Activation of gene expression during hypersensitive response (HR) induced by auxin in the grapevine rootstock cultivar 'Börner'

The cultivar ‘Börner’ is one of the very few grape phylloxera (Daktulosphaira vitifoliae Fitch) resistant rootstocks commercially available. In contrast to tolerant or sensitive rootstocks, ‘Börner’ roots react to grape phylloxera attack with a hypersensitive response leading to necroses around the puncture sites. In this study, we identified genes differentially up-regulated during the HR. HR was chemically induced in root cells by the application of indol-acetic-acid (IAA). After a cDNA subtraction of induced and non-induced material, the subtracted cDNA-samples were also hybridized to Arabidopsis microarray chips to identify differentially expressed candidate genes. The microarray data were analyzed and differentially expressed genes were grouped into different functional categories, e.g. signal transduction pathways, transcription factors, defence associated genes. Primers were designed to target genes of interest putatively involved in the HR. So far, 38 ESTs induced in ‘Börner’ roots undergoing a HR have been sequenced and annotated.

Flowering Phenology of Six Seasonal-Flowering Strawberry Cultivars in a Coordinated European Study

The flowering phenology of six genetically distant strawberry cultivars (‘Candonga®’ (ES), ‘Clery’ (IT), ‘Florence’ (UK), ‘Frida’ (NO), ‘Gariguette’ (FR), and ‘Sonata’ (NL)) was studied for 3 years in relation to climatic parameters in open-field cultivation at three locations (Norway, Poland, Germany) and in soil-less cultivation at two locations (Italy, and France), covering a distance of 16 degrees of latitude. This proved to be a useful approach for unravelling the climatic adaptation and plasticity of strawberry genotypes and their suitability both for profitable cultivation and as a breeding pedigree. Despite the intercorrelated character of the climatic variables, the observed results highlight the importance of global radiation as a powerful modifying phenological factor in strawberry. Generally, early flower initiation was associated with elevated temperature and global radiation. ‘Frida’ revealed the highest dependency on global radiation for flower initiation, while ‘Sonata’ was least affected by temperature and radiation. In general, temperature and global radiation in periods both preceding and following flower initiation had a stronger positive effect on the number of flowers than on crowns, especially under open-field conditions. The influence of these factors was highly variable across the cultivars: ‘Clery’, ‘Florence’, and ‘Gariguette’ were most affected, while ‘Frida’ was least influenced.publishedVersio

Thioredoxin-regulated β-amylase (BAM1) triggers diurnal starch degradation in guard cells, and in mesophyll cells under osmotic stress

BAM1 is a plastid-targeted β-amylase of Arabidopsis thaliana specifically activated by reducing conditions. Among eight different chloroplast thioredoxin isoforms, thioredoxin f1 was the most efficient redox mediator, followed by thioredoxins m1, m2, y1, y2, and m4. Plastid-localized NADPH-thioredoxin reductase (NTRC) was also able partially to restore the activity of oxidized BAM1. Promoter activity of BAM1 was studied by reporter gene expression (GUS and YFP) in Arabidopsis transgenic plants. In young (non-flowering) plants, BAM1 was expressed both in leaves and roots, but expression in leaves was mainly restricted to guard cells. Compared with wild-type plants, bam1 knockout mutants were characterized by having more starch in illuminated guard cells and reduced stomata opening, suggesting that thioredoxin-regulated BAM1 plays a role in diurnal starch degradation which sustains stomata opening. Besides guard cells, BAM1 appears in mesophyll cells of young plants as a result of a strongly induced gene expression under osmotic stress, which is paralleled by an increase in total β-amylase activity together with its redox-sensitive fraction. Osmotic stress impairs the rate of diurnal starch accumulation in leaves of wild-type plants, but has no effect on starch accumulation in bam1 mutants. It is proposed that thioredoxin-regulated BAM1 activates a starch degradation pathway in illuminated mesophyll cells upon osmotic stress, similar to the diurnal pathway of starch degradation in guard cells that is also dependent on thioredoxin-regulated BAM1

Two negative cis-regulatory regions involved in fruit-specific promoter activity from watermelon (Citrullus vulgaris S.)

A 1.8 kb 5′-flanking region of the large subunit of ADP-glucose pyrophosphorylase, isolated from watermelon (Citrullus vulgaris S.), has fruit-specific promoter activity in transgenic tomato plants. Two negative regulatory regions, from –986 to –959 and from –472 to –424, were identified in this promoter region by fine deletion analyses. Removal of both regions led to constitutive expression in epidermal cells. Gain-of-function experiments showed that these two regions were sufficient to inhibit RFP (red fluorescent protein) expression in transformed epidermal cells when fused to the cauliflower mosaic virus (CaMV) 35S minimal promoter. Gel mobility shift experiments demonstrated the presence of leaf nuclear factors that interact with these two elements. A TCCAAAA motif was identified in these two regions, as well as one in the reverse orientation, which was confirmed to be a novel specific cis-element. A quantitative β-glucuronidase (GUS) activity assay of stable transgenic tomato plants showed that the activities of chimeric promoters harbouring only one of the two cis-elements, or both, were ∼10-fold higher in fruits than in leaves. These data confirm that the TCCAAAA motif functions as a fruit-specific element by inhibiting gene expression in leaves

The nature of floral signals in Arabidopsis. I. Photosynthesis and a far-red photoresponse independently regulate flowering by increasing expression of FLOWERING LOCUS T (FT)

Arabidopsis flowers in long day (LD) in response to signals transported from the photoinduced leaf to the shoot apex. These LD signals may include protein of the gene FLOWERING LOCUS T (FT) while in short day (SD) with its slower flowering, signalling may involve sucrose and gibberellin. Here, it is shown that after 5 weeks growth in SD, a single LD up-regulated leaf blade expression of FT and CONSTANS (CO) within 4–8 h, and flowers were visible within 2–3 weeks. Plants kept in SDs were still vegetative 7 weeks later. This LD response was blocked in ft-1 and a co mutant. Exposure to different LD light intensities and spectral qualities showed that two LD photoresponses are important for up-regulation of FT and for flowering. Phytochrome is effective at a low intensity from far-red (FR)-rich incandescent lamps. Independently, photosynthesis is active in an LD at a high intensity from red (R)-rich fluorescent lamps. The photosynthetic role of a single high light LD is demonstrated here by the blocking of the flowering and FT increase on removal of atmospheric CO2 or by decreasing the LD light intensity by 10-fold. These conditions also reduced leaf blade sucrose content and photosynthetic gene expression. An SD light integral matching that in a single LD was not effective for flowering, although there was reasonable FT-independent flowering after 12 SD at high light. While a single photosynthetic LD strongly amplified FT expression, the ability to respond to the LD required an additional but unidentified photoresponse. The implications of these findings for studies with mutants and for flowering in natural conditions are discussed

Patient safety in Dutch primary care: a study protocol

<p>Abstract</p> <p>Background</p> <p>Insight into the frequency and seriousness of potentially unsafe situations may be the first step towards improving patient safety. Most patient safety attention has been paid to patient safety in hospitals. However, in many countries, patients receive most of their healthcare in primary care settings. There is little concrete information about patient safety in primary care in the Netherlands. The overall aim of this study was to provide insight into the current patient safety issues in Dutch general practices, out-of-hours primary care centres, general dental practices, midwifery practices, and allied healthcare practices. The objectives of this study are: to determine the frequency, type, impact, and causes of incidents found in the records of primary care patients; to determine the type, impact, and causes of incidents reported by Dutch healthcare professionals; and to provide insight into patient safety management in primary care practices.</p> <p>Design and methods</p> <p>The study consists of three parts: a retrospective patient record study of 1,000 records per practice type was conducted to determine the frequency, type, impact, and causes of incidents found in the records of primary care patients (objective one); a prospective component concerns an incident-reporting study in each of the participating practices, during two successive weeks, to determine the type, impact, and causes of incidents reported by Dutch healthcare professionals (objective two); to provide insight into patient safety management in Dutch primary care practices (objective three), we surveyed organizational and cultural items relating to patient safety. We analysed the incidents found in the retrospective patient record study and the prospective incident-reporting study by type of incident, causes (Eindhoven Classification Model), actual harm (severity-of-outcome domain of the International Taxonomy of Medical Errors in Primary Care), and probability of severe harm or death.</p> <p>Discussion</p> <p>To estimate the frequency of incidents was difficult. Much depended on the accuracy of the patient records and the professionals' consensus about which types of adverse events have to be recognized as incidents.</p

Multiple circadian clock outputs regulate diel turnover of carbon and nitrogen reserves

Plants accumulate reserves in the daytime to support growth at night. Circadian regulation of diel reserve turnover was investigated by profiling starch, sugars, glucose 6‐phosphate, organic acids, and amino acids during a light–dark cycle and after transfer to continuous light in Arabidopsis wild types and in mutants lacking dawn (lhy cca1), morning (prr7 prr9), dusk (toc1, gi), or evening (elf3) clock components. The metabolite time series were integrated with published time series for circadian clock transcripts to identify circadian outputs that regulate central metabolism. (a) Starch accumulation was slower in elf3 and prr7 prr9. It is proposed that ELF3 positively regulates starch accumulation. (b) Reducing sugars were high early in the T‐cycle in elf3, revealing that ELF3 negatively regulates sucrose recycling. (c) The pattern of starch mobilization was modified in all five mutants. A model is proposed in which dawn and dusk/evening components interact to pace degradation to anticipated dawn. (d) An endogenous oscillation of glucose 6‐phosphate revealed that the clock buffers metabolism against the large influx of carbon from photosynthesis. (e) Low levels of organic and amino acids in lhy cca1 and high levels in prr7 prr9 provide evidence that the dawn components positively regulate the accumulation of amino acid reserves.Research was supported by the Max Planck Society and European Union (Seventh Framework Programme, TiMet, no. 245143), by the Biotechnology and Biological Sciences Research Council (UK) in the form of an Institute Strategic Grant (BB/J004596/1) to the John Innes Centre, and by the John Innes Foundation. We are grateful to Karen Halliday for discussions about the EC‐independent function of ELF3