Project description:Of all the essential nutrients, nitrogen is the one most often limiting for plant growth. Nitrogen can be taken up by plants in two ways. One possibility is through ammonium and nitrate, which are the predominate inorganic forms of nitrogen in soils. The second possibility is the uptake of air-born nitrogen through plant-associated mircoorganisms in root nodules. The majority of plants able to form such nitrogen-fixing root nodules are in the legume family Fabaceae. Here we present a third possibility – a new pathway, termed as nitric oxide (NO)-fixation pathway, which allows plants to fix atmospheric NO and to use it for better growth and development. We identified non-symbiotic hemoglobin class 1 (AtGLB1) and class 2 (AtGLB2) as key proteins of the NO-fixation pathway. In an NO enriched environment NO-fixation is enhanced considerably in plants overexpressing AtGLB1 or AtGLB2 genes. NO uptake resulted in four-fold higher nitrate levels in these plants compared to NO-treated wild-type plants. Correspondingly, the growth parameters like rosettes size and weight, vegetative shoot thickness and also seed yield were 25%, 40%, 30%, and 20% higher, respectively, in the overexpression lines in comparison to wild-type plants. Our results highlight the existence of a NO-fixing pathway in plants. We demonstrated that plant non-symbiotic hemoglobin proteins can fix atmospheric NO and convert it to nitrate, which is further introduced into the N-metabolism. We assume that our results might provide new insights into the field of crop science research and that the NO-fixation capability might serve as a new economically important breeding trait for enhancing biomass, fruit, and seed production. Modifying this pathway might be a promising approach for better and more environment-friendly supply of nitrogen. For example crop plant hemoglobin proteins could be improved for their NO-fixing capability and their expression levels could be increased.
Project description:Of all the essential nutrients, nitrogen is the one most often limiting for plant growth. Nitrogen can be taken up by plants in two ways. One possibility is through ammonium and nitrate, which are the predominate inorganic forms of nitrogen in soils. The second possibility is the uptake of air-born nitrogen through plant-associated mircoorganisms in root nodules. The majority of plants able to form such nitrogen-fixing root nodules are in the legume family Fabaceae. Here we present a third possibility M-bM-^@M-^S a new pathway, termed as nitric oxide (NO)-fixation pathway, which allows plants to fix atmospheric NO and to use it for better growth and development. We identified non-symbiotic hemoglobin class 1 (AtGLB1) and class 2 (AtGLB2) as key proteins of the NO-fixation pathway. In an NO enriched environment NO-fixation is enhanced considerably in plants overexpressing AtGLB1 or AtGLB2 genes. NO uptake resulted in four-fold higher nitrate levels in these plants compared to NO-treated wild-type plants. Correspondingly, the growth parameters like rosettes size and weight, vegetative shoot thickness and also seed yield were 25%, 40%, 30%, and 20% higher, respectively, in the overexpression lines in comparison to wild-type plants. Our results highlight the existence of a NO-fixing pathway in plants. We demonstrated that plant non-symbiotic hemoglobin proteins can fix atmospheric NO and convert it to nitrate, which is further introduced into the N-metabolism. We assume that our results might provide new insights into the field of crop science research and that the NO-fixation capability might serve as a new economically important breeding trait for enhancing biomass, fruit, and seed production. Modifying this pathway might be a promising approach for better and more environment-friendly supply of nitrogen. For example crop plant hemoglobin proteins could be improved for their NO-fixing capability and their expression levels could be increased. WT-Arabidopsis thaliana plants were fumigated with Ambient NO and 3 ppm NO air in three completely independent biological experiments. Total RNA was isolated from four-week old rosette leaves of these plants to determine the gene expression signature of each samples using Agilent one-color microarray. Differences in the gene expression signatures between Ambient NO and 3 ppm NO treated samples were analyzed to see the effect of NO fumigation on the WT Arabidopsis plants at the transcript level.
Project description:The diazotrophic bacterium Rhodobacter capsulatus synthesizes a molybdenum nitrogenase and an alternative iron-only nitrogenase, enabling growth with molecular dinitrogen as sole nitrogen source. Regulation of nitrogen fixation was analyzed by proteome profiling of wild-type and mutant strains lacking the transcriptional regulators NifA, AnfA, and MopAB.
Project description:Cadmium treatment induces slow but long lasting nitric oxide production in plant tissues. This NO production can be suppressed using the commonly used Nitric Oxide Synthase inhibitor L-NAME. This inhibitor tends to partially alleviate Cd toxicity. This effect is correlated with a strong diminution of Cd content in roots of plants treated both with Cd and L-NAME compared to roots from plants treated with Cd only. The main goal of this study is the identification of transcriptionnal changes caused by Cd-induced nitric oxide, and that could potentially result in enhanced Cd root accumulation.
Project description:Non-symbiotic hemoglobins are ubiquitously expressed proteins known to interact with nitric oxide, an inhibitor of mitochondrial respiration and an important signalling component. We evaluated the underlying molecular mechanisms of AtHb1 (also referred as AtGLB1 or AHb1) function, its effects on stress response and the interplay with nitric oxide. For this purpose, AtHb1 was overexpressed in Arabidopsis thaliana under control of the seed-specific promoter LeB4. We performed comparative transcriptome analysis of developing siliques from wild type (WT, Col-0) and transgenic plants subjected to control and moderate hypoxic conditions. The experimental design was used to analyze the underlying molecular mechanisms of AtHb1 function and to assess differences in the hypoxic response between WT and AtHb1-overexpressing seeds/siliques. Hypoxic and control (normoxic) treatment was carried out with complete transgenic (T3) and WT plants 45 days after germination (DPG). Plants were aerated with a gas mixture containing 10.5% oxygen or ambient gas containing 21% oxygen for control samples in darkness. After one hour, plants were decapitated, immediately frozen in liquid nitrogen, and siliques were dissected in liquid nitrogen for RNA extraction and hybridization to Affymetrix microarrays. Hypoxic and control treatment runs with WT and AtHb1-overexpressing plants were repeated twice to provide 3 biological replicates (total number of samples = 12).
Project description:RATIONALE: Chemoprevention is the use of certain drugs to keep cancer from forming, growing, or coming back. The use of nitric oxide-releasing acetylsalicyclic acid may prevent colorectal cancer.
PURPOSE: This randomized phase I trial is studying the side effects and best dose of nitric oxide-releasing acetylsalicyclic acid in preventing colorectal cancer in patients at high risk of colorectal cancer.
Project description:The effectiveness of new cancer therapies such as checkpoint blockade and adoptive cell transfer of activated anti-tumor T cells requires overcoming immunosuppressive tumor microenvironments. We found that the activation of tumor-infiltrating myeloid cells to produce local nitric oxide is a prerequisite for adoptively transferred CD8+ cytotoxic T cells to destroy tumors. These myeloid cells are phenotypically similar to ‘Tip-DCs’ or nitric oxide- and TNFα-producing dendritic cells. The nitric oxide-dependent killing was tempered by coincident arginase 1 expression, which competes with iNOS for arginine, the substrate for nitric oxide production. Depletion of immunosuppressive CSF-1R-dependent arginase 1+ myeloid cells enhanced nitric oxide-dependent tumor killing. Tumor killing via iNOS was independent of the microbiota but dependent on the CD40-CD40L pathway and, in part, lymphotoxin alpha. We extended our findings in mice to uncover a strong correlation between iNOS, CD40 and TNF expression and survival in colorectal cancer patients. Our results identify a network of anti-tumor targets to boost the efficacy of cancer immunotherapies.
Project description:Aerobic methanotrophic bacteria can use methane as their sole energy source. The discovery of ‘Ca. Methylacidiphilum fumariolicum’ strain SolV and other verrucomicrobial methanotrophs has revealed that the ability of bacteria to oxidize CH4 is much more diverse than has previously been assumed in terms of ecology, phylogeny and physiology. A remarkable characteristic of the methane-oxidizing Verrucomicrobia is their extremely acidophilic phenotype, growing even below pH 1. In this study we used RNA-Seq to analyze the metabolic regulation of ‘Ca. M. fumariolicum’ SolV cells growing at μmax in batch culture or under nitrogen fixing or oxygen limited conditions in chemostats, all at pH 2. The analysis showed that two of the three pmoCAB operons each encoding particulate methane monoxygenases were differentially expressed, probably regulated by the available oxygen. The hydrogen produced during N2 fixation is apparently recycled as demonstrated by the upregulation of the genes encoding a Ni/Fe-dependent hydrogenase. These hydrogenase genes were also upregulated under low oxygen conditions. Handling of nitrosative stress was shown by the expression of the nitric oxide reductase encoding genes norB and norC under all conditions tested, the upregulation of nitrite reductase nirK under oxygen limitation and of hydroxylamine oxidoreductase hao in the presence of ammonium. Unraveling the gene regulation of carbon and nitrogen metabolism helps to understand the underlying physiological adaptations of strain SolV in view of the harsh conditions of its natural ecosystem.
Project description:rs05-08_no - no - Cadmium treatment induces slow but long lasting nitric oxide production in plant tissues. This NO production can be suppressed using the commonly used Nitric Oxide Synthase inhibitor L-NAME. This inhibitor tends to partially alleviate Cd toxicity. This effect is correlated with a strong diminution of Cd content in roots of plants treated both with Cd and L-NAME compared to roots from plants treated with Cd only. The main goal of this study is the identification of transcriptionnal changes caused by Cd-induced nitric oxide, and that could potentially result in enhanced Cd root accumulation. - Germination of surface sterilized seeds was performed on solid MS/2 medium. After 2 weeks, the young plantlets were placed on sand for an additional 3 week period in a controlled environment (8 h photoperiod of 300 umol m-2 s-1, 22degreeC, 70% relative humidity) . Plants of similar rosette diameters were then transfered to an hydroponic culture using the nutritive solution described in [Gravot et al. 2004 FEBS Letters 561:22-28] for an acclimation of 3 days. L-NAME and cadmium treatments were respectively started 1 and 2 hours after the start of illumination and root tissues were collected 24 h later and immediatly frozen in liquid nitrogen before RNA extraction. Treatments were : 1 : control 2 : L-NAME 5 mM 3 : Cd 30 uM 4 : L-NAME 5mM + Cd 30 uM 5 : Cd 15 uM For each condition, 3 independent biological repetitions were conducted. RNA were extracted separatly and checked for quality before pooling together. Keywords: treated vs untreated comparison