scholarly journals In vivo analysis of DNA-protein interactions on the human erythropoietin enhancer.

1997 ◽  
Vol 17 (2) ◽  
pp. 851-856 ◽  
Author(s):  
B Hu ◽  
E Wright ◽  
L Campbell ◽  
K L Blanchard
Keyword(s):  
Oxygen Tension ◽  
Protein Interactions ◽  
Proximal Promoter ◽  
Low Oxygen ◽  
Initiation Rate ◽  
Cobalt Exposure ◽  
In Cells ◽  
Dna Protein Interactions

The erythropoietin (EPO) gene is one of the best examples of a mammalian gene controlled by oxygen tension. The DNA elements responsible for hypoxia-induced transcription consist of a short region of the proximal promoter and a <50-bp 3' enhancer. The elements act cooperatively to increase the transcriptional initiation rate approximately 100-fold in response to low oxygen tension in Hep3B cells. Two distinct types of transactivating proteins have been demonstrated to bind the response elements in the human EPO enhancer in vitro: one shows hypoxia-inducible DNA binding activity, while the other activity binds DNA under normoxic and hypoxic conditions. We have investigated the DNA-protein interactions on the human EPO enhancer in living tissue culture cells that produce EPO in a regulated fashion (Hep3B) and in cells that do not express EPO under any conditions tested (HeLa). We have identified in vivo DNA-protein interactions on the control elements in the human EPO enhancer by ligation-mediated PCR technology. We show that the putative protein binding sites in the EPO enhancer are occupied in vivo under conditions of normoxia, hypoxia, and cobalt exposure in EPO-producing cells. These sites are not occupied in cells that do not produce EPO. We also provide evidence for a conformational change in the topography of the EPO enhancer in response to hypoxia and cobalt exposure.

scholarly journals Ways into Understanding HIF Inhibition

Cancers ◽  
2021 ◽  
Vol 13 (1) ◽  
pp. 159
Author(s):  
Tina Schönberger ◽  
Joachim Fandrey ◽  
Katrin Prost-Fingerle
Keyword(s):  
Protein Interactions ◽  
Target Genes ◽  
Protein Protein Interactions ◽  
Low Oxygen ◽  
Drug Candidates ◽  
Open Questions ◽  
Wide Range ◽  
Hif Pathway

Hypoxia is a key characteristic of tumor tissue. Cancer cells adapt to low oxygen by activating hypoxia-inducible factors (HIFs), ensuring their survival and continued growth despite this hostile environment. Therefore, the inhibition of HIFs and their target genes is a promising and emerging field of cancer research. Several drug candidates target protein–protein interactions or transcription mechanisms of the HIF pathway in order to interfere with activation of this pathway, which is deregulated in a wide range of solid and liquid cancers. Although some inhibitors are already in clinical trials, open questions remain with respect to their modes of action. New imaging technologies using luminescent and fluorescent methods or nanobodies to complement widely used approaches such as chromatin immunoprecipitation may help to answer some of these questions. In this review, we aim to summarize current inhibitor classes targeting the HIF pathway and to provide an overview of in vitro and in vivo techniques that could improve the understanding of inhibitor mechanisms. Unravelling the distinct principles regarding how inhibitors work is an indispensable step for efficient clinical applications and safety of anticancer compounds.

scholarly journals The Role of Tissue Oxygen Tension in Dengue Virus Replication

Cells ◽  
2018 ◽  
Vol 7 (12) ◽  
pp. 241 ◽  
Author(s):  
Efseveia Frakolaki ◽  
Panagiota Kaimou ◽  
Maria Moraiti ◽  
Katerina Kalliampakou ◽  
Kalliopi Karampetsou ◽  
...  
Keyword(s):  
Dengue Virus ◽  
Oxygen Tension ◽  
Atmospheric Oxygen ◽  
Rna Replication ◽  
Metabolic Reprogramming ◽  
Anaerobic Glycolysis ◽  
Low Oxygen ◽  
Hypoxia Response

Low oxygen tension exerts a profound effect on the replication of several DNA and RNA viruses. In vitro propagation of Dengue virus (DENV) has been conventionally studied under atmospheric oxygen levels despite that in vivo, the tissue microenvironment is hypoxic. Here, we compared the efficiency of DENV replication in liver cells, monocytes, and epithelial cells under hypoxic and normoxic conditions, investigated the ability of DENV to induce a hypoxia response and metabolic reprogramming and determined the underlying molecular mechanism. In DENV-infected cells, hypoxia had no effect on virus entry and RNA translation, but enhanced RNA replication. Overexpression and silencing approaches as well as chemical inhibition and energy substrate exchanging experiments showed that hypoxia-mediated enhancement of DENV replication depends on the activation of the key metabolic regulators hypoxia-inducible factors 1α/2α (HIF-1α/2α) and the serine/threonine kinase AKT. Enhanced RNA replication correlates directly with an increase in anaerobic glycolysis producing elevated ATP levels. Additionally, DENV activates HIF and anaerobic glycolysis markers. Finally, reactive oxygen species were shown to contribute, at least in part through HIF, both to the hypoxia-mediated increase of DENV replication and to virus-induced hypoxic reprogramming. These suggest that DENV manipulates hypoxia response and oxygen-dependent metabolic reprogramming for efficient viral replication.

134 Effects of Gas Tension During Culture upon Development of Bovine Embryos from Beef (Nellore) and Dairy (Girolando) Breeds

2018 ◽  
Vol 30 (1) ◽  
pp. 207
Author(s):  
J. G. V. Grázia ◽  
L. G. Lacerda ◽  
L. G. B. Siqueira ◽  
C. A. G. Pellegrino ◽  
L. S. Grapiuna ◽  
...  
Keyword(s):  
Oxygen Tension ◽  
Initial Period ◽  
Bos Indicus ◽  
Southeastern Brazil ◽  
Bovine Embryos ◽  
Low Oxygen ◽  
Cleavage Rate ◽  
High O2

Culture of bovine embryos is a critical step during in vitro embryo production (IVEP) and, as such, has been the focus of numerous studies on cattle IVEP. Improvements of culture conditions to mimic the in vivo maternal microenvironment involves studying the optimal gas tension for pre-implantation embryonic development. In the commercial conditions, there is great variability in results, in part because of the difference between breeds and donors. The objective of this study was to evaluate the effects of culture in high or low oxygen tension upon the development of embryos from a crossbred dairy breed (Girolando F1; Gir × Holstein) and a beef Bos indicus breed, Nellore. We collected data from an IVEP commercial operation located in a tropical area of southeastern Brazil (Minas Gerais State) from February to May 2017. The study was designed in a 2 × 2 factorial arrangement of treatments: 2 O2 tensions during culture (5%, low O2 v. 20%, high O2) and 2 breeds (Nellore, beef v. Girolando F1, dairy). Thus, the following 4 groups were studied: Nellore-high O2 (n = 86 donors), Nellore-low O2 (n = 107 donors), Girolando F1-high O2 (n = 114 donors), and Girolando F1-low O2 (n = 110 donors). Outcome variables were the number of cleaved embryos 72 h post-insemination (hpi), cleavage rate relative to the total number cumulus–oocyte complexes (COC) put in culture, number and percentage of blastocysts 192 hpi relative to the structures kept in culture. Variables that were not normally distributed were transformed using the formula log(y + 0.05). Data were analysed using the GLM procedure of SAS (SAS Institute Inc., Cary, NC, USA) for the main effects of gas tension (low v. high O2) and breed (Girolando F1 v. Nelore). Results are shown as mean ± SEM. Gas tension affected the number of cleaved embryos (10.52 ± 0.92 v. 8.33 ± 0.72 for high and low O2, respectively; P < 0.01) and cleavage rates (40.58 ± 2.49 v. 44.41 ± 2.88 for high and low O2; P < 0.01 in Nellore), but did not affect these variables in Girolando F1 donors (13.23 ± 1.33 v. 10.76 ± 0.76 cleaved embryos, for high and low O2; P = 0.63; 58.01 ± 2.00 v. 60.19 ± 1.97 cleavage rate, for high and low O2; P = 0.80). Nonetheless, the number and percentage of blastocysts were not affected by gas tension in either breed. Results for Nellore were 4.99 ± 0.56 v. 3.51 ± 0.38 blastocysts in high and low O2, respectively (P = 0.051) and 41.92 ± 3.91% v. 39.81 ± 3.77% blastocysts, in high and low O2 (P = 0.11). For Girolando F1, numbers of blastocysts were 5.84 ± 0.66 v. 4.24 ± 0.39 in high and low O2 (P = 0.19) and percentage of blastocysts 49.14 ± 2.97% v. 49.11 ± 3.40% in high and low O2 (P = 0.46). These results suggest that oxygen tension during culture affects IVEP differently depending on breed. The initial period of culture, recognised as critical in IVEP, seemed more sensitive to high O2 tension, particularly in Nellore.

Embryo culture at a reduced oxygen concentration of 5%: a mini review

Zygote ◽  
2019 ◽  
Vol 27 (6) ◽  
pp. 355-361 ◽  
Author(s):  
R. Sciorio ◽  
G.D. Smith
Keyword(s):  
Oxygen Tension ◽  
Embryo Development ◽  
Atmospheric Oxygen ◽  
Embryo Implantation ◽  
Critical Role ◽  
Low Oxygen ◽  
Physiological Level ◽  
Low Oxygen Tension

SummaryThe optimum oxygen tension for culturing mammalian embryos has been widely debated by the scientific community. While several laboratories have moved to using 5% as the value for oxygen tension, the majority of modern in vitro fertilization (IVF) laboratory programmes still use 20%. Several in vivo studies have shown the oxygen tension measured in the oviduct of mammals fluctuates between 2% and 8% and in cows and primates this values drops to <2% in the uterine milieu. In human IVF, a non-physiological level of 20% oxygen has been used in the past. However, several studies have shown that atmospheric oxygen introduces adverse effects to embryo development, not limited to numerous molecular and cellular physiology events. In addition, low oxygen tension plays a critical role in reducing the high level of detrimental reactive oxygen species within cells, influences embryonic gene expression, helps with embryo metabolism of glucose, and enhances embryo development to the blastocyst stage. Collectively, this improves embryo implantation potential. However, clinical studies have yielded contradictory results. In almost all reports, some level of improvement has been identified in embryo development or implantation, without any observed drawbacks. This review article will examine the recent literature and discusses ongoing efforts to understand the benefits that low oxygen tension can bring to mammal embryo development in vitro.

scholarly journals Oxygen Tension Strongly Influences Metabolic Parameters and the Release of Interleukin-6 of Human Amniotic Mesenchymal Stromal Cells In Vitro

2018 ◽  
Vol 2018 ◽  
pp. 1-11 ◽  
Author(s):  
Asmita Banerjee ◽  
Andrea Lindenmair ◽  
Ralf Steinborn ◽  
Sergiu Dan Dumitrescu ◽  
Simone Hennerbichler ◽  
...  
Keyword(s):  
Oxygen Tension ◽  
Mesenchymal Stromal Cells ◽  
Stromal Cells ◽  
Human Amniotic Membrane ◽  
Low Oxygen ◽  
Inflammatory Parameters ◽  
Short Time ◽  
Amniotic Cells

The human amniotic membrane (hAM) has been used for tissue regeneration for over a century. In vivo (in utero), cells of the hAM are exposed to low oxygen tension (1–4% oxygen), while the hAM is usually cultured in atmospheric, meaning high, oxygen tension (20% oxygen). We tested the influence of oxygen tensions on mitochondrial and inflammatory parameters of human amniotic mesenchymal stromal cells (hAMSCs). Freshly isolated hAMSCs were incubated for 4 days at 5% and 20% oxygen. We found 20% oxygen to strongly increase mitochondrial oxidative phosphorylation, especially in placental amniotic cells. Oxygen tension did not impact levels of reactive oxygen species (ROS); however, placental amniotic cells showed lower levels of ROS, independent of oxygen tension. In contrast, the release of nitric oxide was independent of the amniotic region but dependent on oxygen tension. Furthermore, IL-6 was significantly increased at 20% oxygen. To conclude, short-time cultivation at 20% oxygen of freshly isolated hAMSCs induced significant changes in mitochondrial function and release of IL-6. Depending on the therapeutic purpose, cultivation conditions of the cells should be chosen carefully for providing the best possible quality of cell therapy.

scholarly journals Comparative Genomics Shows Differences in the Electron Transport and Carbon Metabolic Pathways of Mycobacterium africanum relative to Mycobacterium tuberculosis and suggests an adaptation to low oxygen tension

2019 ◽  
Author(s):  
Boatema Ofori-Anyinam ◽  
Abi Janet Riley ◽  
Tijan Jobarteh ◽  
Ensa Gitteh ◽  
Binta Sarr ◽  
...  
Keyword(s):  
Electron Transport ◽  
Oxygen Tension ◽  
Significant Proportion ◽  
Sensu Stricto ◽  
Low Oxygen ◽  
Low Oxygen Tension ◽  
Genes Encoding ◽  
Mycobacterium Africanum

SummaryThe geographically restricted Mycobacterium africanum lineages (MAF) are primarily found in West Africa, where they account for a significant proportion of tuberculosis. Despite this phenomenon, little is known about the co-evolution of these ancient lineages with West Africans. MAF and M. tuberculosis sensu stricto lineages (MTB) differ in their clinical, in vitro and in vivo characteristics for reasons not fully understood. Therefore, we compared genomes of 289 MAF and 205 MTB clinical isolates from the 6 main human-adapted M. tuberculosis complex lineages, for mutations in their Electron Transport Chain and Central Carbon Metabolic pathway in order to explain these metabolic differences. Furthermore, we determined, in silico, whether each mutation could affect the function of genes encoding enzymes in these pathways.We found more mutations with the potential to affect enzymes in these pathways in MAF lineages compared to MTB lineages. We also found that similar mutations occurred in these pathways between MAF and some MTB lineages.Generally, our findings show further differences between MAF and MTB lineages that may have contributed to the MAF clinical and growth phenotype and indicate potential adaptation of MAF lineages to a distinct ecological niche, which we suggest includes areas characterized by low oxygen tension.

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