An NADH nitrate reductase gene copy appears to have been deleted in barley

Cultivated barley,Hordeum vulgare L., has a single NADH nitrate reductase (NR) gene while diploid wheat,Triticum monococcum, and cultivated hexaploid wheat,Triticum aestivum L., have two NADH NR genes. To determine whether the NADH NR gene was duplicated since the divergence ofTriticum fromHordeum or was deleted from barley, theT. Monococcum NADH NR gene heme-hinge regions were sequenced and compared with the barley NADH NR gene sequence. Sequence identity and phylogenetic analyses showed that one of theT. Monococcum NADH NR genes is more-closely related to the barley NADH NR gene than to the otherT. Monococcum NADH NR gene. The heme-hinge region of all three NR genes appeared to have evolved at a constant rate. These results suggest that the NADH NR gene duplicated before the divergence ofTriticum andHordeum and that a deletion resulted in the loss of one NADH NR gene from cultivated barley.     

Linkage Maps of Lowland and Upland Tetraploid Switchgrass Ecotypes

Switchgrass (Panicum virgatum L.) is a native perennial warm season (C₄) grass that has been identified as a promising species for bioenergy research and production. Consequently, biomass yield and feedstock quality improvements are high priorities for switchgrass research. The objective of this study was to develop a switchgrass genetic linkage map using a full-sib pseudo-testcross mapping population derived from a cross between two heterozygous genotypes selected from the lowland cultivar ‘Alamo’ (AP13) and the upland cultivar ‘Summer’ (VS16). The female parent (AP13) map consists of 515 loci in 18 linkage groups (LGs) and spans 1,733 cM. The male parent (VS16) map arranges 363 loci in 17 LGs and spans 1,508 cM. No obvious cause for the lack of one LG in VS16 could be identified. Comparative analyses between the AP13 and VS16 maps showed that the two major ecotypic classes of switchgrass have highly colinear maps with similar recombination rates, suggesting that chromosomal exchange between the two ecotypes should be able to occur freely. The AP13 and VS16 maps are also highly similar with respect to marker orders and recombination levels to previously published switchgrass maps. The genetic maps will be used to identify quantitative trait loci associated with biomass and quality traits. The AP13 genotype was used for the whole genome-sequencing project and the map will thus also provide a tool for the anchoring of the switchgrass genome assembly.     

Objectively Measured Walking Duration and Sedentary Behaviour and Four-Year Mortality in Older People

BackgroundPhysical activity is an important component of health. Recommendations based on sensor measurements are sparse in older people. The aim of this study was to analyse the effect of objectively measured walking and sedentary duration on four-year mortality in community-dwelling older people.MethodsBetween March 2009 and April 2010, physical activity of 1271 participants (≥65 years, 56.4% men) from Southern Germany was measured over one week using a thigh-worn uni-axial accelerometer (activPAL; PAL Technologies, Glasgow, Scotland). Mortality was assessed during a four-year follow-up. Cox-proportional-hazards models were used to estimate the associations between walking (including low to high intensity) and sedentary duration with mortality. Models were adjusted for age and sex, additional epidemiological variables, and selected biomarkers.ResultsAn inverse relationship between walking duration and mortality with a minimum risk for the 3rd quartile (102.2 to128.4 minutes walking daily) was found even after multivariate adjustment with HRs for quartiles 2 to 4 compared to quartile 1 of 0.45 (95%-CI: 0.26; 0.76), 0.18 (95%-CI: 0.08; 0.41), 0.39 (95%-CI: 0.19; 0.78), respectively. For sedentary duration an age- and sex-adjusted increased mortality risk was observed for the 4th quartile (daily sedentary duration ≥1137.2 min.) (HR 2.05, 95%-CI: 1.13; 3.73), which diminished, however, after full adjustment (HR 1.63, 95%-CI: 0.88; 3.02). Furthermore, our results suggest effect modification between walking and sedentary duration, such that in people with low walking duration a high sedentary duration was noted as an independent factor for increased mortality.ConclusionsIn summary, walking duration was clearly associated with four-year overall mortality in community-dwelling older people.     

P2X7-mediated ATP secretion is accompanied by depletion of cytosolic ATP

Purinergic Signalling - ATP and its metabolites are important extracellular signal transmitters acting on purinergic P2 and P1 receptors. Most cells can actively secrete ATP in response to a...     

Rice-barley synteny and its application to saturation mapping of the barley Rpg1 region.

In order to facilitate the map-based cloning of the barley stem rust resistance gene Rpg1, we have demonstrated a high degree of synteny at a micro level between the telomeric region of barley chromosome 1P and rice chromosome 6. We have also developed and applied a simple and efficient method for selecting useful probes from large insert genomic YAC and cosmid clones. The gene order within the most terminal 6.5 cM of barley chromosome 1P was compared with the most terminal 2.7 cM of rice chromosome 6. Nine rice probes, previously mapped in rice or isolated from YAC or cosmid clones from this region, were mapped in barley. All, except one, were in synteny with the rice gene order. The exception, probe Y617R, was duplicated in barley. One copy was located on a different chromosome and the other in a non-syntenic position on barley chromosome 1P. The barley probes from this region could not be mapped to rice, but two of them were inferred to be in a syntenic location based on their position on a rice YAC. This work demonstrates the utility of applying the results of genetic and physical mapping of the small genome cereal rice to map-based cloning of interesting genes from large genome relatives.     

Detergent solubilization of the interleukin 1 receptor

Interleukin 1 (IL 1) receptors were solubilized from membranes prepared from murine EL-4 thymoma cells with the zwitterionic detergent 3[(3-cholamidopropyl) dimethylammonio]-1-propanesulfonate (CHAPS). Binding of IL 1 to the solubilized receptor was detected by a polyethylene glycol (PEG) precipitation procedure. Concentrations of CHAPS from 4 to 8 mM were effective in solubilizing the IL 1 receptor. At 10 mM CHAPS, there was some loss in binding activity, whereas 2 mM CHAPS was completely ineffective in solubilizing the receptor. Detergent concentrations of 4 mM were routinely used. The solubilized receptor retains the ability to bind 125I-IL 1 in a specific and saturable manner. Scatchard analysis reveals a single type of high affinity binding site having an apparent dissociation constant (KD) of approximately 1.2 X 10(-10) M. Nearly identical KD values are observed for membrane fractions. There are approximately 400 to 500 fmol receptor/mg protein in the detergent extract, corresponding to a two- to threefold enrichment in the Bmax observed for membranes. There is no loss in receptor activity as determined by complete recovery of the total number of binding sites from membranes after solubilization. Binding kinetics show that apparent steady state for the solubilized receptor is reached after 60 min at 37 degrees C. The binding of 125I-IL 1 is essentially irreversible because relatively little bound ligand can be dissociated from the receptor on the addition of excess unlabeled IL 1 at 37 degrees C. Both human IL 1 alpha and IL 1 beta compete for binding of 125I-IL 1 to the soluble receptor, confirming that IL 1 alpha and IL 1 beta bind to the same receptor. Other recombinant proteins, including interferon-alpha A, interferon-gamma, and interleukin 2 have no inhibitory effect.     

Nanocrystalline Nickel and Cobalt Sulfides Formed by a Heavy Metal-Tolerant,Sulfate-Reducing Enrichment Culture

A soil enrichment culture of the sulfate-reducers Desulfosporosinus auripigmenti and Citrobacter freundii and of fermentative bacteria from a former uranium-mining site was studied for its metal retention potential by promoting metal sulfide precipitation. The culture could tolerate up to 30 mM Ni and 40 mM Co. XRD and TEM analyses revealed the formation of amorphous NiS together with nanocrystalline, metastable α-NiS, and nanocrystalline cobalt pentlandite. The α-NiS with a grain size of 5 nm shows probably an example of size-dependent phase stability and/or specific biomineralization precipitation paths. Detailed mineralogical characterizations are necessary to correctly assess the mineral inventory and thus metal bioavailability.     

Spatio-temporal propagation of Ca2+ signals by cyclic ADP-ribose in 3T3 cells stimulated via purinergic P2Y receptors

The role of cyclic ADP-ribose in the amplification of subcellular and global Ca2+ signaling upon stimulation of P2Y purinergic receptors was studied in 3T3 fibroblasts. Either (1) 3T3 fibroblasts (CD38- cells), (2) 3T3 fibroblasts preloaded by incubation with extracellular cyclic ADP-ribose (cADPR), (3) 3T3 fibroblasts microinjected with ryanodine, or (4) 3T3 fibroblasts transfected to express the ADP-ribosyl cyclase CD38 (CD38+ cells) were used. Both preincubation with cADPR and CD38 expression resulted in comparable intracellular amounts of cyclic ADP-ribose (42.3 +/- 5.2 and 50.5 +/- 8.0 pmol/mg protein). P2Y receptor stimulation of CD38- cells yielded a small increase of intracellular Ca2+ concentration and a much higher Ca2+ signal in CD38-transfected cells, in cADPR-preloaded cells, or in cells microinjected with ryanodine. Confocal Ca2+ imaging revealed that stimulation of ryanodine receptors by cADPR or ryanodine amplified localized pacemaker Ca2+ signals with properties resembling Ca2+ quarks and triggered the propagation of such localized signals from the plasma membrane toward the internal environment, thereby initiating a global Ca2+ wave.