114 research outputs found
Diethylpyrocarbonate, a Histidine Selective Reagent, Causes Structural Alteration of Rat Ovarian LH/hCG Receptor
Abstract. Treatment of íat ovanan membiane-bound and Tnton X-100 solubilized LH/hCG íeceptoi with a histidme-specific icagent diethylpyiocaibonate (DEPC) íesulted m macti\ation of the ability of the íeccptoi to bind hCG The paitial íe-veisibihťv of this inhibition by hydioxylamme domonstiated that histidine losidues aie involved m hCG-ieceptoi binding Fhioies(ence quenching expeiiments nidi cated that DEPC did not change the accessibihty of fiuoiophoies foi aciylamide Alterations of quenchiiig íate goneially suggest exposuie of tivptophanvl íesidues Modification of histidjl íesidues was connected with an alteiation of the physical state of ovanan membianes Membiane lipid iigicht\ was dec leased aftei DEPC íe action Theimal peitmbation techniques won used to monitoi stnutuial (lianges m the ícceptoi clue to the action of DEPC on membianes Heat mactnation of hCG-bmdmg sites demonstiated that theie was a signmc ant destabilization of the LH/hCG íeeeptoi stmctuie when the inembianes weio tieated with DEPC Thei mal dcstabih/ation pioduced h\ 5 mmol/1 DEPC ( ausc d a dec lease m T^, 0 \ allies bv about 12°C These íesults suggest that histidine lesidues aie lo< ated at the binding sites of the íeeeptoi and that they aie also imohed m alteiations of membiane piotems the stiuctuial mtcgiitv of w Inch sec ondanly influences the ae ce^ssibihty oi the LH/hCG íeceptoi Key words: Diethv lpyiocaibonate LH/hCG nceptois Theimal mat tnation Fluoiesc enc e polaii/atio
Compared to conventional, ecological intensive management promotes beneficial proteolytic soil microbial communities for agro-ecosystem functioning under climate change-induced rain regimes
Projected climate change and rainfall variability will affect soil microbial communities, biogeochemical cycling and agriculture. Nitrogen (N) is the most limiting nutrient in agroecosystems and its cycling and availability is highly dependent on microbial driven processes. In agroecosystems, hydrolysis of organic nitrogen (N) is an important step in controlling soil N availability. We analyzed the effect of management (ecological intensive vs. conventional intensive) on N-cycling processes and involved microbial communities under climate change-induced rain regimes. Terrestrial model ecosystems originating from agroecosystems across Europe were subjected to four different rain regimes for 263 days. Using structural equation modelling we identified direct impacts of rain regimes on N-cycling processes, whereas N-related microbial communities were more resistant. In addition to rain regimes, management indirectly affected N-cycling processes via modifications of N-related microbial community composition. Ecological intensive management promoted a beneficial N-related microbial community composition involved in N-cycling processes under climate change-induced rain regimes. Exploratory analyses identified phosphorus-associated litter properties as possible drivers for the observed management effects on N-related microbial community composition. This work provides novel insights into mechanisms controlling agro-ecosystem functioning under climate change
NO, ROS, and cell death associated with caspase-like activity increase in stress-induced microspore embryogenesis of barley
Under specific stress treatments (cold, starvation), in vitro microspores can be induced to deviate from their gametophytic development and switch to embryogenesis, forming haploid embryos and homozygous breeding lines in a short period of time. The inductive stress produces reactive oxygen species (ROS) and nitric oxide (NO), signalling molecules mediating cellular responses, and cell death, modifying the embryogenic microspore response and therefore, the efficiency of the process. This work analysed cell death, caspase 3-like activity, and ROS and NO production (using fluorescence probes and confocal analysis) after inductive stress in barley microspore cultures and embryogenic suspension cultures, as an in vitro system which permitted easy handling for comparison. There was an increase in caspase 3-like activity and cell death after stress treatment in microspore and suspension cultures, while ROS increased in non-induced microspores and suspension cultures. Treatments of the cultures with a caspase 3 inhibitor, DEVD-CHO, significantly reduced the cell death percentages. Stress-treated embryogenic suspension cultures exhibited high NO signals and cell death, while treatment with S-nitrosoglutathione (NO donor) in control suspension cultures resulted in even higher cell death. In contrast, in microspore cultures, NO production was detected after stress, and, in the case of 4-day microspore cultures, in embryogenic microspores accompanying the initiation of cell divisions. Subsequent treatments of stress-treated microspore cultures with ROS and NO scavengers resulted in a decreasing cell death during the early stages, but later they produced a delay in embryo development as well as a decrease in the percentage of embryogenesis in microspores. Results showed that the ROS increase was involved in the stress-induced programmed cell death occurring at early stages in both non-induced microspores and embryogenic suspension cultures; whereas NO played a dual role after stress in the two in vitro systems, one involved in programmed cell death in embryogenic suspension cultures and the other in the initiation of cell division leading to embryogenesis in reprogrammed microspores
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The role of macro-aggregation in regulating enzymatic depolymerization of soil organic nitrogen
Extracellular enzymatic depolymerization of polymeric organic nitrogen (PON) is a rate-limiting step in N mineralization. However, enzymatic accessibility to PON might be regulated by physical occlusion of the PON resulting from the architectural packing of soil minerals during aggregate formation. To examine the extent to which enzymatic accessibility to PON is regulated by soil aggregation, we put forward a new approach involving the comparison of relationships between potential N depolymerase activity (protease and β-glucosaminidase; as an estimate of the potential to produce depolymerized products) and net N mineralization (as a bioassay for actual low molecular weight dissolved ON production) in aggregated and corresponding disaggregated soil. Soils were sampled from grassland (GL) and arable land (AL), separated by dry sieving into fractions (4.75-2, 2-0.25 and 0.25-0.063 mm) and fractions mixed (4:4:1 by mass, respectively) to obtain constructed aggregated soils. Corresponding disaggregated soils were prepared using a mortar and pestle. This procedure mainly disrupted the 4.75-2 mm (large macro-aggregate) fraction. Disaggregation significantly promoted (p<0.05) net N mineralization rates by 1.3 times and 1.5 times in GL and AL soil, respectively. When net N mineralization - potential N depolymerase relationships for GL were examined, a greater slope parameter for disaggregated compared to aggregated soil (p=0.001; ANCOVA) quantified the extent to which this promoted N mineralization could be attributed to disruption of macroaggregate-increased enzymatic accessibility to PON. For AL, which had low protease and β-glucosaminidase activity, promoted N mineralization rate could not be attributed to increased protease + β-glucosaminidase accessibility to PON reflecting a possible role for other N depolymerases and/or osmolyte/lysate effects. By proposing how differences between mineralization-depolymerase relationships for soils differing in aggregation status might, with assumptions, be interpreted to identify the role of physical occlusion in protection of PON, we give new insight on the regulation of enzymatic depolymerization by physical protection through macro-aggregation for soils from contrasting land use
Species-Specific Expansion and Molecular Evolution of the 3-hydroxy-3-methylglutaryl Coenzyme A Reductase (HMGR) Gene Family in Plants
Kazakh dandelion (Taraxacum kok-saghyz, Tk) is a rubber-producing plant currently being investigated as a source of natural rubber for industrial applications. Like many other isoprenoids, rubber is a downstream product of the mevalonate pathway. The 3-hydroxy-3-methylglutaryl-CoA reductase (HMGR) enzyme catalyzes the conversion of 3-hydroxy-3-methylglutaryl-CoA to mevalonic acid, a key regulatory step in the MVA pathway. Such regulated steps provide targets for increases in isoprenoid and rubber contents via genetic engineering to increase enzyme activities. In this study, we identify a TkHMGR1 gene that is highly expressed in the roots of Kazakh dandelion, the main tissue where rubber is synthesized and stored. This finding paves the way for further molecular and genetic studies of the TkHMGR1 gene, and its role in rubber biosynthesis in Tk and other rubber-producing plants
A Polyadenylation Factor Subunit Implicated in Regulating Oxidative Signaling in Arabidopsis thaliana
BACKGROUND: Plants respond to many unfavorable environmental conditions via signaling mediated by altered levels of various reactive oxygen species (ROS). To gain additional insight into oxidative signaling responses, Arabidopsis mutants that exhibited tolerance to oxidative stress were isolated. We describe herein the isolation and characterization of one such mutant, oxt6. METHODOLOGY/PRINCIPAL FINDINGS: The oxt6 mutation is due to the disruption of a complex gene (At1g30460) that encodes the Arabidopsis ortholog of the 30-kD subunit of the cleavage and polyadenylation specificity factor (CPSF30) as well as a larger, related 65-kD protein. Expression of mRNAs encoding Arabidopsis CPSF30 alone was able to restore wild-type growth and stress susceptibility to the oxt6 mutant. Transcriptional profiling and single gene expression studies show elevated constitutive expression of a subset of genes that encode proteins containing thioredoxin- and glutaredoxin-related domains in the oxt6 mutant, suggesting that stress can be ameliorated by these gene classes. Bulk poly(A) tail length was not seemingly affected in the oxt6 mutant, but poly(A) site selection was different, indicating a subtle effect on polyadenylation in the mutant. CONCLUSIONS/SIGNIFICANCE: These results implicate the Arabidopsis CPSF30 protein in the posttranscriptional control of the responses of plants to stress, and in particular to the expression of a set of genes that suffices to confer tolerance to oxidative stress
Fortunella margarita Transcriptional Reprogramming Triggered by Xanthomonas citri subsp. citri
<p>Abstract</p> <p>Background</p> <p>Citrus canker disease caused by the bacterial pathogen <it>Xanthomonas citri </it>subsp. <it>citri (</it>Xcc) <it>has </it>become endemic in areas where high temperature, rain, humidity, and windy conditions provide a favourable environment for the dissemination of the bacterium. Xcc is pathogenic on many commercial citrus varieties but appears to elicit an incompatible reaction on the citrus relative <it>Fortunella margarita </it>Swing (kumquat), in the form of a very distinct delayed necrotic response. We have developed subtractive libraries enriched in sequences expressed in kumquat leaves during both early and late stages of the disease. The isolated differentially expressed transcripts were subsequently sequenced. Our results demonstrate how the use of microarray expression profiling can help assign roles to previously uncharacterized genes and elucidate plant pathogenesis-response related mechanisms. This can be considered to be a case study in a citrus relative where high throughput technologies were utilized to understand defence mechanisms in <it>Fortunella </it>and citrus at the molecular level.</p> <p>Results</p> <p><b>cDNAs from sequenced kumquat libraries (ESTs) made from subtracted RNA populations, healthy vs. infected, were used to make this microarray</b>. Of 2054 selected genes on a customized array, 317 were differentially expressed (P < 0.05) in Xcc challenged kumquat plants compared to mock-inoculated ones. This study identified components of the incompatible interaction such as reactive oxygen species (ROS) and programmed cell death (PCD). Common defence mechanisms and a number of resistance genes were also identified. In addition, there were a considerable number of differentially regulated genes that had no homologues in the databases. This could be an indication of either a specialized set of genes employed by kumquat in response to canker disease or new defence mechanisms in citrus.</p> <p>Conclusion</p> <p>Functional categorization of kumquat Xcc-responsive genes revealed an enhanced defence-related metabolism as well as a number of resistant response-specific genes in the kumquat transcriptome in response to Xcc inoculation. Gene expression profile(s) were analyzed to assemble a comprehensive and inclusive image of the molecular interaction in the kumquat/Xcc system. This was done in order to elucidate molecular mechanisms associated with the development of the hypersensitive response phenotype in kumquat leaves. These data will be used to perform comparisons among citrus species to evaluate means to enhance the host immune responses against bacterial diseases.</p
Heterologous expression of the isopimaric acid pathway in Nicotiana benthamiana and the effect of N-terminal modifications of the involved cytochrome P450 enzyme
Isolation and characterization of Arabidopsis mutants with enhanced tolerance to oxidative stress
Jasmonic acid elicits oxidative defense and detoxification systems in Cucumis melo L. cells
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