bluuebunny/crossref_metadata_2025_split huggingface dataset:
Interactions of Two Transcriptional Repressors and Two Transcriptional Activators in Modulating Gibberellin Signaling in Aleurone Cells
Authors: Zou Xiaolu , Neuman Dawn , Shen Qingxi J. 
Published: July 2008

AbstractGibberellins (GAs) regulate many aspects of plant development, such as germination, growth, and flowering. The barley (Hordeum vulgare) Amy32b α-amylase promoter contains at least five cis-acting elements that govern its GA-induced expression. Our previous studies indicate that a barley WRKY gene, HvWRKY38, and its rice (Oryza sativa) ortholog, OsWRKY71, block GA-induced expression of Amy32b-GUS. In this work, we investigated the functional and physical interactions of HvWRKY38 with another repressor and two activators in barley. HvWRKY38 blocks the inductive activities of SAD (a DOF protein) and HvGAMYB (a R2R3 MYB protein) when either of these proteins is present individually. However, SAD and HvGAMYB together overcome the inhibitory effect of HvWRKY38. Yet, the combination of HvWRKY38 and BPBF (another DOF protein) almost diminishes the synergistic effect of SAD and HvGAMYB transcriptional activators. Electrophoretic mobility shift assays indicate that HvWRKY38 blocks the GA-induced expression of Amy32b by interfering with the binding of HvGAMYB to the cis-acting elements in the α-amylase promoter. The physical interaction of HvWRKY38 and BPBF repressors is demonstrated via bimolecular fluorescence complementation assays. These data suggest that the expression of Amy32b is modulated by protein complexes that contain either activators (e.g. HvGAMYB and SAD) or repressors (e.g. HvWRKY38 and BPBF). The relative amounts of the repressor or activator complexes binding to the Amy32b promoter regulate its expression level in barley aleurone cells.
Geminivirus-Mediated Gene Silencing fromCotton Leaf Crumple VirusIs Enhanced by Low Temperature in Cotton
Authors: Tuttle John R. , Idris A.M. , Brown Judith K. , Haigler Candace H. , Robertson Dominique 
Published: July 2008

AbstractA silencing vector for cotton (Gossypium hirsutum) was developed from the geminivirus Cotton leaf crumple virus (CLCrV). The CLCrV coat protein gene was replaced by up to 500 bp of DNA homologous to one of two endogenous genes, the magnesium chelatase subunit I gene (ChlI) or the phytoene desaturase gene (PDS). Cotyledons of cotton cultivar ‘Deltapine 5415’ bombarded with the modified viral vectors manifested chlorosis due to silencing of either ChlI or PDS in approximately 70% of inoculated plants after 2 to 3 weeks. Use of the green fluorescence protein gene showed that replication of viral DNA was restricted to vascular tissue and that the viral vector could transmit to leaves, roots, and the ovule integument from which fibers originate. Temperature had profound effects on vector DNA accumulation and the spread of endogenous gene silencing. Consistent with reports that silencing against viruses increases at higher temperatures, plants grown at a 30°C/26°C day/night cycle had a greater than 10-fold reduction in viral DNA accumulation compared to plants grown at 22°C/18°C. However, endogenous gene silencing decreased at 30°C/26°C. There was an approximately 7 d delay in the onset of gene silencing at 22°C/18°C, but silencing was extensive and persisted throughout the life of the plant. The extent of silencing in new growth could be increased or decreased by changing temperature regimes at various times following the onset of silencing. Our experiments establish the use of the CLCrV silencing vector to study gene function in cotton and show that temperature can have a major impact on the extent of geminivirus-induced gene silencing.
The Coenzyme A Biosynthetic Enzyme Phosphopantetheine Adenylyltransferase Plays a Crucial Role in Plant Growth, Salt/Osmotic Stress Resistance, and Seed Lipid Storage
Authors: Rubio Silvia , Whitehead Lynne , Larson Tony R. , Graham Ian A. , Rodriguez Pedro L. 
Published: July 2008

Abstract
Coenzyme A (CoA) is an essential cofactor in the metabolism of both prokaryotic and eukaryotic organisms and a universal five-step pathway is utilized to synthesize CoA from pantothenate. Null mutations in two of the five steps of this pathway led to embryo lethality and therefore viable reduction-of-function mutations are required to further study its role in plant biology. In this article, we have characterized a viable Arabidopsis (Arabidopsis thaliana) T-DNA mutant affected in the penultimate step of the CoA biosynthesis pathway, which is catalyzed by the enzyme phosphopantetheine adenylyltransferase (PPAT). This ppat-1 knockdown mutation showed an approximately 90% reduction in PPAT transcript levels and was severely impaired in plant growth and seed production. The sum of CoA and acetyl-CoA levels was severely reduced (60%–80%) in ppat-1 seedlings compared to wild type, and catabolism of storage lipids during seedling establishment was delayed. Conversely, PPAT overexpressing lines showed, on average, approximately 1.6-fold higher levels of CoA + acetyl-CoA levels, as well as enhanced vegetative and reproductive growth and salt/osmotic stress resistance. Interestingly, dry seeds of overexpressing lines contained between 35% to 50% more fatty acids than wild type, which suggests that CoA biosynthesis plays a crucial role in storage oil accumulation. Finally, biochemical analysis of the recombinant PPAT enzyme revealed an inhibitory effect of CoA on PPAT activity. Taken together, these results suggest that the reaction catalyzed by PPAT is a regulatory step in the CoA biosynthetic pathway that plays a key role for plant growth, stress resistance, and seed lipid storage.
Mutation of the Plastidial α-Glucan Phosphorylase Gene in Rice Affects the Synthesis and Structure of Starch in the Endosperm
Authors: Satoh Hikaru , Shibahara Kensuke , Tokunaga Takashi , Nishi Aiko , Tasaki Mikako , Hwang Seon-Kap , Okita Thomas W. , Kaneko Nanae , Fujita Naoko , Yoshida Mayumi , Hosaka Yuko , Sato Aya , Utsumi Yoshinori , Ohdan Takashi , Nakamura Yasunori 
Published: July 2008

Abstract
Plastidial phosphorylase (Pho1) accounts for ∼96% of the total phosphorylase activity in developing rice (Oryza sativa) seeds. From mutant stocks induced by N-methyl-N-nitrosourea treatment, we identified plants with mutations in the Pho1 gene that are deficient in Pho1. Strikingly, the size of mature seeds and the starch content in these mutants showed considerable variation, ranging from shrunken to pseudonormal. The loss of Pho1 caused smaller starch granules to accumulate and modified the amylopectin structure. Variation in the morphological and biochemical phenotype of individual seeds was common to all 15 pho1-independent homozygous mutant lines studied, indicating that this phenotype was caused solely by the genetic defect. The phenotype of the pho1 mutation was temperature dependent. While the mutant plants grown at 30°C produced mainly plump seeds at maturity, most of the seeds from plants grown at 20°C were shrunken, with a significant proportion showing severe reduction in starch accumulation. These results strongly suggest that Pho1 plays a crucial role in starch biosynthesis in rice endosperm at low temperatures and that one or more other factors can complement the function of Pho1 at high temperatures.
RXLR-Mediated Entry of Phytophthora sojae Effector Avr1b into Soybean Cells Does Not Require Pathogen-Encoded Machinery
Authors: Dou Daolong , Kale Shiv D. , Wang Xia , Jiang Rays H.Y. , Bruce Nathan A. , Arredondo Felipe D. , Zhang Xuemin , Tyler Brett M. 
Published: July 2008

Abstract
Effector proteins secreted by oomycete and fungal pathogens have been inferred to enter host cells, where they interact with host resistance gene products. Using the effector protein Avr1b of Phytophthora sojae, an oomycete pathogen of soybean (Glycine max), we show that a pair of sequence motifs, RXLR and dEER, plus surrounding sequences, are both necessary and sufficient to deliver the protein into plant cells. Particle bombardment experiments demonstrate that these motifs function in the absence of the pathogen, indicating that no additional pathogen-encoded machinery is required for effector protein entry into host cells. Furthermore, fusion of the Avr1b RXLR-dEER domain to green fluorescent protein (GFP) allows GFP to enter soybean root cells autonomously. The conclusion that RXLR and dEER serve to transduce oomycete effectors into host cells indicates that the &gt;370 RXLR-dEER–containing proteins encoded in the genome sequence of P. sojae are candidate effectors. We further show that the RXLR and dEER motifs can be replaced by the closely related erythrocyte targeting signals found in effector proteins of Plasmodium, the protozoan that causes malaria in humans. Mutational analysis of the RXLR motif shows that the required residues are very similar in the motifs of Plasmodium and Phytophthora. Thus, the machinery of the hosts (soybean and human) targeted by the effectors may be very ancient.
Calmodulin7 Plays an Important Role as Transcriptional Regulator inArabidopsisSeedling Development
Authors: Kushwaha Ritu , Singh Aparna , Chattopadhyay Sudip 
Published: July 2008

AbstractAlthough calmodulin (CaM) is known to play multiple regulatory roles in eukaryotes, its direct function as transcriptional regulator is unknown. Furthermore, the physiological functions of CaM are largely unknown in plants. Here, we show that one of the four Arabidopsis thaliana CaM isoforms, CAM7, is a transcriptional regulator that directly interacts with the promoters of light-inducible genes and promotes photomorphogenesis. CAM7 overexpression causes hyperphotomorphogenic growth and an increase in the expression of light-inducible genes. Mutations in CAM7 produce no visible effects on photomorphogenic growth, indicating likely redundant gene functions. However, cam7 mutants display reduced expression of light-inducible genes, and cam7 hy5 double mutants show an enhancement of the hy5 phenotype. Moreover, overexpression of CAM7 can partly suppress the hy5 phenotype, indicating that the two factors work together to control light-induced seedling development. The mutational and transgenic studies, together with physiological analyses, illustrate the concerted function of CAM7 and HY5 basic leucine zipper transcription factor in Arabidopsis seedling development.
A universal method for fishing target proteins from mixtures of biomolecules using isothermal titration calorimetry
Authors: Zhou Xingding , Sun Qingxiang , Kini R. Manjunatha , Sivaraman J. 
Published: July 2008

AbstractThe most challenging tasks in biology include the identification of (1) the orphan receptor for a ligand, (2) the ligand for an orphan receptor protein, and (3) the target protein(s) for a given drug or a lead compound that are critical for the pharmacological or side effects. At present, several approaches are available, including cell‐ or animal‐based assays, affinity labeling, solid‐phase binding assays, surface plasmon resonance, and nuclear magnetic resonance. Most of these techniques are not easy to apply when the target protein is unknown and the compound is not amenable to labeling, chemical modification, or immobilization. Here we demonstrate a new universal method for fishing orphan target proteins from a complex mixture of biomolecules using isothermal titration calorimetry (ITC) as a tracking tool. We took snake venom, a crude mixture of several hundred proteins/peptides, as a model to demonstrate our proposed ITC method in tracking the isolation and purification of two distinct target proteins, a major component and a minor component. Identities of fished out target proteins were confirmed by amino acid sequencing and inhibition assays. This method has the potential to make a significant advancement in the area of identifying orphan target proteins and inhibitor screening in drug discovery and characterization.
The hemoglobins of the trematodes Fasciola hepatica and Paramphistomum epiclitum: A molecular biological, physico‐chemical, kinetic, and vaccination study
Authors: Dewilde Sylvia , Ioanitescu A. Iulia , Kiger Laurent , Gilany Kambiz , Marden Michael C. , Van Doorslaer Sabine , Vercruysse Jozef , Pesce Alessandra , Nardini Marco , Bolognesi Martino , Moens Luc 
Published: July 2008

AbstractThe trematode Fasciola hepatica (Fa.he.) is a common parasite of human and livestock. The hemoglobin (Hb) of Fa.he., a potential immunogen, was chosen for characterization in the search for an effective vaccine. Characterization of trematode Hbs show that they are intracellular single‐domain globins with the following remarkable features: (1) Fa.he. expresses two Hb isoforms that differ at two amino acid sites (F1: 119Y/123Q; F2: 119F/123L). Both isoforms are monoacetylated at their N‐termini; (2) the genes coding for Fa.he. and Paramphistomum epiclitum (Pa.ep.) Hbs are interrupted by two introns at the conserved positions B12.2 and G7.0.; (3) UV/VIS and resonance Raman spectroscopy identify the recombinant Fa.he. HbF2 as a pentacoordinated high‐spin ferrous Hb; (4) electron paramagnetic resonance spectroscopy of cyano‐met Fa.he. HbF2 proves that the endogenously bound imidazole has no imidazolate character; (5) the major structural determinants of the globin fold are present, they contain a TyrB10/TyrE7 residue pair on the distal side. Although such distal‐site pair is a signature for high oxygen affinity, as shown for Pa.ep. Hb, the oxygen‐binding rate parameters for Fa.he. Hb are intermediate between those of myoglobin and those of other trematode Hbs; (6) the three‐dimensional structure of recombinant Fa.he. HbF2 from this study closely resembles the three‐dimensional structure of Pa.ep. determined earlier. The set of distal‐site polar interactions observed in Pa.ep. Hb is matched with small but significant structural adjustments; (7) despite the potential immunogenic character of the fluke Hb, vaccination of calves with recombinant Fa.he. HbF2 failed to promote protection against parasitic infection.
The novel p53 isoform “delta p53” is a misfolded protein and does not bind the p21 promoter site
Authors: García‐Alai Maria M. , Tidow Henning , Natan Eviatar , Townsley Fiona M. , Veprintsev Dmitry B. , Fersht Alan R. 
Published: July 2008

AbstractThe tumor suppressor p53 can be expressed as different isoforms because of promoter selection and mRNA editing. One isoform, “delta p53” (Δp53), results from what would be an unusual alternative splicing of exons 7/8 of the p53 gene, conserving the reading frame and generating a novel protein with proposed transcriptional activity essential for the intra S‐phase checkpoint. Here, we show that the deletion of the 66 residues that correspond to strand β10 and the C‐terminal helix of the core domain and the interconnecting linker to the tetramerization domain occurring in the Δp53 isoform leads to a misfolded and unstable protein, prone to form soluble aggregates, which does not bind the p21 promoter site. The complex of coexpressed Δp53 and flp53 is soluble in vitro and binds poorly to DNA. Our results provide a structural explanation for the dominant‐negative effect of Δp53 and its lack of transcriptional activity.
Estimating the size and dynamics of an injecting drug user population and implications for health service coverage: comparison of indirect prevalence estimation methods
Authors: Kimber Jo , Hickman Matthew , Degenhardt Louisa , Coulson Tim , Van Beek Ingrid 
Published: July 2008

ABSTRACTAims  (i) To compare indirect estimation methods to obtain mean injecting drug use (IDU) prevalence for a confined geographic location; and (ii) to use these estimates to calculate IDU and injection coverage of a medically supervised injecting facility.Design  Multiple indirect prevalence estimation methods.Setting  Kings Cross, Sydney, Australia.Participants  IDUs residing in Kings Cross area postcodes recorded in surveillance data of the Sydney Medically Supervised Injecting Centre (MSIC) between November 2001 and October 2002.Measurements  Two closed and one open capture–recapture (CRC) models (Poisson regression, truncated Poisson and Jolly–Seber, respectively) were fitted to the observed data. Multiplier estimates were derived from opioid overdose mortality data and a cross‐sectional survey of needle and syringe programme attendees. MSIC client injection frequency and the number of needles and syringes distributed in the study area were used to estimate injection prevalence and injection coverage.Findings  From three convergent estimates, the mean estimated size of the IDU population aged 15–54 years was 1103 (range 877–1288), yielding a population prevalence of 3.6% (2.9–4.3%). Mean IDU coverage was 70.7% (range 59.1–86.7%) and the mean adjusted injection coverage was 8.8% (range 7.3–10.8%). Approximately 11.3% of the total IDU population were estimated to be new entrants to the population per month.Conclusions  Credible local area IDU prevalence estimates using MSIC surveillance data were obtained. MSIC appears to achieve high coverage of the local IDU population, although only an estimated one in 10 injections occurs at MSIC. Future prevalence estimation efforts should incorporate open models to capture the dynamic nature of IDU populations.
Acute liver failure following therapeutic paracetamol administration in patients with muscular dystrophies
Authors: Pearce B. , Grant I. S. 
Published: July 2008

SummaryClinically significant liver damage in patients taking therapeutic doses of paracetamol is very rare. We report two cases of fulminant hepatic failure caused by therapeutic (4 g.day−1) paracetamol administration on our Intensive Care Unit. Both patients had a muscular dystrophy and presented with a chest infection on a background of endstage neuromuscular respiratory failure. We also noted one further similar case in the literature and suggest a relationship between muscular dystrophy and paracetamol‐induced hepatotoxicity. In this report we discuss in detail possible mechanisms that may account for this apparent association, which include altered pharmacokinetics, reduced glutathione stores, malnutrition and hypoxic injury.
Patients' understanding of the reasons for starting and discontinuing inhaled corticosteroids
Authors: Menckeberg Tanja T. , Bouvy Marcel L. , Bracke Madelon , Hugtenburg Jacqueline G. , Lammers Jan‐Willem , Raaijmakers Jan A. M. 
Published: July 2008

WHAT IS ALREADY KNOWN ABOUT THIS SUBJECT • Early discontinuation of treatment in new users of inhaled corticosteroids (ICS) has been widely discussed, but patients' reasons for this have not been investigated in depth. WHAT THIS STUDY ADDS • In contrast to what might have been expected, the majority of patients mentioned a wide range of symptoms or conditions as the reason for the start of ICS therapy.• There is no clear indication for prescribing ICS in these patients.• Most of these conditions are self limiting and may be expected to be of short duration, consequently a decrease in symptoms was the main and justifiable reason for discontinuing ICS.• However, a non‐negligible proportion of patients has significant residual symptoms and seems to be in need of continuous ICS use.AIM Although early discontinuation of treatment in new users of inhaled corticosteroids (ICS) has been widely discussed, the reasons for stopping have not been investigated in depth. We aimed to describe reasons for discontinuation from a patient's perspective in relation to their experience of symptoms at the time of the investigation.METHODS A cross‐sectional study among new users that discontinued ICS use in the Netherlands was performed. Patients were interviewed by telephone, aiming to identify the symptoms for which they were prescribed ICS, the reasons for discontinuing treatment and the respiratory symptoms patients still experienced at the time of the survey. In addition, automated dispensing records of all patients were retrieved.RESULTS From 287 eligible patients, 230 (80.1%) were interviewed. Only 22 patients (9.6%) mentioned asthma as the reason for a first ICS prescription. A decrease in symptoms was the main reason for discontinuation (45%). Thirty patients (13%) reported clinically significant residual symptoms. These patients reported more seasonal variation of symptoms and were more often prescribed short‐acting β2‐agonists.CONCLUSIONS The majority of patients mentioned a wide range of symptoms and conditions, other than asthma or chronic obstructive pulmonary disease, as the reason for the start of ICS therapy. Most of these conditions may be expected to be of short duration. Not surprisingly a decrease in symptoms was the main, and justifiable, reason for discontinuing ICS. However, a non‐negligible proportion of patients reported residual symptoms that suggest the need of continued ICS use. Physicians and pharmacists could cooperate in identifying those patients for which ICS are really indicated and motivate them to continue the use of ICS.
Evaluation of minimal residual disease in multiple myeloma patients by fluorescent‐polymerase chain reaction: the prognostic impact of achieving molecular response
Authors: Martínez‐Sánchez Pilar , Montejano Laura , Sarasquete Maria Eugenia , García‐Sanz Ramón , Fernández‐Redondo Elena , Ayala Rosa , Montalbán María Angeles , Martínez Rafael , García Laraña José , Alegre Adrian , Hernández Belen , Lahuerta Juan José , Martínez‐López Joaquín 
Published: July 2008

SummaryThis study aimed to standardize a simple molecular method for evaluating the response to treatment in multiple myeloma (MM) patients after high dose chemotherapy. Fifty three patients enrolled in the GEM2000 protocol were studied for minimal residual disease (MRD) using both fluorescent‐polymerase chain reaction (F‐PCR) and flow cytometry. Most patients had achieved complete remission or very good response after autologous stem cell transplantation. The molecular analysis of immunoglobulin gene rearrangements at diagnosis and during the follow‐up was carried out by F‐PCR according to the Biomed‐2 protocols. F‐PCR could be used in 91% of the patients and the results were similar to flow cytometry. F‐PCR was able to identify a group of patients with a better prognosis [progression‐free survival (PFS) 67·86% in patients with negative F‐PCR vs. 28%; P = 0·001], even amongst patients who achieved a complete response with negative immunofixation (PFS 75% vs. 25%; P = 0·002). Multivariate analysis identified the F‐PCR result as the only variable to show a prognostic value when PFS was analysed. F‐PCR of DHJ and light chain rearrangements of immunoglobulin genes is a feasible method for evaluating MRD in MM patients after intensive therapy. Achieving molecular response by F‐PCR shows prognostic value.
Recent advances in the understanding of inherited sideroblastic anaemia
Authors: Camaschella Clara 
Published: July 2008

SummarySideroblastic anaemia includes a heterogeneous group of rare conditions, characterized by decreased haem synthesis and mitochondrial iron overload, which are diagnosed by the presence of ringed sideroblasts in the bone marrow aspirate. The most frequent form is X‐linked sideroblastic anaemia, caused by mutations of delta‐aminolevulinic acid synthase 2 (ALAS2), the enzyme that catalyses the first and regulatory step of haem synthesis in erythroid precursors and is post‐transcriptionally controlled by the iron regulatory proteins. Impaired haem production causes variable degrees of anaemia and mitochondrial iron accumulation as ringed sideroblasts. The heterogeneity and complexity of sideroblastic anaemia is explained by an increasing number of recognized molecular defects. New forms have been recognized as being linked to the deficient function of mitochondrial proteins involved in iron–sulphur cluster biogenesis, such as ABCB7 and GLRX5, which are extremely rare but represent important biological models. Local mitochondrial iron overload is present in all sideroblastic anaemias, whereas systemic iron overload occurs only in the forms because of primary or secondary deficiency of ALAS2.