Dioxygenase Activity Research Articles

Effects of Anti-inflammatory Medication on Indoleamine 2,3 Dioxygenase Activity

Objective: To study the effects of anti-inflammatory medication on Indoleamine 2,3 dioxygenase activity.
 Research Design: This was an investigational study.
 Methodology: Eighteen fully grown albino rats separated into control and two treated sets, both treated sets were given indomethacin (50mg/1000g) orally. For acute treatment first treated set was sacrificed after 3.5 hrs & for chronic treatment second set was sacrificed after 3 days. However, control set animals were given an equivalent amount of vehicle.
 Results: Outcomes shows that serum Indoleamine 2,3 dioxygenase (IDO) enzyme activity was suppressed after acute treatment while serum IDO activity were increased after chronic treatment however no significant effect was seen on brain IDO.
 Conclusion: It is concluded that indomethacin has not shown any significant effect on brain IDO. But inhibits serum IDO activity.

Transcriptomic Analysis Elaborates the Resistance Mechanism of Grapevine Rootstocks against Salt Stress.

Grapes are subject to a wide range of climatic conditions during their life cycle, but the use of rootstocks can effectively ameliorate the effects of abiotic stress. However, the tolerance mechanism of different grape rootstock varieties varies under various stresses, and systematic research on this aspect is limited. On the basis of previous research, transcriptome sequencing was performed on three tolerant grape rootstock varieties (3309C, 520A, 1103P) and three intolerant grape rootstock varieties (5BB, 101–14, Beta). In total, 56,478,468 clean reads were obtained. One hundred and ten genes only existed in all combinations during P1 with a downregulated trend, and 178 genes existed only in P1 of tolerant grape rootstock varieties. Salt treatment firstly affected the photosynthesis of leaves, and tolerant varieties weakened or even eliminated this effect through their own mechanisms in the later stage. Tolerant varieties mobilized a large number of MFs during the P2 stage, such as hydrolase activity, carboxypeptidase activity, and dioxygenase activity. Carbon metabolism was significantly enriched in P1, while circadian rhythm and flavonoid biosynthesis were only enriched in tolerant varieties. In the intolerant varieties, photosynthesis-related pathways were always the most significantly enriched. There were large differences in the gene expression of the main signal pathways related to salt stress in different varieties. Salt stress affected the expression of genes related to plant abiotic stress, biotic stress, transcription factors, hormones, and secondary metabolism. Tolerant varieties mobilized more bHLH, WRKY, and MYB transcription factors to respond to salt stress than intolerant varieties. In the tolerant rootstocks, SOS was co-expressed. Among these, SOS1 and SOS2 were upregulated, and the SOS3 and SOS5 components were downregulated. The genes of heat shock proteins and the phenylalanine pathway were upregulated in the tolerant varieties. These findings outline a tolerance mechanism model for rootstocks for coping with osmotic stress, providing important information for improving the resistance of grapes under global climate change.

Iron(III) complexes of pyrrolidine and piperidine appended tridentate 3N donor ligands as models for catechol dioxygenase enzymes

Six new mononuclear iron(III) complexes [Fe(L1)Cl3]-[Fe(L6)Cl3] bearing tridentate all-nitrogen donor ligands (where L1 = N-(pyridin-2-ylmethyl)-2-(pyrrolidin-1-yl)ethanamine, L2 = 2-(piperidin-1-yl)-N-(pyridin-2-ylmethyl)ethanamine, L3 = 1-(piperidin-2-yl)-N-(pyridin-2-ylmethyl)methanamine, L4 = N-((1-methyl-1H-imidazol-2-yl)methyl)-2-(pyrrolidin-1-yl)ethan-1-amine, L5 = N-((1-methyl-1H-imidazol-2-yl)methyl)-2-(piperidin-1-yl)ethan-1-amine), L6 = 1-(1-methyl-1H-imidazol-2-yl)-N-(piperidin-2-ylmethyl)methanamine) have been prepared, characterized and evaluated as the functional models for the carbon–carbon bond-cleaving catechol dioxygenase enzymes. The molecular structure of complexes 1 and 3 revealed a distorted octahedral geometry around iron(III) center with facial coordination of ligands via three nitrogen atoms and three chloride ions occupy the remaining sites of the octahedron. Iron(III) catecholate adducts were generated in situ by adding one equivalent of various catechols pretreated with two equivalent of triethylamine. The 3,5-di-tert-butylcatecholate adduct of the complexes 1–6 show two intense bands in visible region (540–900 nm) as the result of DBC2--to-iron(III) ligand-to-metal charge-transfer transitions in acetonitrile. During electrochemical analysis, all the complexes display FeIII→FeII redox couple in the similar potential range (−0.260 to −0.336 V). The dioxygenase activity of present complexes produces major amount of extradiol cleavage products (75–92%) over the intradiol cleavage products (4–14%) at moderate rate (0.561–1.744 × 10−2 M−1 s−1). Interestingly, the iron(III) complexes 3 and 6 with facially coordinating tridentate ligands with heterocyclic piperidine amine (NH) nitrogen donor moiety produce major amounts of extradiol-cleavage products (3, 89%; 6, 75%) along with only minor amounts of other product (10–24%) without any intradiol-cleavage products.

Improving Toluene Dioxygenase Activity for Ester‐Functionalized Substrates through Enzyme Engineering

AbstractRieske dioxygenases have a history of utility in organic synthesis, owing to their ability to catalyze the asymmetric dihydroxylation of aromatics to produce chiral diene‐diol metabolites. However, their utility as green‐chemical tools has been limited by steric and electronic constraints on their substrate scopes and their activity. Herein we report the rational engineering of a widely used Rieske dioxygenase, toluene dioxygenase (TDO), to improve the activity of this enzyme system for the dihydroxylation of a synthetically valuable substrate class for which the wild‐type enzyme possesses low activity, the ester‐functionalized aromatics. Through active site targeted mutagenesis and application of a recently reported high throughput screening platform, engineered TDO variants with significantly increased activity in the dihydroxylation of these valuable substrates were identified and characterized, revealing key active site residues that modulate the enzyme's activity and selectivity.

Additive and synergistic interactions of 4-hydroxyphenylpyruvate dioxygenase (HPPD) and photosystem II (PSII) inhibitors for the control of glyphosate-resistant horseweed (Conyza canadensis) in corn

AbstractGlyphosate-resistant (GR) horseweed [Conyza canadensis (L.) Cronquist; syn.: Erigeron canadensis L.] interference can substantially reduce corn (Zea mays L.) yield. The complementary activity of 4-hydroxyphenylpyruvate dioxygenase (HPPD) and photosystem II (PSII) inhibitors has been investigated for the control of several weed species, and in many cases has been synergistic; however, there is little information on the interaction of HPPD- and PSII-inhibiting herbicides for postemergence control of GR C. canadensis in corn. Four field trials were studied over 2 yr (2019, 2020) in Ontario, Canada, in commercial corn fields with natural infestations of GR C. canadensis to evaluate the level of GR C. canadensis control with three HPPD-inhibiting herbicides (mesotrione, tolpyralate, and topramezone) and three PSII-inhibiting herbicides (atrazine, bromoxynil, and bentazon) applied individually and in tank-mix combinations, and to document the interaction of the three HPPD inhibitors tank mixed with the three PSII inhibitors. Mesotrione, tolpyralate, and topramezone controlled GR C. canadensis 83%, 84%, and 72%, respectively, at 8 wk after application (WAA). Bromoxynil and bentazon controlled GR C. canadensis 71% and 79%, respectively, while atrazine provided only 31% control at 8 WAA. The joint application of atrazine, bromoxynil, or bentazon with mesotrione increased GR C. canadensis control from 83% to 100% at 8 WAA. Tolpyralate tank mixed with atrazine, bromoxynil, or bentazon controlled GR C. canadensis 96%, 98%, and 98%, respectively, which was comparable to the mesotrione tank mixes at 8 WAA. Topramezone plus atrazine, bromoxynil, or bentazon controlled GR C. canadensis 91%, 93%, and 95%, respectively, at 8 WAA. Interactions between HPPD and PSII inhibitors were synergistic for all combinations of mesotrione or tolpyralate with atrazine, bromoxynil, or bentazon. The interaction between topramezone and PSII inhibitors was additive. All nine tank mixes controlled GR C. canadensis >90%. This study concludes that bromoxynil or bentazon, instead of atrazine, can be co-applied with mesotrione, tolpyralate, or topramezone without compromising GR C. canadensis control in corn.

Increasing atmospheric CO2 differentially supports arsenite stress mitigating impact of arbuscular mycorrhizal fungi in wheat and soybean plants

Arbuscular mycorrhizal fungi (AMF) are beneficial for the plant growth under heavy metal stress. Such beneficial effect is improved by elevated CO2 (eCO2). However, the mechanisms by which eCO2 improves AMF symbiotic associations under arsenite (AsIII) toxicity are hardly studied. Herein, we compared these regulatory mechanisms in species from two agronomical important plant families – grasses (wheat) and legumes (soybean). AsIII decreased plant growth (i.e., 53.75 and 60.29% of wheat and soybean, respectively) and photosynthesis. It also increased photorespiration and oxidative injury in both species, but soybean was more sensitive to oxidative stress as indicated by higher H2O2 accumulation and oxidation of protein and lipid. eCO2 significantly improved AMF colonization by increasing auxin levels, which induced high carotenoid cleavage dioxygenase (CCDs) activity, particularly in soybean roots. The improved sugar metabolism in plant shoots by co-application of eCO2 and AsIII allocated more sugars to roots sequentially. Sugar accumulation in plant roots is further induced by AMF, resulting in more C skeletons to produce organic acids, which are effectively exudated into the soil to reduce AsIII uptake. Exposure to eCO2 reduced oxidative damage and this mitigation was stronger in soybean. This could be attributed to a greater reduction in photorespiration as well as a stronger antioxidant and detoxification defence systems. The grass/legume-specificity was supported by principal component analysis, which revealed that soybean was more affected by AsIII stress and more responsive to AMF and eCO2. This study provided a mechanistic understanding of the impact of AMF, eCO2 and their interaction on As-stressed grass and legume plants, allowing better practical strategies to mitigate AsIII phytotoxicity.

Te(IV) bioreduction in the sulfur autotrophic reactor: Performance, kinetics and synergistic mechanism

A laboratory-scale sulfur autotrophic reactor (SAR) was first constructed for treating tellurite [Te(IV)] wastewater. The SAR had excellent Te(IV) bioreduction efficiency (90-96%) at 5-30 mg/L and conformed to the First-order kinetic model. The Te(IV) bioreduction was elaborated deeply from extracellular polymeric substances (EPS) functions, microbial metabolic activity, key enzyme activity, microbial community succession and quorum sensing. Te(IV) stimulated the increase of redox substances in EPS and the improved cell membrane permeability led to the increase of electron transport system activity. Catalase and reduced nicotinamide adenine dinucleotide (NADH) alleviated the oxidative stress caused by Te(IV) toxicity to maintain metabolic activity. The increase of sulfur dioxygenase activity (SDO) suggested that more ATP produced by sulfur oxidation might provide energy for various physiological activities. Meanwhile, nitrate reductase (NAR), nitrite reductase (NIR) and sulfide: quinone oxidoreductase (SQR) played an active role in sulfur oxidation and Te(IV) bioreduction. Combined with the above results and dynamic succession of three functional microbial communities, a synergistic mechanism was proposed to explain the excellent performance of SAR. This work provided a promising strategy for Te(IV) wastewater treatment process and Te(IV) bioreduction mechanism.

Eltrombopag inhibits TET dioxygenase to contribute to hematopoietic stem cell expansion in aplastic anemia.

Eltrombopag, an FDA-approved non-peptidyl thrombopoietin receptor agonist, is clinically used for the treatment of aplastic anemia, a disease characterized by hematopoietic stem cell failure and pancytopenia, to improve platelet counts and stem cell function. Eltrombopag treatment results in a durable trilineage hematopoietic expansion in patients. Some of the eltrombopag hematopoietic activity has been attributed to its off-target effects, including iron chelation properties. However, the mechanism of action for its full spectrum of clinical effects is still poorly understood. Here, we report that eltrombopag bound to the TET2 catalytic domain and inhibited its dioxygenase activity, which was independent of its role as an iron chelator. The DNA demethylating enzyme TET2, essential for hematopoietic stem cell differentiation and lineage commitment, is frequently mutated in myeloid malignancies. Eltrombopag treatment expanded TET2-proficient normal hematopoietic stem and progenitor cells, in part because of its ability to mimic loss of TET2 with simultaneous thrombopoietin receptor activation. On the contrary, TET inhibition in TET2 mutant malignant myeloid cells prevented neoplastic clonal evolution in vitro and in vivo. This mechanism of action may offer a restorative therapeutic index and provide a scientific rationale to treat selected patients with TET2 mutant–associated or TET deficiency–associated myeloid malignancies.

In silico analysis of Leishmania proteomes and protein-protein interaction network: Prioritizing therapeutic targets and drugs for repurposing to treat leishmaniasis

Leishmaniasis is a serious world health problem and its current therapies have several limitations demanding to develop novel therapeutics for this disease. The present study aims to prioritize novel broad-spectrum targets using proteomics and protein-protein interaction network (PPIN) data for 11 Leishmania species. Proteome comparison and host non-homology analysis resulted in 3605 pathogen-specific conserved core proteins. Gene ontology analysis indicated their involvement in major molecular functions like DNA binding, transportation, dioxygenase, and catalytic activity. PPIN analysis of these core proteins identified eight hub proteins (viz., vesicle-trafficking protein (LBRM2903_190011800), ribosomal proteins S17 (LBRM2903_34004790) and L2 (LBRM2903_080008100), eukaryotic translation initiation factor 3 (LBRM2903_350086700), replication factor A (LBRM2903_150008000), U3 small nucleolar RNA-associated protein (LBRM2903_340025600), exonuclease (LBRM2903_200021800), and mitochondrial RNA ligase (LBRM2903_200074100)). Among the hub proteins, six were classified as drug targets and two as vaccine candidates. Further, druggability analysis indicated three hub proteins, namely eukaryotic translation initiation factor 3, ribosomal proteins S17 and L2 as druggable. Their three-dimensional structures were modelled and docked with the identified ligands (2-methylthio-N6-isopentenyl-adenosine-5′-monophosphate, artenimol and omacetaxine mepesuccinate). These ligands could be experimentally validated (in vitro and in vivo) and repurposed for the development of novel antileishmanial agents.

Degradation profile and dioxygenase activity of Carnobacterium gallinarum (mt350233) and Enterococcus faecalis (mt345788) isolated from landfill soil in Benin City

Landfill is a potential microbial hub of hydrocarbon (HC) degrading bacterial population owing to nutrient availability and continuous enrichment by organic materials. The degradation and dioxygenase activity of Carnobacterium gallinarum and Enterococcus faecalis isolated from landfill soil in Benin City was investigated. Soil samples were collected from a Government approved landfill in Benin City at a depth of 0 to 10 cm. Standard microbiological and molecular methods were followed for the isolation and characterization of bacterial population. Bacterial isolates were standardized for degradation using spectrophotometer optical density (OD) 0.08 at 600 nm (equivalent to 1×108cfu/ml). The gas chromatography with flame ionization detector (GC-FID) method was used to determine the total petroleum hydrocarbon (TPH) of waste engine oil (WEO) containing samples. The spectrophotometric methods were used for the assay of catechol 1,2-dioxygenase (C12O) and catechol 2,3-dioxygenase (C23O) activity. The result of the 16S rRNA analyses of the bacterial isolates confirmed the identity of Carnobacterium gallinarum as 98.71 % and Enterococcus faecalis as 100 %. The degradation profile of Carnobacterium gallinarum and Enterococcus faecalis at varied concentrations (20, 40, 60, 80 and 100 mg/L) of WEO showed significant decline in optical density (OD) values from day 1 to day 7. The decline in OD is an indication of utilization of Polycyclic Aromatic Hydrocarbons (PAHs) present in the WEO. The result of the gas Chromatography with Flame Ionization Detector (GC-FID analyses) of degraded WEO containing samples revealed the absence of low molecular weight and high molecular weight PAHs after the 7-days degradation study. The presence of HC degrading enzyme in the bacterial isolates was confirmed by the production of C12O and C23O as a result of the formation of cis, cis muconic acid and 2-hydroxymuconic semi-aldehyde respectively. This study revealed that the landfill soil is a potential natural microcosm for WEO degrading bacteria and possesses biologically active HC degrading bacteria population such as Carnobacterium gallinarum and Enterococcus faecalis.

3,4-Dichlorobenzoic acid biodegradation by the Edwardsiella tarda: Effect of Some Growth Conditions

The biodegradation of 3,4-DiChlorobenzoic acid was investigated by using Edwardsiella tarda and it used 3,4-DCBA as sole carbon and energy source. Several concentrations of 3,4-D CBAs (1mM, 2mM ,3mM ,4mM and 5mM) were used. The highest rate of degradation of 3,4-D CBAs was obtained at a concentration (2mM). The experiments were included substrate concentration, temperature, pH, starvation, adaptation, carbon and nitrogen sources. The degradation ability was monitored through the release of chloride disappearance of the substrate and finally the growth of bacterial cells on that substrate. The optimal temperature and pH for the bacteria were 42ºC and 7.5, respectively. Adaptation of the cells on 3,4-DCBA for 48 hours and cells starvation for 24 hours and 48 hours increasing the initial degradation rate. The carbon sources affected the 3,4 –DCBA degradation differently from that on chloride and cell mass production. Nitrogen sources supplied (yeast extract, L-proline, casein, NH4, K-Nitrate, arginine, urea and glycine). Urea and casine caused a repression in 3,4-DCBA degradation. Catechol 1,2 dioxygenase activity was found to be present in cell free extracts suggesting that 3,4-DCBA is catabolized by ortho-ring cleavage pathway.

Flanking sequences influence the activity of TET1 and TET2 methylcytosine dioxygenases and affect genomic 5hmC patterns

TET dioxygenases convert 5-methylcytosine (5mC) preferentially in a CpG context into 5-hydroxymethylcytosine (5hmC) and higher oxidized forms, thereby initiating DNA demethylation, but details regarding the effects of the DNA sequences flanking the target 5mC site on TET activity are unknown. We investigated oxidation of libraries of DNA substrates containing one 5mC or 5hmC residue in randomized sequence context using single molecule readout of oxidation activity and sequence and show pronounced 20 and 70-fold flanking sequence effects on the catalytic activities of TET1 and TET2, respectively. Flanking sequence preferences were similar for TET1 and TET2 and also for 5mC and 5hmC substrates. Enhanced flanking sequence preferences were observed at non-CpG sites together with profound effects of flanking sequences on the specificity of TET2. TET flanking sequence preferences are reflected in genome-wide and local patterns of 5hmC and DNA demethylation in human and mouse cells indicating that they influence genomic DNA modification patterns in combination with locus specific targeting of TET enzymes.

Indoleamine dioxygenase and tryptophan dioxygenase activities are regulated through GAPDH- and Hsp90-dependent control of their heme levels

Indoleamine-2, 3-dioxygenase (IDO1) and Tryptophan-2, 3-dioxygense (TDO) are heme-containing dioxygenases that catalyze the conversion of tryptophan to N-formyl-kynurenine and thus enable generation of l-kynurenine and related metabolites that govern the immune response and broadly impact human biology. Given that TDO and IDO1 activities are directly proportional to their heme contents, it is important to understand their heme delivery and insertion processes. Early studies established that TDO and IDO1 heme levels are sub-saturating in vivo and subject to change but did not identify the cellular mechanisms that provide their heme or enable dynamic changes in their heme contents. We investigated the potential involvement of GAPDH and chaperone Hsp90, based on our previous studies linking these proteins to intracellular heme allocation. We studied heme delivery and insertion into IDO1 and TDO expressed in both normal and heme-deficient HEK293T cells and into IDO1 naturally expressed in HeLa cells in response to IFN-γ, and also investigated the interactions of TDO and IDO1 with GAPDH and Hsp90 in cells and among their purified forms. We found that GAPDH delivered both mitochondrially-generated and exogenous heme to apo-IDO1 and apo-TDO in cells, potentially through a direct interaction with either enzyme. In contrast, we found Hsp90 interacted with apo-IDO1 but not with apo-TDO, and was only needed to drive heme insertion into apo-IDO1. By uncovering the cellular processes that allocate heme to IDO1 and TDO, our study provides new insight on how their activities and l-kynurenine production may be controlled in health and disease.

Aerobic polychlorinated biphenyl-degrading bacteria isolated from the Tohoku region of Japan are not regionally endemic.

In the Tohoku region of Japan, 72% of the land comprises mountain forest zones. During winter, severe climatic conditions include heavy snowfall. In such an environment, which is considered high in biodiversity, we assumed that aerobic bacteria would be diverse and would possess the ability to degrade polychlorinated biphenyls (PCBs). In this study, 78 environmental samples were collected from the Tohoku region and 56 aerobic PCB-degrading bacterial strains were isolated. They belonged to the genera Achromobacter, Rhodococcus, Pseudomonas, Stenotrophomonas, Comamonas, Pigmentiphaga, Xenophilus, Acinetobacter, and Pandoraea. Previously reported aerobic PCB-degrading bacterial strains isolated in Japan belonged to the same genera, except that the genera Acidovorax and Bacillus were not identified in the present study. In particular, the isolated Comamonas testosteroni strains YAZ2 and YU14-111 had high PCB-degrading abilities. Analysis of the sequences of the YAZ2 and YU14-111 strains showed that the gene structures of the bph operon, which encode enzymes associated with PCB degradation, were the same as those of the Acidovorax sp. KKS102 strain. Moreover, 2,3-biphenyl dioxygenase activity was responsible for the degradation characteristics of all the isolated strains. Overall, this study suggests that aerobic PCB-degrading bacteria are not specifically endemic to the Tohoku region but distributed across Japan.

Determination of the metabolic pathways for degradation of naphthalene and pyrene in Amycolatopsis sp. Poz14

Polycyclic aromatic hydrocarbons (PAHs) constitute important soil contaminants derived from petroleum. Poz14 strain can degrade pyrene and naphthalene. Its genome presented 9333 genes, among them those required for PAHs degradation. By phylogenomic analysis, the strain might be assigned to Amycolatopsis nivea. The strain was grown in glucose, pyrene, and naphthalene to compare their proteomes; 180 proteins were detected in total, and 90 of them were exclusives for xenobiotic conditions. Functions enriched with the xenobiotics belonged to transcription, translation, modification of proteins and transport of inorganic ions. Enriched pathways were pentose phosphate, proteasome and RNA degradation; in contrast, in glucose were glycolysis/gluconeogenesis and glyoxylate cycle. Proteins proposed to participate in the upper PAHs degradation were multicomponent oxygenase complexes, Rieske oxygenases, and dioxygenases; in the lower pathways were ortho-cleavage of catechol, phenylacetate, phenylpropionate, benzoate, and anthranilate. The catechol dioxygenase activity was measured and found increased when the strain was grown in naphthalene. Amycolatopsis sp. Poz14 genome and proteome revealed the PAHs degradation pathways and functions helping to contend the effects of such process.

Enhancement of norisoprenoid and acetoin production for improving the aroma of fermented mango juice by Bacillus subtilis-HNU-B3

Mango juice (MJ) has high nutritional value and decent flavor. However, its aroma could be easily changed by heat treatment during processing, which affects its acceptability and quality. In this study, Bacillus subtilis-HNU-B3 that could transfer carotenoid, was obtained from orchard soil to improve the aroma of fermented mango juice (FMJ). Compared with MJ, the organic acid content of FMJ by B. subtilis-HNU-B3 was significantly changed, and the contents of succinic acid and malic acid were increased to 3.68 mg/mL and 2.04 mg/mL, respectively. Moreover, volatile compounds identified by headspace solid-phase micro-extraction gas-phase mass spectrometry in FMJ were norisoprenoid and acetoin, accounting for 51.52 % of the total volatile substances. In addition, B. subtilis-HNU-B3 fermentation significantly increased the antioxidant capacity and total phenol contents of MJ, and the activity of carotenoid cleavage dioxygenase was 188 U/L. In conclusion, the aroma of FMJ could be improved by B. subtilis-HNU-B3 through increasing the content of norisoprenoid and acetoin. Our results provide theatrical support for Bacillus subtilis to improve the aroma quality of FMJ.

Development of a microtiter plate-based analysis method of nitric oxide dioxygenase activity.

Nitric oxide (NO) functions in cell protection or cell death, depending on its concentration. Therefore, regulation of the intracellular concentrations of NO by its degradation systems is important for cellular functions. One of the NO degrading enzymes, flavohemoglobin (FHb), which has NO dioxygenase (NOD) activity, is a promising target for antibiotics, based on the finding that FHb-deficient pathogens exhibited reduced host toxicity. Here, we developed a high-throughput method to measure the NOD activity. Our newly developed method could contribute to the screening of potential antibiotics with NOD inhibitory activity.

Plant growth promoting endophytic Pseudomonas aeruginosa RD10 isolated from water hyacinth for phenol degradation

A total of twelve endophytic bacteria were isolated from the root of E. crassipes. All these bacterial strains were analyzed for phenol degradation potential. Twelve endophytic bacteria were subjected to polyphenol resistance analysis and all showed phenol tolerance up to 300 ppm. Among the twelve bacterial strains, the strain RD10 was resistant up to 800 ppm, and reduced growth were observed at higher concentrations. The plant growth-promoting properties of endophytic bacterial strains were analyzed. Among the isolated bacterial strain, only five strains produced ACC deaminase. IAA production was detected in seven bacterial isolates, while siderophore production was determined from only five bacterial strains. Among the isolated endophytes, the strain RD10 exhibited ACC deaminase, IAA, and siderophore activity in submerged fermentation. ACC deaminase production of 125.2 ± 4.8 m/ml was shown by strain RD10 and IAA production was 89.7 ± 5.1 μg/ml. Moreover, maximum siderophore production was observed (53.9 ± 4.9% siderophore units) in strain RD3. Catechol 2,3-dioxygenase activity of the bacterial strains was analyzed. Catechol 2,3 dioxygenase activity ranges from 0.03 ± 0.01 U/mg to 35.2 ± 1.1 U/mg. Phenol degradation by the isolated endophytes was studied and the initial phenol concentration was 500 ppm. The strain RD1 degraded 1.2 ± 0.2% phenol and more than 50% phenol degradation was observed in strains RD3, RD4, RD6, RD7, RD9, RD10, and RD11, respectively. The endophytic bacterial strain was Gram-negative, rodshaped and motile. It was non-spore-forming bacteria, capsulated and, showed a positive reaction to oxidase and catalase. In MacConkey agar medium, strain RD10 forms smooth and flat colonies, and the diameter of the colonies ranged from 2 to 3 mm. The colonies have an alligator skin-like appearance on top view and have regular margins. The strain RD10 was streaked on blood agar medium and the strain RD10 produced strong pigmentation on blood agar. Antibiotic sensitivity of strain RD10 against various antibiotics in terms of zone of inhibition was analyzed. The endophytic strain RD10 was sensitive to all selected antibiotics. Based on biochemical characters, pigment analysis and, molecular characterization, the strain was confirmed as P. aeruginosa RD10. In this study, P. aeruginosa RD10 was cultured at 400, 500 and, 600 mg/l phenol concentration for 96 h. Optimum growth was achieved at 500 mg/l concentration and an inhibitory effect was observed at 600 mg/l phenol concentration. The results confirmed that the selected strain can withstand and grow well at 500 mg/l phenol concentration. The strain RD10 was cultured at various incubation temperatures with 500 mg/l phenol concentration. Optimization of environmental factors can lead to maximum degradation of phenol. The results revealed that phenol degradation was influenced by temperature. The highest phenol degradation was achieved at 30 °C in Erlenmeyer flask culture and decreased at high temperatures. P. aeruginosa RD10 could degrade phenol at pH 5.5 (10.2 ± 0.5% removal), at pH 6.0, 15.4 ± 1.1% of the phenol was degraded after incubation for 72 h at 30 °C. Phenol removal (%) declined to 70.4 ± 2.9% at higher initial pH value (9.0).

Comparison of quintrione and quinclorac on mechanism of action

Quintrione is a new post-emergence herbicide developed for use in rice; however, the mechanism of action remains unclear. We determined the phytotoxicity of quintrione, and the contributions of hormone levels and lipid peroxidation to phytotoxicity, by comparing them to those induced by quinclorac. We also investigated 4-hydroxyphenylpyruvate dioxygenase (HPPD) activity and carotenoid content following treatment with quintrione by comparing them to those induced by quinclorac and mesotrione. We found that quintrione and quinclorac both inhibited the growth of Echinochloa crusgalli var. zelayensis, but that quinclorac was a little more effective. At 24 h, quintrione and quinclorac significantly increased ethylene production and the contents of abscisic acid (ABA) and indole acetic acid (IAA) compared with the control. No significant differences were observed between quintrione and quinclorac on the three plant hormones. Quintrione and quinclorac also induced the formation of malondialdehyde (MDA), which is associated with lipid peroxidation, with no significant difference between them. Carotenoid content was reduced in E. crusgalli var. zelayensis following treatments with quintrione, quinclorac, and mesotrione. At 120 h, carotenoid contents were significantly higher following the quintrione and quinclorac treatments, in comparison with mesotrione treatment. There were no significant differences between quintrione and quinclorac in the inhibition of HPPD activity, and the effects of both were significantly less than the effect of mesotrione. In summary, E. crusgalli var. zelayensis was susceptible to both quintrione and quinclorac. The mechanism of action of quintrione, like that of quinclorac, was related to levels of plant hormones and lipid peroxidation; however, quintrione was a poor inhibitor of HPPD activity compared to mesotrione.

TET2 mutations as a part of DNA dioxygenase deficiency in myelodysplastic syndromes

AbstractDecrease in DNA dioxygenase activity generated by TET2 gene family is crucial in myelodysplastic syndromes (MDS). The general downregulation of 5-hydroxymethylcytosine (5-hmC) argues for a role of DNA demethylation in MDS beyond TET2 mutations, which albeit frequent, do not convey any prognostic significance. We investigated TETs expression to identify factors which can modulate the impact of mutations and thus 5-hmC levels on clinical phenotypes and prognosis of MDS patients. DNA/RNA-sequencing and 5-hmC data were collected from 1665 patients with MDS and 91 controls. Irrespective of mutations, a significant fraction of MDS patients exhibited lower TET2 expression, whereas 5-hmC levels were not uniformly decreased. In searching for factors explaining compensatory mechanisms, we discovered that TET3 was upregulated in MDS and inversely correlated with TET2 expression in wild-type cases. Although TET2 was reduced across all age groups, TET3 levels were increased in a likely feedback mechanism induced by TET2 dysfunction. This inverse relationship of TET2 and TET3 expression also corresponded to the expression of L-2-hydroxyglutarate dehydrogenase, involved in agonist/antagonist substrate metabolism. Importantly, elevated TET3 levels influenced the clinical phenotype of TET2 deficiency whereby the lack of compensation by TET3 (low TET3 expression) was associated with poor outcomes of TET2 mutant carriers.