scholarly journals Cloning of thioredoxin h reductase and characterization of the thioredoxin reductase–thioredoxin h system from wheat

2002 ◽  
Vol 367 (2) ◽  
pp. 491-497 ◽  
Author(s):  
Antonio J. SERRATO ◽  
Juan M. PÉREZ-RUIZ ◽  
Francisco J. CEJUDO
Keyword(s):  
Developmental Stages ◽  
Thioredoxin Reductase ◽  
Wide Distribution ◽  
Binding Domains ◽  
Reduction Activity ◽  
Lack Of Information ◽  
Thioredoxin H ◽  
Single Polypeptide ◽  
High Level

Thioredoxins h are ubiquitous proteins reduced by NADPH— thioredoxin reductase (NTR). They are able to reduce disulphides in target proteins. In monocots, thioredoxins h accumulate at high level in seeds and show a predominant localization in the nucleus of seed cells. These results suggest that the NTR—thioredoxin h system probably plays an important role in seed physiology. To date, the study of this system in monocots is limited by the lack of information about NTR. In the present study, we describe the cloning of a full-length cDNA encoding NTR from wheat (Triticum aestivum). The polypeptide deduced from this cDNA shows close similarity to NTRs from Arabidopsis, contains FAD- and NADPH-binding domains and a disulphide probably interacting with the disulphide at the active site of thioredoxin h. Wheat NTR was expressed in Escherichia coli as a His-tagged protein. The absorption spectrum of the purified recombinant protein is typical of flavoenzymes. Furthermore, it showed NADPH-dependent thioredoxin h reduction activity, thus confirming that the cDNA clone reported in the present study encodes wheat NTR. Using the His-tagged NTR and TRXhA (wheat thioredoxin h), we successfully reconstituted the wheat NTR—thioredoxin h system in vitro, as shown by the insulin reduction assay. A polyclonal antibody was raised against wheat NTR after immunization of rabbits with the purified His-tagged protein. This antibody efficiently detected a single polypeptide of the corresponding molecular mass in seed extracts and it allowed the analysis of the pattern of accumulation of NTR in different wheat organs and developmental stages. NTR shows a wide distribution in wheat, but, surprisingly, its accumulation in seeds is low, in contrast with the level of thioredoxins h.

scholarly journals Identification and Regulation of Bone Morphogenetic Protein Antagonists Associated with Preantral Follicle Development in the Ovary

Endocrinology ◽  
2011 ◽  
Vol 152 (9) ◽  
pp. 3515-3526 ◽  
Author(s):  
Mark A. Fenwick ◽  
Yosef T. Mansour ◽  
Stephen Franks ◽  
Kate Hardy
Keyword(s):  
Bone Morphogenetic Proteins ◽  
Developmental Stages ◽  
Follicle Development ◽  
Preantral Follicles ◽  
Twisted Gastrulation ◽  
Morphogenetic Protein ◽  
Gene Transcripts ◽  
Independent Effects ◽  
High Level

The TGFβ superfamily comprises several bone morphogenetic proteins (BMP) capable of exerting gonadotropin-independent effects on the development of small preantral follicles. In embryonic tissues, BMP concentration gradients, partly formed by antagonistic factors, are essential for establishing phenotypic fate. By examining the expression of candidate genes whose protein products are known to interact with BMP ligands, we set out to determine which antagonists would most likely contribute toward regulation of paracrine signaling during early follicle development. Juvenile mouse ovaries of 4, 8, 12, and 21 d of age enriched with follicles at successive developmental stages were used to assess changes in candidate gene transcripts by quantitative RT-PCR. Although some antagonists were found to be positively associated with the emergence of developing follicles (Nog, Htra1, Fst, Bmper, Vwc2), two (Sostdc1, Chrd) showed a corresponding reduction in expression. At each age, twisted gastrulation homolog 1 (Twsg1), Htra1, Nbl1, and Fst were consistently highly expressed and localization of these genes by in situ hybridization, and immunohistochemistry further highlighted a clear pattern of expression in granulosa cells of developing follicles. Moreover, with the exception of Nbl1, levels of these antagonists did not change in preantral follicles exposed to FSH in vitro, suggesting regulation by local factors. The presence of multiple antagonists in the juvenile ovary and their high level of expression in follicles imply the actions of certain growth factors are subject to local modulation and further highlights another important level of intraovarian regulation of follicle development.

scholarly journals Selenium Nanoparticle Synthesized by Proteus mirabilis YC801: An Efficacious Pathway for Selenite Biotransformation and Detoxification

2018 ◽  
Vol 19 (12) ◽  
pp. 3809 ◽  
Author(s):  
Yuting Wang ◽  
Xian Shu ◽  
Jinyan Hou ◽  
Weili Lu ◽  
Weiwei Zhao ◽  
...  
Keyword(s):  
Proteus Mirabilis ◽  
Thioredoxin Reductase ◽  
Hydrodynamic Diameter ◽  
Elemental Selenium ◽  
Selenite Reduction ◽  
Reduction Activity ◽  
Selenium Nanoparticle ◽  
Infrared Spectral ◽  
Average Hydrodynamic Diameter

Selenite is extremely biotoxic, and as a result of this, exploitation of microorganisms able to reduce selenite to non-toxic elemental selenium (Se0) has attracted great interest. In this study, a bacterial strain exhibiting extreme tolerance to selenite (up to 100 mM) was isolated from the gut of adult Monochamus alternatus and identified as Proteus mirabilis YC801. This strain demonstrated efficient transformation of selenite into red selenium nanoparticles (SeNPs) by reducing nearly 100% of 1.0 and 5.0 mM selenite within 42 and 48 h, respectively. Electron microscopy and energy dispersive X-ray analysis demonstrated that the SeNPs were spherical and primarily localized extracellularly, with an average hydrodynamic diameter of 178.3 ± 11.5 nm. In vitro selenite reduction activity assays and real-time PCR indicated that thioredoxin reductase and similar proteins present in the cytoplasm were likely to be involved in selenite reduction, and that NADPH or NADH served as electron donors. Finally, Fourier-transform infrared spectral analysis confirmed the presence of protein and lipid residues on the surfaces of SeNPs. This is the first report on the capability of P. mirabilis to reduce selenite to SeNPs. P. mirabilis YC801 might provide an eco-friendly approach to bioremediate selenium-contaminated soil/water, as well as a bacterial catalyst for the biogenesis of SeNPs.

scholarly journals Selenite Reduction and the Biogenesis of Selenium Nanoparticles by Alcaligenes faecalis Se03 Isolated from the Gut of Monochamus alternatus (Coleoptera: Cerambycidae)

2018 ◽  
Vol 19 (9) ◽  
pp. 2799 ◽  
Author(s):  
Yuting Wang ◽  
Xian Shu ◽  
Qing Zhou ◽  
Tao Fan ◽  
Taichu Wang ◽  
...  
Keyword(s):  
Thioredoxin Reductase ◽  
Alcaligenes Faecalis ◽  
Selenium Nanoparticles ◽  
Synthesis Process ◽  
Hydrodynamic Diameter ◽  
Monochamus Alternatus ◽  
Selenite Reduction ◽  
Reduction Activity ◽  
Liquid Culture Medium

In this study, a bacterial strain exhibiting high selenite (Na2SeO3) tolerance and reduction capacity was isolated from the gut of Monochamus alternatus larvae and identified as Alcaligenes faecalis Se03. The isolate exhibited extreme tolerance to selenite (up to 120 mM) when grown aerobically. In the liquid culture medium, it was capable of reducing nearly 100% of 1.0 and 5.0 mM Na2SeO3 within 24 and 42 h, respectively, leading to the formation of selenium nanoparticles (SeNPs). Electron microscopy and energy dispersive X-ray analysis demonstrated that A. faecalis Se03 produced spherical electron-dense SeNPs with an average hydrodynamic diameter of 273.8 ± 16.9 nm, localized mainly in the extracellular space. In vitro selenite reduction activity and real-time PCR indicated that proteins such as sulfite reductase and thioredoxin reductase present in the cytoplasm were likely to be involved in selenite reduction and the SeNPs synthesis process in the presence of NADPH or NADH as electron donors. Finally, using Fourier transform infrared spectroscopy, protein and carbohydrate residues were detected on the surface of the biogenic SeNPs. Based on these observations, A. faecalis Se03 has the potential to be an eco-friendly candidate for the bioremediation of selenium-contaminated soil/water and a bacterial catalyst for the biogenesis of SeNPs.

Correlated Studies of Cellular Interactions Using TEM, Freeze Etching, and SEM

1974 ◽  
Vol 32 ◽  
pp. 320-321
Author(s):  
J. P. Revel
Keyword(s):  
Developmental Stages ◽  
Three Dimensional ◽  
Cell Movement ◽  
Cellular Interactions ◽  
Serial Sections ◽  
Cell Sheets ◽  
Freeze Etching ◽  
Early Developmental

Movement of individual cells or of cell sheets and complex patterns of folding play a prominent role in the early developmental stages of the embryo. Our understanding of these processes is based on three- dimensional reconstructions laboriously prepared from serial sections, and from autoradiographic and other studies. Many concepts have also evolved from extrapolation of investigations of cell movement carried out in vitro. The scanning electron microscope now allows us to examine some of these events in situ. It is possible to prepare dissections of embryos and even of tissues of adult animals which reveal existing relationships between various structures more readily than used to be possible vithout an SEM.

RAPID IN VITRO PROPAGATION OF WESTERN GHATS CULTIVAR - COLOCASIA ESCULENTA FOR THE HIGH FREQUENCY OF PLANTLETS

2019 ◽  
Vol 7 (2) ◽  
Author(s):  
Jyothi R ◽  
Srinivasa Murthy K M ◽  
Hossein . ◽  
Veena .
Keyword(s):  
Tissue Culture ◽  
Wide Distribution ◽  
Colocasia Esculenta ◽  
Limiting Factor ◽  
Promising Alternative ◽  
Rapid Multiplication ◽  
Medical Plant ◽  
Total Fat ◽  
Planting Materials

Colocasia esculenta is commonly known as Taro, it is referred to as cocoyam in Nigeria. They are cherished for their rich taste, nutritional and medicinal properties. Every 100 g of taro corms possess 112 Kcal, 26.46 g carbohydrate, 1.50 g protein, 0.20 g total fat and 4.1g fiber (USDA National Nutrient Data Base). Besides its nutritional value, taro is used as a medical plant and provides bioactive compounds used as an anti-cancer drugs. Traditionally, cocoyams are vegetative propagated from tuber fragments, a practice that encourages pathogen distribution. Colocasia esculenta is a widely distributed food crop in the humid tropics and subtropics. Despite of its wide distribution, Taro plants are commonly infected with DsMV and other pathogens. This virus induces conspicuous mosaic, malformation, dwarfing or feathering on leaves in taro. As the results of infection, it reduces the quality and yield of taro production greatly. This virus is thus considered as a major limiting factor in the production of taro. Here plays the importance of  tissue culture plays a major role in producing the disease resistant plants round the year with high quality. For rapid multiplication and production of quality planting materials, tissue culture technology offers promising alternative compared to the traditional production methods. KEYWORDS: Colocasia esculenta, Virus, Pathogens, Conventional propagation, Micropropagation, Yield, Rapid multiplication, Quality

In Vitro Effect of Chlorquine and Picroliv on Plasmodium Berghei Induced Alterations in the Activity of Adenosine Triphosphatase, Aryl Hyrocarbon Hydroxylase Enzymes and Malondialdehyde in spleen Explant Culture

2020 ◽  
Vol 20 ◽  
Author(s):  
Anchal Trivedi ◽  
Aparna Misra ◽  
Esha Sarkar ◽  
Anil K. Balapure
Keyword(s):  
Drug Resistance ◽  
Plasmodium Berghei ◽  
Adenosine Triphosphatase ◽  
Explant Culture ◽  
Need To Evaluate ◽  
Mda Content ◽  
High Level ◽  
Vitro Effect ◽  
Positive Effect

Background: In recent years, great progress has been made in reducing the high level of malaria suffering worldwide. There is a great need to evaluate drug resistance reversers and consider new medicines against malaria. There are many approaches to the development of antimalarial drugs. Specific concerns must be taken in to account in these approaches, in particular there requirement for very in expensive and simple use of new therapies and the need to limit drug discovery expenses. Important ongoing efforts are the optimisation of treatment with available medications, including the use of combination therapy. The production of analogs of known agents and the identification of natural products, the use of compounds originally developed against other diseases, the assessment of overcoming drug resistance and the consideration of new therapeutic targets. Liver and spleen are the important organs which are directly associated with malarial complications. Aim: An analysis the Activity of Adenosine Triphosphatase, Aryl Hyrocarbon Hydroxylase Enzymes and Malondialdehyde in spleen Explant Culture. Objective: To determine in-Vitro Effect of Chlorquine and Picroliv on Plasmodium Berghei Induced Alterations in the Activity of Adenosine Triphosphatase, Aryl Hyrocarbon Hydroxylase Enzymes and Malondialdehyde in spleen Explant Culture. Material and method: 1-Histological preparation of spleen explants for paraplast embedding 2-Biochemicalstudies (Enzymes (Atpase, ALP&GST) and the level of protein, Malondialdehyde (MDA). Result: Splenomegalyis one of the three main diagnostic parameters of malaria infection besides fever and anaemia. Many enzymes present in the liver and spleen may also be altered or liberated under different pathological conditions. Enzymes (ATPase, ALP&GST) and the level of protein, Malondialdehyde (MDA) content was found to increase in the liver and spleen explants during malarial infection. In the liver and spleen derived from parasitized CQ treated animals, the activity of all the above enzymes (ATPase, ALP&GST) and the level of protein & MDA of liver/spleen reversed towards the normal for all the 4or3 days of incubations. Picroliv efficacy decreased with the increment of parasitaemia and at 60%parasitaemia. Conclusion: Alkalinephosphatase (ALP) was found to increase with increasing parasitaemia. After the addition of Picroliv to the medium, a decrement in the activity was observed up to day 4 of culture.A similar positive effect of Picroliv was observed on the ATPase and ALP activity of spleen explants.DNA and protein contents also increased in the parasitized liver cultured in the presence of picroliv.On the contrary, in the spleen explants DNA, protein and MDA content were found to decrease after Picroliv supplementation to the culture medium.

scholarly journals Mutations in fbiD (Rv2983) as a Novel Determinant of Resistance to Pretomanid and Delamanid in Mycobacterium tuberculosis

2020 ◽  
Vol 65 (1) ◽  
pp. e01948-20
Author(s):  
Dalin Rifat ◽  
Si-Yang Li ◽  
Thomas Ioerger ◽  
Keshav Shah ◽  
Jean-Philippe Lanoix ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Clinical Evidence ◽  
Clinical Samples ◽  
Loss Of Function ◽  
Wild Type ◽  
Content Type ◽  
Solid Media ◽  
High Level ◽  
Level Resistance

ABSTRACTThe nitroimidazole prodrugs delamanid and pretomanid comprise one of only two new antimicrobial classes approved to treat tuberculosis (TB) in 50 years. Prior in vitro studies suggest a relatively low barrier to nitroimidazole resistance in Mycobacterium tuberculosis, but clinical evidence is limited to date. We selected pretomanid-resistant M. tuberculosis mutants in two mouse models of TB using a range of pretomanid doses. The frequency of spontaneous resistance was approximately 10−5 CFU. Whole-genome sequencing of 161 resistant isolates from 47 mice revealed 99 unique mutations, of which 91% occurred in 1 of 5 genes previously associated with nitroimidazole activation and resistance, namely, fbiC (56%), fbiA (15%), ddn (12%), fgd (4%), and fbiB (4%). Nearly all mutations were unique to a single mouse and not previously identified. The remaining 9% of resistant mutants harbored mutations in Rv2983 (fbiD), a gene not previously associated with nitroimidazole resistance but recently shown to be a guanylyltransferase necessary for cofactor F420 synthesis. Most mutants exhibited high-level resistance to pretomanid and delamanid, although Rv2983 and fbiB mutants exhibited high-level pretomanid resistance but relatively small changes in delamanid susceptibility. Complementing an Rv2983 mutant with wild-type Rv2983 restored susceptibility to pretomanid and delamanid. By quantifying intracellular F420 and its precursor Fo in overexpressing and loss-of-function mutants, we provide further evidence that Rv2983 is necessary for F420 biosynthesis. Finally, Rv2983 mutants and other F420H2-deficient mutants displayed hypersusceptibility to some antibiotics and to concentrations of malachite green found in solid media used to isolate and propagate mycobacteria from clinical samples.

scholarly journals Allelic Exchange and Mutant Selection Demonstrate that Common Clinical embCAB Gene Mutations Only Modestly Increase Resistance to Ethambutol in Mycobacterium tuberculosis

2009 ◽  
Vol 54 (1) ◽  
pp. 103-108 ◽  
Author(s):  
Hassan Safi ◽  
Robert D. Fleischmann ◽  
Scott N. Peterson ◽  
Marcus B. Jones ◽  
Behnam Jarrahi ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
In Vitro Selection ◽  
Gene Mutations ◽  
Wild Type ◽  
Mutant Selection ◽  
Preferential Growth ◽  
Allelic Exchange ◽  
High Level ◽  
Isogenic Mutants

ABSTRACT Mutations within codon 306 of the Mycobacterium tuberculosis embB gene modestly increase ethambutol (EMB) MICs. To identify other causes of EMB resistance and to identify causes of high-level resistance, we generated EMB-resistant M. tuberculosis isolates in vitro and performed allelic exchange studies of embB codon 406 (embB406) and embB497 mutations. In vitro selection produced mutations already identified clinically in embB306, embB397, embB497, embB1024, and embC13, which result in EMB MICs of 8 or 14 μg/ml, 5 μg/ml, 12 μg/ml, 3 μg/ml, and 4 μg/ml, respectively, and mutations at embB320, embB324, and embB445, which have not been identified in clinical M. tuberculosis isolates and which result in EMB MICs of 8 μg/ml, 8 μg/ml, and 2 to 8 μg/ml, respectively. To definitively identify the effect of the common clinical embB497 and embB406 mutations on EMB susceptibility, we created a series of isogenic mutants, exchanging the wild-type embB497 CAG codon in EMB-susceptible M. tuberculosis strain 210 for the embB497 CGG codon and the wild-type embB406 GGC codon for either the embB406 GCC, embB406 TGC, embB406 TCC, or embB406 GAC codon. These new mutants showed 6-fold and 3- to 3.5-fold increases in the EMB MICs, respectively. In contrast to the embB306 mutants, the isogenic embB497 and embB406 mutants did not have preferential growth in the presence of isoniazid or rifampin (rifampicin) at their MICs. These results demonstrate that individual embCAB mutations confer low to moderate increases in EMB MICs. Discrepancies between the EMB MICs of laboratory mutants and clinical M. tuberculosis strains with identical mutations suggest that clinical EMB resistance is multigenic and that high-level EMB resistance requires mutations in currently unknown loci.

scholarly journals Aberrantly high activation of a FoxM1–STMN1 axis contributes to progression and tumorigenesis in FoxM1-driven cancers

2021 ◽  
Vol 6 (1) ◽  
Author(s):  
Jun Liu ◽  
Jipeng Li ◽  
Ke Wang ◽  
Haiming Liu ◽  
Jianyong Sun ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Poor Prognosis ◽  
Soft Agar ◽  
Transcriptional Level ◽  
Regulation Mechanism ◽  
Strong Positive Correlation ◽  
Cell Viability Assay ◽  
High Level

AbstractFork-head box protein M1 (FoxM1) is a transcriptional factor which plays critical roles in cancer development and progression. However, the general regulatory mechanism of FoxM1 is still limited. STMN1 is a microtubule-binding protein which can inhibit the assembly of microtubule dimer or promote depolymerization of microtubules. It was reported as a major responsive factor of paclitaxel resistance for clinical chemotherapy of tumor patients. But the function of abnormally high level of STMN1 and its regulation mechanism in cancer cells remain unclear. In this study, we used public database and tissue microarrays to analyze the expression pattern of FoxM1 and STMN1 and found a strong positive correlation between FoxM1 and STMN1 in multiple types of cancer. Lentivirus-mediated FoxM1/STMN1-knockdown cell lines were established to study the function of FoxM1/STMN1 by performing cell viability assay, plate clone formation assay, soft agar assay in vitro and xenograft mouse model in vivo. Our results showed that FoxM1 promotes cell proliferation by upregulating STMN1. Further ChIP assay showed that FoxM1 upregulates STMN1 in a transcriptional level. Prognostic analysis showed that a high level of FoxM1 and STMN1 is related to poor prognosis in solid tumors. Moreover, a high co-expression of FoxM1 and STMN1 has a more significant correlation with poor prognosis. Our findings suggest that a general FoxM1-STMN1 axis contributes to cell proliferation and tumorigenesis in hepatocellular carcinoma, gastric cancer and colorectal cancer. The combination of FoxM1 and STMN1 can be a more precise biomarker for prognostic prediction.

scholarly journals Genome-Wide Transcriptomic Identification and Functional Insight of Lily WRKY Genes Responding to Botrytis Fungal Disease

Plants ◽  
2021 ◽  
Vol 10 (4) ◽  
pp. 776
Author(s):  
Shipra Kumari ◽  
Bashistha Kumar Kanth ◽  
Ju young Ahn ◽  
Jong Hwa Kim ◽  
Geung-Joo Lee
Keyword(s):  
Abiotic Stress ◽  
Biotic Stress ◽  
Developmental Stages ◽  
Expression Patterns ◽  
Lilium Longiflorum ◽  
Biotic And Abiotic Stress ◽  
Biotic Stresses ◽  
Genome Wide

Genome-wide transcriptome analysis using RNA-Seq of Lilium longiflorum revealed valuable genes responding to biotic stresses. WRKY transcription factors are regulatory proteins playing essential roles in defense processes under environmental stresses, causing considerable losses in flower quality and production. Thirty-eight WRKY genes were identified from the transcriptomic profile from lily genotypes, exhibiting leaf blight caused by Botrytis elliptica. Lily WRKYs have a highly conserved motif, WRKYGQK, with a common variant, WRKYGKK. Phylogeny of LlWRKYs with homologous genes from other representative plant species classified them into three groups- I, II, and III consisting of seven, 22, and nine genes, respectively. Base on functional annotation, 22 LlWRKY genes were associated with biotic stress, nine with abiotic stress, and seven with others. Sixteen unique LlWRKY were studied to investigate responses to stress conditions using gene expression under biotic and abiotic stress treatments. Five genes—LlWRKY3, LlWRKY4, LlWRKY5, LlWRKY10, and LlWRKY12—were substantially upregulated, proving to be biotic stress-responsive genes in vivo and in vitro conditions. Moreover, the expression patterns of LlWRKY genes varied in response to drought, heat, cold, and different developmental stages or tissues. Overall, our study provides structural and molecular insights into LlWRKY genes for use in the genetic engineering in Lilium against Botrytis disease.

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