scholarly journals Treatment with surfactants enables quantification of translational activity by O-propargyl-puromycin labelling in yeast

2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Jennifer Staudacher ◽  
Corinna Rebnegger ◽  
Brigitte Gasser
Keyword(s):  
Mammalian Cells ◽  
Growth Conditions ◽  
Non Invasive ◽  
Translational Activity ◽  
Wide Range ◽  
Simple Protocol ◽  
Set Up ◽  
Different Strains ◽  
Negative Controls ◽  
Global Translation

Abstract Background Translation is an important point of regulation in protein synthesis. However, there is a limited number of methods available to measure global translation activity in yeast. Recently, O-propargyl-puromycin (OPP) labelling has been established for mammalian cells, but unmodified yeasts are unsusceptible to puromycin. Results We could increase susceptibility by using a Komagataella phaffii strain with an impaired ergosterol pathway (erg6Δ), but translation measurements are restricted to this strain background, which displayed growth deficits. Using surfactants, specifically Imipramine, instead, proved to be more advantageous and circumvents previous restrictions. Imipramine-supplemented OPP-labelling with subsequent flow cytometry analysis, enabled us to distinguish actively translating cells from negative controls, and to clearly quantify differences in translation activities in different strains and growth conditions. Specifically, we investigated K. phaffii at different growth rates, verified that methanol feeding alters translation activity, and analysed global translation in strains with genetically modified stress response pathways. Conclusions We set up a simple protocol to measure global translation activity in yeast on a single cell basis. The use of surfactants poses a practical and non-invasive alternative to the commonly used ergosterol pathway impaired strains and thus impacts a wide range of applications where increased drug and dye uptake is needed.

scholarly journals Co-feeding glucose with either gluconate or galacturonate during clostridial fermentations provides metabolic fine-tuning capabilities

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Theresah N. K. Zu ◽  
Sanchao Liu ◽  
Elliot S. Gerlach ◽  
Wais Mojadedi ◽  
Christian J. Sund
Keyword(s):  
Growth Conditions ◽  
Fine Tuning ◽  
Oxidation States ◽  
Mixed Fermentation ◽  
Wide Range ◽  
Commodity Chemicals ◽  
Different Strains ◽  
Metabolic Output ◽  
Electron Carriers ◽  
Strain Dependent

AbstractClostridiumacetobutylicum ATCC 824 effectively utilizes a wide range of substrates to produce commodity chemicals. When grown on substrates of different oxidation states, the organism exhibits different recycling needs of reduced intracellular electron carrying co-factors. Ratios of substrates with different oxidation states were used to modulate the need to balance electron carriers and demonstrate fine-tuned control of metabolic output. Three different oxidized substrates were first fed singularly, then in different ratios to three different strains of Clostridium sp. Growth was most robust when fed glucose in exclusive fermentations. However, the use of the other two more oxidized substrates was strain-dependent in exclusive feeds. In glucose-galacturonate mixed fermentation, the main products (acetate and butyrate) were dependant on the ratios of the substrates. Exclusive fermentation on galacturonate was nearly homoacetic. Co-utilization of galacturonate and glucose was observed from the onset of fermentation in growth conditions using both substrates combined, with the proportion of galacturonate present dictating the amount of acetate produced. For all three strains, increasing galacturonate content (%) in a mixture of galacturonate and glucose from 0 to 50, and 100, resulted in a corresponding increase in the amount of acetate produced. For example, C.acetobutylicum increased from ~ 10 mM to ~ 17 mM, and then ~ 23 mM. No co-utilization was observed when galacturonate was replaced with gluconate in the two substrate co-feed.

Settlers or Movers? The Temporality of Past Migrations, Political Inaction and its Consequences, 1945–1985

2018 ◽  
Author(s):  
Jozefien De Bock
Keyword(s):  
Temporary Workers ◽  
Guest Workers ◽  
Temporary Migration ◽  
Wide Range ◽  
Settler Societies ◽  
Future Policy ◽  
Set Up ◽  
Legal Frameworks ◽  
Temporary Migrants ◽  
Multicultural Policies

Historically, those societies that have the longest tradition in multicultural policies are settler societies. The question of how to deal with temporary migrants has only recently aroused their interest. In Europe, temporary migration programmes have a much longer history. In the period after WWII, a wide range of legal frameworks were set up to import temporary workers, who came to be known as guest workers. In the end, many of these ‘guests’ settled in Europe permanently. Their presence lay at the basis of European multicultural policies. However, when these policies were drafted, the former mobility of guest workers had been forgotten. This chapter will focus on this mobility of initially temporary workers, comparing the period of economic growth 1945-1974 with the years after the 1974 economic crisis. Further, it will look at the kind of policies that were developed towards guest workers in the era before multiculturalism. This way, it shows how their consideration as temporary residents had far-reaching consequences for the immigrants, their descendants and the receiving societies involved. The chapter will finish by suggesting a number of lessons from the past. If the mobility-gap between guest workers and present-day migrants is not as big as generally assumed, then the consequences of previous neglect should serve as a warning for future policy making.

Role of Cardiac MRI in Detecting Familial Hypertrophic Cardiomyopathy: Review

2016 ◽  
Vol 1 (1) ◽  
pp. 4
Author(s):  
Marymol Koshy ◽  
Bushra Johari ◽  
Mohd Farhan Hamdan ◽  
Mohammad Hanafiah
Keyword(s):  
Hypertrophic Cardiomyopathy ◽  
Magnetic Resonance ◽  
Genetic Disorder ◽  
Familial Hypertrophic Cardiomyopathy ◽  
Non Invasive ◽  
Wide Range ◽  
Proper Diagnosis ◽  
Magnetic Resonance Imaging Mri ◽  
Potential Use

Hypertrophic cardiomyopathy (HCM) is a global disease affecting people of various ethnic origins and both genders. HCM is a genetic disorder with a wide range of symptoms, including the catastrophic presentation of sudden cardiac death. Proper diagnosis and treatment of this disorder can relieve symptoms and prolong life. Non-invasive imaging is essential in diagnosing HCM. We present a review to deliberate the potential use of cardiac magnetic resonance (CMR) imaging in HCM assessment and also identify the risk factors entailed with risk stratification of HCM based on Magnetic Resonance Imaging (MRI).

ORAL HISTORY SOURCES AS LEARNING MATERIALS: A CASE STUDY OF THE NATIONAL UNIVERSITY OF SCIENCE AND TECHNOLOGY

2016 ◽  
Vol 3 (2) ◽  
pp. 82-93
Author(s):  
Gugulethu Shamaine Nkala ◽  
Rodreck David
Keyword(s):  
Oral History ◽  
Learning Experience ◽  
Science And Technology ◽  
Group Discussion ◽  
Human Interaction ◽  
Learning Material ◽  
Wide Range ◽  
National Archives ◽  
National University ◽  
Set Up

Knowledge presented by Oral History (OH) is unique in that it shares the tacit perspective, thoughts, opinions and understanding of the interviewee in its primary form. While teachers, lecturers and other education specialists have at their disposal a wide range of primary, secondary and tertiary sources upon which to relate and share or impart knowledge, OH presents a rich source of information that can improve the learning and knowledge impartation experience. The uniqueness of OH is presented in the following advantages of its use: it allows one to learn about the perspectives of individuals who might not otherwise appear in the historical record; it allows one to compensate for the digital age; one can learn different kinds of information; it provides historical actors with an opportunity to tell their own stories in their own words; and it offers a rich opportunity for human interaction. This article discusses the placement of oral history in the classroom set-up by investigating its use as a source of learning material presented by the National Archives of Zimbabwe to students in the Department of Records and Archives Management at the National University of Science and Technology (NUST). Interviews and a group discussion were used to gather data from an archivist at the National Archives of Zimbabwe, lecturers and students in the Department of Records and Archives Management at NUST, respectively. These groups were approached on the usability, uniqueness and other characteristics that support this type of knowledge about OH in a tertiary learning experience. The findings indicate several qualities that reflect the richness of OH as a teaching source material in a classroom set-up. It further points to weak areas that may be addressed where the source is considered a viable strategy for knowledge sharing and learning. The researchers present a possible model that can be used to champion the use of this rich knowledge source in classroom education at this university and in similar set-ups. 

Nanofiltration of endocrine disrupting compounds

2003 ◽  
Vol 3 (5-6) ◽  
pp. 321-327 ◽  
Author(s):  
M. Gallenkemper ◽  
T. Wintgens ◽  
T. Melin
Keyword(s):  
Wastewater Treatment ◽  
Municipal Wastewater ◽  
Wastewater Treatment Plants ◽  
Endocrine Disrupting Compounds ◽  
Municipal Wastewater Treatment ◽  
Water And Wastewater Treatment ◽  
Wide Range ◽  
Endocrine Disrupting ◽  
Municipal Wastewater Treatment Plants ◽  
Set Up

Endocrine disrupting compounds can affect the hormone system in organisms. A wide range of endocrine disrupters were found in sewage and effluents of municipal wastewater treatment plants. Toxicological evaluations indicate that conventional wastewater treatment plants are not able to remove these substances sufficiently before disposing effluent into the environment. Membrane technology, which is proving to be an effective barrier to these substances, is the subject of this research. Nanofiltration provides high quality permeates in water and wastewater treatment. Eleven different nanofiltration membranes were tested in the laboratory set-up. The observed retention for nonylphenol (NP) and bisphenol A (BPA) ranged between 70% and 100%. The contact angle is an indicator for the hydrophobicity of a membrane, whose influence on the permeability and retention of NP was evident. The retention of BPA was found to be inversely proportional to the membrane permeability.

Evaluation of the nucleopolyhedrovirus of Anticarsia gemmatalis as a vector for gene therapy in mammals

2020 ◽  
Vol 20 ◽  
Author(s):  
Cintia N. Parsza ◽  
Diego L. Mengual Gómez ◽  
Jorge Alejandro Simonin ◽  
Mariano Nicolás Belaich ◽  
Pablo Daniel Ghiringhelli
Keyword(s):  
Gene Therapy ◽  
Mammalian Cells ◽  
Insect Cells ◽  
Autographa Californica ◽  
Anticarsia Gemmatalis ◽  
Agricultural Pests ◽  
Mammalian Cell Lines ◽  
Delivery Vector ◽  
Wide Range ◽  
Insect Pathogens

Background: Baculoviruses are insect pathogens with important biotechnological applications that transcend their use as biological controllers of agricultural pests. One species, Autographa californica multiple nucleopolhyedrovirus (AcMNPV) has been extensively exploited as a molecular platform to produce recombinant proteins and as a delivery vector for genes in mammals, because it can transduce a wide range of mammalian cells and tissues without replicating or producing progeny. Objective/Method: To investigate if the budded virions of Anticarsia gemmatalis multiple nucleopolhyedrovirus (AgMNPV) species has the same ability, the viral genome was modified by homologous recombination into susceptible insect cells to integrate reporter genes and then it was evaluated on mammalian cell lines in comparative form with respect to equivalent viruses derived from AcMNPV. Besides, the replicative capacity of AgMNPV´s virions in mammals was determined. Results: The experiments carried out showed that the recombinant variant of AgMNPV transduces and support the expression of delivered genes but not replicates in mammalian cells. Conclusion: Consequently, this insect pathogen is proposed as an alternative of non-infectious viruses in humans to explore new approaches in gene therapy and other applications based on the use of mammalian cells.

scholarly journals A Metzincin and TIMP-Like Protein Pair of a Phage Origin Sensitize Listeria monocytogenes to Phage Lysins and Other Cell Wall Targeting Agents

2021 ◽  
Vol 9 (6) ◽  
pp. 1323
Author(s):  
Etai Boichis ◽  
Nadejda Sigal ◽  
Ilya Borovok ◽  
Anat A. Herskovits
Keyword(s):  
Cell Wall ◽  
Listeria Monocytogenes ◽  
Mammalian Cells ◽  
Protein Pair ◽  
Growth Conditions ◽  
Wall Structure ◽  
Antibiotic Resistant ◽  
Extracellular Milieu ◽  
Virion Release ◽  
Genes Encoding

Infection of mammalian cells by Listeria monocytogenes (Lm) was shown to be facilitated by its phage elements. In a search for additional phage remnants that play a role in Lm’s lifecycle, we identified a conserved locus containing two XRE regulators and a pair of genes encoding a secreted metzincin protease and a lipoprotein structurally similar to a TIMP-family metzincin inhibitor. We found that the XRE regulators act as a classic CI/Cro regulatory switch that regulates the expression of the metzincin and TIMP-like genes under intracellular growth conditions. We established that when these genes are expressed, their products alter Lm morphology and increase its sensitivity to phage mediated lysis, thereby enhancing virion release. Expression of these proteins also sensitized the bacteria to cell wall targeting compounds, implying that they modulate the cell wall structure. Our data indicate that these effects are mediated by the cleavage of the TIMP-like protein by the metzincin, and its subsequent release to the extracellular milieu. While the importance of this locus to Lm pathogenicity remains unclear, the observation that this phage-associated protein pair act upon the bacterial cell wall may hold promise in the field of antibiotic potentiation to combat antibiotic resistant bacterial pathogens.

scholarly journals A System-Agnostic, Adaptable and Extensible Animal Support Cradle System for Cardio-Respiratory-Synchronised, and Other, Multi-Modal Imaging of Small Animals

Tomography ◽  
2021 ◽  
Vol 7 (1) ◽  
pp. 39-54
Author(s):  
Veerle Kersemans ◽  
Stuart Gilchrist ◽  
Philip Danny Allen ◽  
Sheena Wallington ◽  
Paul Kinchesh ◽  
...  
Keyword(s):  
Scientific Data ◽  
Cardiac Monitoring ◽  
Small Animals ◽  
Maintenance System ◽  
Wide Range ◽  
Set Up ◽  
Animal Handling ◽  
Imaging Performance ◽  
Significant Effort

Standardisation of animal handling procedures for a wide range of preclinical imaging scanners will improve imaging performance and reproducibility of scientific data. Whilst there has been significant effort in defining how well scanners should operate and how in vivo experimentation should be practised, there is little detail on how to achieve optimal scanner performance with best practices in animal welfare. Here, we describe a system-agnostic, adaptable and extensible animal support cradle system for cardio-respiratory-synchronised, and other, multi-modal imaging of small animals. The animal support cradle can be adapted on a per application basis and features integrated tubing for anaesthetic and tracer delivery, an electrically driven rectal temperature maintenance system and respiratory and cardiac monitoring. Through a combination of careful material and device selection, we have described an approach that allows animals to be transferred whilst under general anaesthesia between any of the tomographic scanners we currently or have previously operated. The set-up is minimally invasive, cheap and easy to implement and for multi-modal, multi-vendor imaging of small animals.

scholarly journals Autophagy Deficiency by Atg4B Loss Leads to Metabolomic Alterations in Mice

Metabolites ◽  
2021 ◽  
Vol 11 (8) ◽  
pp. 481
Author(s):  
Gemma G. Martínez-García ◽  
Raúl F. Pérez ◽  
Álvaro F. Fernández ◽  
Sylvere Durand ◽  
Guido Kroemer ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Amino Acids ◽  
Mammalian Cells ◽  
Tissue Homeostasis ◽  
In Vivo Studies ◽  
Protective Mechanism ◽  
Cardiac Damage ◽  
Wide Range ◽  
First Time

Autophagy is an essential protective mechanism that allows mammalian cells to cope with a variety of stressors and contributes to maintaining cellular and tissue homeostasis. Due to these crucial roles and also to the fact that autophagy malfunction has been described in a wide range of pathologies, an increasing number of in vivo studies involving animal models targeting autophagy genes have been developed. In mammals, total autophagy inactivation is lethal, and constitutive knockout models lacking effectors of this route are not viable, which has hindered so far the analysis of the consequences of a systemic autophagy decline. Here, we take advantage of atg4b−/− mice, an autophagy-deficient model with only partial disruption of the process, to assess the effects of systemic reduction of autophagy on the metabolome. We describe for the first time the metabolic footprint of systemic autophagy decline, showing that impaired autophagy results in highly tissue-dependent alterations that are more accentuated in the skeletal muscle and plasma. These changes, which include changes in the levels of amino-acids, lipids, or nucleosides, sometimes resemble those that are frequently described in conditions like aging, obesity, or cardiac damage. We also discuss different hypotheses on how impaired autophagy may affect the metabolism of several tissues in mammals.

scholarly journals SARS-CoV-2 specific antibody and neutralization assays reveal the wide range of the humoral immune response to virus

2021 ◽  
Vol 4 (1) ◽  
Author(s):  
Mikail Dogan ◽  
Lina Kozhaya ◽  
Lindsey Placek ◽  
Courtney Gunter ◽  
Mesut Yigit ◽  
...  
Keyword(s):  
Specific Antibody ◽  
Neutralizing Antibodies ◽  
Vaccine Development ◽  
High Specificity ◽  
Spike Protein ◽  
Humoral Immune ◽  
Specificity And Sensitivity ◽  
Wide Range ◽  
Specific Igg ◽  
Negative Controls

AbstractDevelopment of antibody protection during SARS-CoV-2 infection is a pressing question for public health and for vaccine development. We developed highly sensitive SARS-CoV-2-specific antibody and neutralization assays. SARS-CoV-2 Spike protein or Nucleocapsid protein specific IgG antibodies at titers more than 1:100,000 were detectable in all PCR+ subjects (n = 115) and were absent in the negative controls. Other isotype antibodies (IgA, IgG1-4) were also detected. SARS-CoV-2 neutralization was determined in COVID-19 and convalescent plasma at up to 10,000-fold dilution, using Spike protein pseudotyped lentiviruses, which were also blocked by neutralizing antibodies (NAbs). Hospitalized patients had up to 3000-fold higher antibody and neutralization titers compared to outpatients or convalescent plasma donors. Interestingly, some COVID-19 patients also possessed NAbs against SARS-CoV Spike protein pseudovirus. Together these results demonstrate the high specificity and sensitivity of our assays, which may impact understanding the quality or duration of the antibody response during COVID-19 and in determining the effectiveness of potential vaccines.

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