit BCG1, the phospholipase C (BcPLC1) and the calcineurin phosphatase co‐ordinately regulate gene expression in the grey mould fungusBotrytis cinerea
Authors: Schumacher Julia , Viaud Muriel , Simon Adeline , Tudzynski Bettina
Published: February 2008

The Gα subunit BCG1 is essential for pathogenicity of the grey mould fungusBotrytis cinerea. Several processes such as the transition from primary infection to secondary invasive growth and the production of the phytotoxin botrydial are regulated by BCG1 via a cAMP‐independent pathway. Our recent finding that the botrydial biosynthesis genes belong to the group of Ca2+/calcineurin‐dependent genes suggested for the first time a connection between this Gα subunit and the calcineurin signalling pathway. To investigate whether this co‐regulation of genes by BCG1 and calcineurin is a common feature, a cDNA macroarray approach was used to compare the gene expression pattern of the wild‐type and the Δbcg1mutant, non‐treated or treated with the calcineurin inhibitor cyclosporin A. We identified three sets of genes whose expression was regulated either by both BCG1 and calcineurin, or only by one of them. Among the BCG1/calcineurin‐co‐regulated genes, we found a new gene cluster coding for a yet unknown polyketide secondary metabolite. Furthermore, we show for the first time in a phytopathogenic fungus that the phospholipase C (BcPLC1) is a component of the BCG1‐ and calcineurin‐dependent signalling pathway as several BCG1‐ and calcineurin‐dependent genes were downregulated inbcplc1knock‐down mutants.
ABC‐type amino acid uptake transporters Bgt and N‐II of Anabaena sp. strain PCC 7120 share an ATPase subunit and are expressed in vegetative cells and heterocysts
Authors: Pernil Rafael , Picossi Silvia , Mariscal Vicente , Herrero Antonia , Flores Enrique
Published: February 2008

Anabaena sp. strain PCC 7120 is a filamentous cyanobacterium that can fix N2 in differentiated cells called heterocysts. Anabaena open reading frames alr4167 and alr3187 encode, respectively, an ATPase subunit, BgtA, and a composite protein bearing periplasmic substrate‐binding and transmembrane domains, BgtB, of an ABC‐type high‐affinity basic amino acid uptake transporter (Bgt). Open reading frame alr4167 is clustered with open reading frames alr4164, alr4165 and alr4166 that encode a periplasmic substrate‐binding protein, NatF, and transmembrane proteins NatG and NatH respectively. The NatF, NatG, NatH and BgtA proteins constitute an ABC‐type uptake transporter for acidic and neutral polar amino acids (N‐II). The Bgt and N‐II transport systems thus share the ATPase subunit, BgtA. These transporters together with the previously characterized ABC‐type uptake transporter for proline and hydrophobic amino acids (N‐I) account for more than 98% of the amino acid transport activity exhibited by Anabaena sp. strain PCC 7120. In contrast to N‐I that is expressed only in vegetative cells, the Bgt and N‐II systems are present in both vegetative cells and heterocysts. Whereas Bgt is dispensable for diazotrophic growth, N‐II appears to contribute together with N‐I to the diazotrophic physiology of this cyanobacterium.
Mitochondrial fatty acid synthesis is required for normal mitochondrial morphology and function in Trypanosoma brucei
Authors: Guler Jennifer L. , Kriegova Eva , Smith Terry K. , Lukeš Julius , Englund Paul T.
Published: February 2008

Trypanosoma brucei use microsomal elongases for de novo synthesis of most of its fatty acids. In addition, this parasite utilizes an essential mitochondrial type II synthase for production of octanoate (a lipoic acid precursor) as well as longer fatty acids such as palmitate. Evidence from other organisms suggests that mitochondrially synthesized fatty acids are required for efficient respiration but the exact relationship remains unclear. In procyclic form trypanosomes, we also found that RNAi depletion of the mitochondrial acyl carrier protein, an important component of the fatty acid synthesis machinery, significantly reduces cytochrome‐mediated respiration. This reduction was explained by RNAi‐mediated inhibition of respiratory complexes II, III and IV, but not complex I. Other effects of RNAi, such as changes in mitochondrial morphology and alterations in membrane potential, raised the possibility of a change in mitochondrial membrane composition. Using mass spectrometry, we observed a decrease in total and mitochondrial phosphatidylinositol and mitochondrial phosphatidylethanolamine. Thus, we conclude that the mitochondrial synthase produces fatty acids needed for maintaining local phospholipid levels that are required for activity of respiratory complexes and preservation of mitochondrial morphology and function.
Compensatory role for Rad52 during recombinational repair in Ustilago maydis
Authors: Kojic Milorad , Mao Ninghui , Zhou Qingwen , Lisby Michael , Holloman William K.
Published: February 2008

A single Rad52‐related protein is evident by blast analysis of the Ustilago maydis genome database. Mutants created by disruption of the structural gene exhibited few discernible defects in resistance to UV, ionizing radiation, chemical alkylating or cross‐linking agents. No deficiency was noted in spontaneous mutator activity, allelic recombination or meiosis. GFP‐Rad51 foci were formed in rad52 cells following DNA damage, but were initially less intense than normal suggesting a possible role for Rad52 in formation of the Rad51 nucleoprotein filament. A search for interacting genes that confer a synthetic fitness phenotype with rad52 after DNA damage by UV irradiation identified the genes for Mph1, Ercc1 and the Rad51 paralogue Rec2. Testing known mutants in recombinational repair revealed an additional interaction with the BRCA2 orthologue Brh2. Suppression of the rec2 mutant's UV sensitivity by overexpressing Brh2 was found to be dependent on Rad52. The results suggest that Rad52 serves in an overlapping, compensatory role with both Rec2 and Brh2 to promote and maintain formation of the Rad51 nucleoprotein filament.
How the early sporulation sigma factor σF delays the switch to late development in Bacillus subtilis
Authors: Karmazyn‐Campelli Céline , Rhayat Lamya , Carballido‐López Rut , Duperrier Sandra , Frandsen Niels , Stragier Patrick
Published: February 2008

Sporulation in Bacillus subtilis is a primitive differentiation process involving two cell types, the forespore and the mother cell. Each cell implements two successive transcription programmes controlled by specific sigma factors. We report that activity of σG, the late forespore sigma factor, is kept in check by Gin, the product of csfB, a gene controlled by σF, the early forespore sigma factor. Gin abolishes σG transcriptional activity when σG is artificially synthesized during growth, but has no effect on σF. Gin interacts strongly with σG but not with σF in a yeast two‐hybrid experiment. The absence of Gin allows σG to be active during sporulation independently of the mother‐cell development to which it is normally coupled. Premature σG activity leads to the formation of slow‐germinating spores, and complete deregulation of σG synthesis is lethal when combined with gin inactivation. Gin allows σF to delay the switch to the late forespore transcription programme by preventing σG to take over before the cell has reached a critical stage of development. A similar strategy, following a completely unrelated route, is used by the mother cell.
Mid-infrared imaging of 18 planetary nebulae using the Spitzer Space Telescope
Authors: Phillips J. P. , Ramos-Larios G.
Published: February 2008

We present 3.6-, 4.5-, 5.8- and 8.0-μm photometric mapping of 18 galactic planetary nebulae, based on observations taken with the Spitzer Space Telescope. These are shown to have morphologies which are sometimes quite different from those observed in the visible, with much of the emission arising outside the ionized shells. There is also evidence for a change in nebular sizes between the differing photometric bands. An analysis of mid-infrared (MIR) colours suggests that many nebulae have dust/polycyclic aromatic hydrocarbon (PAH) emission components, and it seems likely that longer wave MIR fluxes (in particular) are associated with PAH emission features. Such features are likely to be associated, in addition, with photodissociative regimes, where shock fragmentation of dust may lead to the replenishment of smaller PAH grains.

Finally, we note that the source Ap 2–1 possesses a particularly interesting structure, and may represent a case in which the nebular shell is ploughing through an enveloping H ii region. Similarly, it appears that central emission in M 2–48 may arise from a warm and dusty collimating disc.
Chemical coding and central projections of gastric vagal afferent neurons
Authors: Young R. L. , Cooper N. J. , Blackshaw L. A.
Published: February 2008

Abstract  Vagal afferents that innervate gastric muscle or mucosa transmit distinct sensory information from their endings to the nucleus of the tractus solitarius (NTS). While these afferent subtypes are functionally distinct, no neurochemical correlate has been described and it is unknown whether they terminate in different central locations. This study aimed to identify gastric vagal afferent subtypes in the nodose ganglion (NG) of ferrets, their terminal areas in NTS and neurochemistry for isolectin‐B4 (IB4) and calcitonin gene‐related peptide (CGRP). Vagal afferents were traced from gastric muscle or mucosa and IB4 and CGRP labelling assessed in NG and NTS. 7 ± 1% and 6 ± 1% of NG neurons were traced from gastric muscle or mucosa respectively; these were more likely to label for CGRP or for both CGRP and IB4 than other NG neurons (P < 0.01). Muscular afferents were also less likely than others to label with IB4 (P < 0.001). Less than 1% of NG neurons were traced from both muscle and mucosa. Central terminals of both afferent subtypes occurred in the subnucleus gelatinosus of the NTS, but did not overlap completely. This region also labelled for CGRP and IB4. We conclude that while vagal afferents from gastric muscle and mucosa differ little in their chemical coding for CGRP and IB4, they can be traced selectively from their peripheral endings to NG and to overlapping and distinct regions of NTS. Thus, there is an anatomical substrate for convergent NTS integration for both types of afferent input.
Alvimopan and COX‐2 inhibition reverse opioid and inflammatory components of postoperative ileus
Authors: Schmidt J. , Stoffels B. , Nazir A. , Dehaven‐hudkins D. L. , Bauer A. J.
Published: February 2008

Abstract  Our objective was to investigate the therapeutic potential of peripheral opioid antagonism with alvimopan and anti‐inflammatory cyclooxygenase 2 (COX‐2) inhibition in an animal model of postoperative ileus with pain management. Intestinal manipulation was conducted in mice and rats with or without postoperative morphine injection. Rodents were orally fed non‐digestible fluorescein (FITC)‐labelled dextran and transit measured after a period of 90 min. The immunomodulatory effects of morphine and alvimopan were determined on nitric oxide released from the organ cultured muscularis externa. Surgical manipulation of the intestine resulted in a delay in gastrointestinal transit after 24 h that worsened with exogenous morphine. Alvimopan did not significantly alter transit of control or manipulated animals, but significantly antagonized the transit delaying effects of morphine. However, when the inflammatory component was robust enough to obscure a further opioid induced delay in gastrointestinal transit, alvimopan ceased to be effective in improving postoperative intestinal function. Cyclooxygenase 2 inhibition significantly diminished the inflammatory component of postoperative ileus. Surgical manipulation resulted in an increased release of nitric oxide from the inflamed isolated muscularis externa in 24‐h organ culture which was not altered by morphine or alvimopan. Two distinct mechanisms exist which participate in postoperative bowel dysfunction: a local inflammatory response which is antagonized by COX‐2 inhibition, and a morphine‐induced alteration in neural function which can be blocked with alvimopan.
Transgenic Leishmania and the immune response to infection
Authors: BEATTIE L. , EVANS K. J. , KAYE P. M. , SMITH D. F.
Published: February 2008

Genetic manipulation of single‐celled organisms such as the Leishmania parasite enables in depth analysis of the consequences of genotypic change on biological function. In probing the immune responses to infection, use of transgenic Leishmania has the potential to unravel both the contribution of the parasite to the infection process and the cellular interactions and mechanisms that characterize the innate and adaptive immune responses of the host. Here, we briefly review recent technical advances in parasite genetics and explore how these methods are being used to investigate parasite virulence factors, elucidate immune regulatory mechanisms and contribute to the development of novel therapeutics for the leishmaniases. Recent developments in imaging technology, such as bioluminescence and intravital imaging, combined with parasite transfection with fluorescent or enzyme‐encoding marker genes, provides a rich opportunity for novel assessment of intimate, real‐time host–parasite interactions at a previously unexplored level. Further advances in transgenic technology, such as the introduction of robust inducible gene cassettes for expression in intracellular parasite stages or the development of RNA interference methods for down‐regulation of parasite gene expression in the host, will further advance our ability to probe host–parasite interactions and unravel disease‐promoting mechanisms in the leishmaniases.
Chilling induces a decrease in pyrophosphate‐dependent H+‐accumulation associated with a ΔpHvac‐stat in mung bean, a chill‐sensitive plant
Authors: KAWAMURA YUKIO
Published: February 2008

Chilling leads to cytoplasmic acidification in chill‐sensitive plants. A possible explanation for this observation is that a ΔpH‐stat between the cytosol and vacuole (ΔpHvac‐stat) is perturbed by chilling. To understand the nature of this ΔpHvac‐stat, the effect of temperature, between 20 and 0 °C, on pyrophosphate (PPi)‐ or ATP‐dependent acidification of vacuolar vesicles, isolated from mung bean hypocotyls, was determined. Over the temperature range investigated, the H+‐influx mediated by PPase was balanced with the H+‐efflux, which was PPi‐dependently suppressed, and consequently a constant pH in vesicles (pHin) of ca. 5 was maintained against temperature changes. However, the ΔpHin driven by ATP decreased as the temperature dropped. Thus, the PPi‐dependent H+‐accumulation may function as an essential factor to form a ΔpHvac‐stat against temperature changes. Next, to study the chilling sensitivity of PPi‐dependent H+‐accumulation, vacuolar vesicles were isolated from control seedlings or from seedlings chilled at 0 °C for 1 d. Chilling treatment resulted in a decrease in the H+‐accumulation rate and in the steady‐state ΔpHin formed by PPi, the causes of which were enhanced by PPi‐dependent H+‐efflux and reduced by H+‐influx driven by PPase. Together, the results suggest that the decrease of PPi‐dependent H+‐accumulation associated with the ΔpHvac‐stat could result in cytoplasmic acidification.
The role of cytokinins in responses to water deficit in tobacco plants over‐expressing trans‐zeatin O‐glucosyltransferase gene under 35S or SAG12 promoters
Authors: HAVLOVÁ MARIE , DOBREV PETRE I. , MOTYKA VÁCLAV , ŠTORCHOVÁ HELENA , LIBUS JIŘÍ , DOBRÁ JANA , MALBECK JIŘÍ , GAUDINOVÁ ALENA , VANKOVÁ RADOMIRA
Published: February 2008

The impact of water deficit progression on cytokinin (CK), auxin and abscisic acid (ABA) levels was followed in upper, middle and lower leaves and roots of Nicotiana tabacum L. cv. Wisconsin 38 plants [wild type (WT)]. ABA content was strongly increased during drought stress, especially in upper leaves. In plants with a uniformly elevated total CK content, expressing constitutively the trans‐zeatin O‐glucosyltransferase gene (35S::ZOG1), a delay in the increase of ABA was observed; later on, ABA levels were comparable with those of WT.

As drought progressed, the bioactive CK content in leaves gradually decreased, being maintained longer in the upper leaves of all tested genotypes. Under severe stress (11 d dehydration), a large stimulation of cytokinin oxidase/dehydrogenase (CKX) activity was monitored in lower leaves, which correlated well with the decrease in bioactive CK levels. This suggests that a gradient of bioactive CKs in favour of upper leaves is established during drought stress, which might be beneficial for the preferential protection of these leaves.

During drought, significant accumulation of CKs occurred in roots, partially because of decreased CKX activity. Simultaneously, auxin increased in roots and lower leaves. This indicates that both CKs and auxin play a role in root response to severe drought, which involves the stimulation of primary root growth and branching inhibition.
Characterization of growth‐phase‐specific responses to cold in Arabidopsis thaliana suspension‐cultured cells
Authors: SASAKI YUTAKA , TAKAHASHI KAZUE , OONO YOKO , SEKI MOTOAKI , YOSHIDA RIICHIRO , SHINOZAKI KAZUO , UEMURA MATSUO
Published: February 2008

To understand complex responses of plant cells to low temperatures, suspension‐cultured cells of Arabidopsis thaliana (line T87) were characterized during cold treatment. Freezing tolerance of cells collected at the lag or log phase was quite different: an increase in freezing tolerance during cold treatment was only detectable with cells at the lag phase. Although there were little differences in the osmolality of cells at the two growth phases, sugar content increased during cold treatment only in lag phase cells. Abscisic acid (ABA) content was greater at the lag phase than at the log phase throughout the cold treatment, and increased only in lag phase cells. Interestingly, ABA treatment resulted in an increase in freezing tolerance only in lag phase cells. Expression of cold‐responsive genes such as DREB1A/CBF3, COR15a and RD29A occurred in cells at the two growth phases; however, the extent of the induction of COR15a and RD29A was somewhat greater in cells at the lag phase than at the log phase. Microarray analysis revealed that cold‐regulated genes (COR genes) were categorized into three groups: up‐ or down‐regulated consistently during cold treatment, transiently after 1 d and later after 2 d of cold treatment. Among genes that were up‐regulated throughout or transiently after 1 d of cold treatment only in lag phase cells, functions in upstream of signal transduction such as kinase and transcription factor were often deduced. These results collectively suggest that these genes seem to be associated with induction of freezing tolerance by cold treatment in cells at the lag phase.
Metabolomic and proteomic changes in the xylem sap of maize under drought
Authors: ALVAREZ SOPHIE , MARSH ELLEN L. , SCHROEDER STEVE G. , SCHACHTMAN DANIEL P.
Published: February 2008

Plants produce compounds in roots that are transported to shoots via the xylem sap. Some of these compounds are vital for signalling and adaptation to environmental stress such as drought. In this study, we screened the xylem sap using mass spectrometry to quantify the changes in new and previously identified sap constituents under extended drought. We detected and quantified the changes in the concentration of 31 compounds present in the xylem sap under progressively increasing drought stress. We found changes in the hormones abscisic acid (ABA) and cytokinin, and the presence of high concentrations of the aromatic cytokinin 6‐benzylaminopurine (BAP). Several phenylpropanoid compounds (coumaric, caffeic and ferulic acids) were found in xylem sap. The concentrations of some of these phenylpropanoid compounds changed under drought. In parallel, an analysis of the xylem sap proteome was conducted. We found a higher abundance of cationic peroxidases, which with the increase in phenylpropanoids may lead to a reduction in lignin biosynthesis in the xylem vessels and could induce cell wall stiffening. The application of new methodologies provides insights into the range of compounds in sap and how alterations in composition may lead to changes in development and signalling during adaptation to drought.
Comparing model predictions and experimental data for the response of stomatal conductance and guard cell turgor to manipulations of cuticular conductance, leaf‐to‐air vapour pressure difference and temperature: feedback mechanisms are able to account for all observations
Authors: EAMUS DEREK , TAYLOR DANIEL T. , MACINNIS‐NG CATRIONA M. O. , SHANAHAN STEVE , DE SILVA LIONEL
Published: February 2008

Stomata respond to increasing leaf‐to‐air vapour pressure difference (LAVPD) (D) by closing. The mechanism by which this occurs is debated. A role for feedback and peristomatal transpiration has been proposed. In this paper, we apply a recent mechanistic model of stomatal behaviour, and compare model and experimental data for the influence of increasing D on stomatal conductance.

We manipulated cuticular conductance (gc) by three independent methods. First, we increased gc by using a solvent mixture applied to both leaf surfaces prior to determining stomatal responses to D; second, we increased gc by increasing leaf temperature at constant D; and third, we coated a small area of leaf with a light oil to decrease gc. In all three experiments, experimental data and model outputs showed very close agreement.

We conclude, from the close agreement between model and experimental data and the fact that manipulations of gc, and hence cuticular transpiration, influenced gs in ways consistent with a feedback mechanism, that feedback is central in determining stomatal responses to D.
Short‐term responses of leaf growth rate to water deficit scale up to whole‐plant and crop levels: an integrated modelling approach in maize
Authors: CHENU KARINE , CHAPMAN SCOTT C. , HAMMER GRAEME L. , MCLEAN GREG , SALAH HALIM BEN HAJ , TARDIEU FRANÇOIS
Published: February 2008

Physiological and genetic studies of leaf growth often focus on short‐term responses, leaving a gap to whole‐plant models that predict biomass accumulation, transpiration and yield at crop scale. To bridge this gap, we developed a model that combines an existing model of leaf 6 expansion in response to short‐term environmental variations with a model coordinating the development of all leaves of a plant. The latter was based on: (1) rates of leaf initiation, appearance and end of elongation measured in field experiments; and (2) the hypothesis of an independence of the growth between leaves. The resulting whole‐plant leaf model was integrated into the generic crop model APSIM which provided dynamic feedback of environmental conditions to the leaf model and allowed simulation of crop growth at canopy level. The model was tested in 12 field situations with contrasting temperature, evaporative demand and soil water status. In observed and simulated data, high evaporative demand reduced leaf area at the whole‐plant level, and short water deficits affected only leaves developing during the stress, either visible or still hidden in the whorl. The model adequately simulated whole‐plant profiles of leaf area with a single set of parameters that applied to the same hybrid in all experiments. It was also suitable to predict biomass accumulation and yield of a similar hybrid grown in different conditions. This model extends to field conditions existing knowledge of the environmental controls of leaf elongation, and can be used to simulate how their genetic controls flow through to yield.
Molecular and biological characterization of Macroptilium yellow mosaic virus from Jamaica
Authors: Amarakoon I. I. , Roye M. E. , Briddon R. W. , Bedford I. D. , Stanley J.
Published: February 2008

The molecular and biological characterization of a begomovirus infecting the common weed Macroptilium lathyroides from Jamaica are reported. The virus showed 92% sequence identity to an isolate of Macroptilium yellow mosaic virus (MaYMV) from Cuba, but was distinct from the two other begomoviruses isolated from M. lathyroides, namely Macroptilium yellow mosaic Florida virus (80% identity) and Macroptilium mosaic Puerto Rico virus (68% identity). Hence, the Jamaican begomovirus was considered an isolate of MaYMV and called Macroptilium yellow mosaic virus‐[Jamaica] (MaYMV‐[JM]). In infectivity studies using cloned DNA‐A and DNA‐B genomic components, MaYMV‐[JM] infected red kidney bean (Phaseolus vulgaris) and produced mild symptoms in Scotch Bonnet pepper (Capsicum chinense), but did not infect cabbage (Brassica oleracea). This information has implications for the development of strategies to control begomovirus diseases in Jamaica and elsewhere.
Human B Cells and Macrophages Cooperate in T‐cell‐independent Type 2 Response
Authors: Mättö M. , Raunio A.‐R. , Postila V. , Huttunen K. , Hirvonen M.‐R. , Pelkonen J.
Published: February 2008

We have analysed separately the role of B‐cell receptor (BCR) stimulation and the soluble second signal in the T‐cell‐independent type 2 (TI‐2) B‐cell response. We were able to show that human B cells and macrophages (Mφ) could function together in TI‐type microbial response. Interestingly, BCR cross‐linking of peripheral blood (PB) B cells enhanced IgG production induced by Mφ‐derived growth factors whereas interleukin (IL)‐12 + IL‐18 had milder effect on IgG production. We demonstrated that B‐cell‐derived soluble mediators primed lipopolysaccharide (LPS)‐stimulated Mφ for tumour necrosis factor‐α (TNF‐α) and IL‐6 production significantly better than IFN‐γ, confirming the role of B cells in the activation of Mφ. We could show that human PB B cells were active cytokine producers and could be induced to produce interferon (IFN)‐γ mRNA in the presence of known Mφ cytokines, like IL‐12 and IL‐18. BCR stimulation also stabilized and enhanced the IFN‐γ mRNA production induced by IL‐12 and IL‐18. In addition, our novel finding was that a known Mφ cytokine, IL‐10, induced the expression of IFN‐γ mRNA from human B‐cell line (HF28R0) cells. In summary, we propose a model for the active role of B cells in the induction of the inflammatory response during TI antigen challenge in close collaboration with Mφ.
Susceptibility of Human Monocyte‐macrophages to Chlamydia pneumoniae Infection In Vitro is Highly Variable and Associated with levels of Soluble CD14 and C. pneumoniae IgA and Human HSP‐IgG Antibodies in Serum
Authors: Poikonen K. , Lajunen T. , Silvennoinen‐Kassinen S. , Paldanius M. , Leinonen M. , Saikku P.
Published: February 2008

Chlamydia pneumoniae, an intracellular microbe, causes respiratory infections and may participate in the development of atherosclerosis. It is able to survive and multiply in macrophages. The susceptibility of monocyte‐macrophages from healthy individuals to C. pneumoniae infection in vitro was studied. Intracellular growth of C. pneumoniae, as an indicator of susceptibility to infection, was compared to serum levels of C‐reactive protein, soluble CD14 (sCD14), human heat shock protein (HSP)‐IgG, human HSP‐IgA, C. pneumoniae IgG and IgA antibodies. The production of C. pneumoniae in infected macrophages was highly variable, ranging from 0 to 638 chlamydial genomes per human genome. Chlamydia pneumoniae production associated positively with serum C. pneumoniae IgA (titre: ≥10) and hHSP‐IgG and negatively with sCD14 concentration. The association between sCD14 concentration, C. pneumoniae IgA and human HSP‐IgG antibodies and C. pneumoniae production was statistically significant only among males. Age and gender did not correlate with the production. We hypothesize that persons whose macrophages cannot restrict the growth of C. pneumoniae are more prone to chronic infection by this agent.
CD4+ T Cells Transfer Resistance Against Citrobacter rodentium‐Induced Infectious Colitis by Induction of Th 1 Immunity
Authors: Spahn T. W. , Ross M. , Von Eiff C. , Maaser C. , Spieker T. , Kannengiesser K. , Domschke W. , Kucharzik T.
Published: February 2008

Citrobacter rodentium induces an acute, self‐limited colitis in mice which is histologically associated with crypt hyperplasia. The infection serves as a model for human infectious colitis induced by enteropathogenic Escherichia coli. We investigated if Balb/c mice, which had spontaneously cleared C. rodentium infection, were protected against re‐infection and if resistance against intestinal infection can be systemically transferred using spleen cells. The course of infection was monitored by faecal excretion. Spleen cells, splenic CD3+ and CD4+ cells were transferred from resistant mice to non‐infected recipients prior to infection. Cytokine secretion, serum and faecal antibody titres and histological disease severity were assessed. Balb/c mice were resistant against re‐infection. The course of infection was shorter in mice receiving primed spleen cells, CD3+ and CD4+ cells. Transfer of CD4+ T cells from resistant mice induced γ‐interferon, interleukin (IL)‐2 and IL‐17 secretion and suppressed IL‐10 secretion. Anti‐Citrobacter serum IgG1 and IgG2a enzyme‐linked immunosorbent assay OD levels were increased. Faecal IgA secretion was increased while serum IgA was suppressed in recipients of CD4+ cells. Large bowel histology showed protection from colitis in recipients of primed cells as indicated by normal colonic epithelium. In Balb/c mice, C. rodentium infection is followed by resistance, which can be transferred by CD4+ cells. Transfer of protection is associated with IL‐17 secretion, enhanced serum IgG and faecal IgA secretion. This is the first study to demonstrate the mechanisms by which systemic resistance from previously C. rodentium‐infected mice can be transferred to non‐infected animals.
Antigen Analysis of Mycobacterium tuberculosis H37Rv Culture Filtrate Proteins
Authors: Målen H. , Søfteland T. , Wiker H. G.
Published: February 2008

Culture filtrates of Mycobacterium tuberculosis H37Rv highly enriched with secreted proteins were used to identify antigens recognized by a serum pool from tuberculosis patients. Two different approaches were used to separate the culture filtrate protein mixture: (i) proteins were fractionated according to their hydrophobicity using an HPLC‐C18 chromatography column followed by separation based on their molecular mass by SDS‐PAGE and subsequent immunoblotting or (ii) proteins were separated by two‐dimensional gel electrophoresis, based on their isoelectric point and their molecular mass. Twenty serologically reactive proteins were ultimately identified by both methods, including four novel antigens. Further, to estimate the immunogenicity of the identified culture filtrate proteins, the relative antibody quantities were measured using Image master software. Our results show that the antibodies against proteins belonging to the antigen 85 complex were the most abundant in the serum of patients with active tuberculosis. The most immunogenic proteins in terms of high antibody‐to‐protein‐ratio were Rv3881c and three lipoproteins Rv0934 (the 38 kDa antigen), Rv0932c (pstS2), and Rv3006 (LppZ). Rv3881c is located in the region of difference 1 (RD1) which is deleted from Mycobacterium bovis BCG, and is therefore a particularly promising candidate for development of serodiagnostic assays to detect active tuberculosis. The proteins from the M. tuberculosis H37Rv culture filtrate are strong candidates to be evaluated for improvement of the serodiagnostic tests of tuberculosis.
Absence of In vitro Responses to Type II Collagen by Splenocytes from Arthritic BALB/c Mice is Possibly Caused by Intrinsic CD25+ Regulatory Cells
Authors: Bäckström F. , Dahlgren U.
Published: February 2008

Collagen‐induced arthritis‐resistant BALB/c mice develop arthritis if a foreign protein is added to an emulsion of type II collagen (CII) and adjuvant. The IgG autoantibody activity to CII is increased, whereas no CII autoreactive T cells in vitro can be recorded. In this study, we have explored whether CD25+ cells inhibit T‐cell autoreactivity to CII. We also followed the IgG anti‐CII autoantibody activity and the IL‐6 level in serum during the development of arthritis. BALB/c mice were coimmunized with bovine CII (BCII) and keyhole limpet haemocyanin (KLH) in complete Freund’s adjuvant and boostered 3 weeks later. Control animals were immunized with either BCII or KLH. Sera were collected prior to and during the development of arthritis and examined for IgG anti‐CII antibody activity and IL‐6 content. When all BCII–KLH immunized mice had developed arthritis, splenocytes were prepared, with and without CD25+ cells, and tested for BCII reactivity in vitro. The serum IgG, IgG1 and IgG2a anti‐CII antibody activities and the IL‐6 level were significantly higher in BCII–KLH immunized mice than in BCII‐immunized animals that failed to develop arthritis. The BCII‐specific IL‐2 secretion in vitro was significantly increased in CD25‐depleted splenocyte cultures prepared from arthritic BCII–KLH‐immunized mice. Development of arthritis in BALB/c mice induced by coimmunization with BCII/KLH results in increased levels of circulating IL‐6 and IgG autoantibodies to CII. The arthritogenic BCII–KLH immunization potentiates BCII‐specific IL‐2 secretion by CD25‐depleted splenocytes, but CD25+ cells hamper the outcome of their action, at least in vitro.
Complex variations in sediment transport at three large river bifurcations during discharge waves in the river Rhine
Authors: FRINGS ROY M. , KLEINHANS MAARTEN G.
Published: February 2008

River bifurcations strongly control the distribution of water and sediment over a river system. A good understanding of this distribution process is crucial for river management. In this paper, an extensive data set from three large bifurcations in the Dutch Rhine is presented, containing data on bed‐load transport, suspended bed sediment transport, dune development and hydrodynamics. The data show complex variations in sediment transport during discharge waves. The objective of this paper is to examine and explain these measured variations in sediment transport. It is found that bend sorting upstream of the bifurcations leads to supply limitation, particularly in the downstream branch that originates in the outer bend of the main channel. Tidal water level variations lead to cyclical variations in the sediment distribution over the downstream branches. Lags in dune development cause complex hysteresis patterns in flow parameters and sediment transport. All bifurcations show evidence of sediment waves, which probably are intrinsic bifurcation phenomena. The complex transport processes at the three bifurcations cause distinct discontinuities in the downstream fining trend of the river. Differences among the studied river bifurcations are mainly due to differences in sediment mobility (Shields value). Because the variations in sediment transport are complex and poorly correlated with the flow discharge, prediction of the sediment distribution with existing relationships for one‐dimensional models is problematic.
How to usefully compare homologous plant genes and chromosomes as DNA sequences
Authors: Lyons Eric , Freeling Michael
Published: February 2008

There are four sequenced and publicly available plant genomes to date. With many more slated for completion, one challenge will be to use comparative genomic methods to detect novel evolutionary patterns in plant genomes. This research requires sequence alignment algorithms to detect regions of similarity within and among genomes. However, different alignment algorithms are optimized for identifying different types of homologous sequences. This review focuses on plant genome evolution and provides a tutorial for using several sequence alignment algorithms and visualization tools to detect useful patterns of conservation: conserved non‐coding sequences, false positive noise, subfunctionalization, synteny, annotation errors, inversions and local duplications. Our tutorial encourages the reader to experiment online with the reviewed tools as a companion to the text.
Gene silencing in plants using artificial microRNAs and other small RNAs
Authors: Ossowski Stephan , Schwab Rebecca , Weigel Detlef
Published: February 2008

Comprehensive analysis of gene function requires the detailed examination of mutant alleles. In Arabidopsis thaliana, large collections of sequence‐indexed insertion and chemical mutants provide potential loss‐of‐function alleles for most annotated genes. However, limitations for phenotypic analysis include gametophytic or early sporophytic lethality, and the ability to recombine mutant alleles in closely linked genes, especially those present as tandem duplications. Transgene‐mediated gene silencing can overcome some of these shortcomings through tissue‐specific, inducible and partial gene inactivation, or simultaneous targeting of several, sequence‐related genes. In addition, gene silencing is a convenient approach in species or varieties for which exhaustive mutant collections are not yet available. Typically, gene function is reduced post‐transcriptionally, effected by small RNAs that act in a sequence‐specific manner by base pairing to complementary mRNA molecules. A recently introduced approach is the use of artificial microRNAs (amiRNAs). Here, we review various strategies for small RNA‐based gene silencing, and describe in detail the design and application of amiRNAs in many plant species.