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Article ; Online: Reduced DNAJC3 Expression Affects Protein Translocation across the ER Membrane and Attenuates the Down-Modulating Effect of the Translocation Inhibitor Cyclotriazadisulfonamide.

Pauwels, Eva / Provinciael, Becky / Camps, Anita / Hartmann, Enno / Vermeire, Kurt

International journal of molecular sciences

2022 Volume 23, Issue 2

Abstract: One of the reported substrates for the endoplasmic reticulum (ER) translocation inhibitor cyclotriazadisulfonamide (CADA) is DNAJC3, a chaperone of the unfolded protein response during ER stress. In this study, we investigated the impact of altered ... ...

Abstract	One of the reported substrates for the endoplasmic reticulum (ER) translocation inhibitor cyclotriazadisulfonamide (CADA) is DNAJC3, a chaperone of the unfolded protein response during ER stress. In this study, we investigated the impact of altered DNAJC3 protein levels on the inhibitory activity of CADA. By comparing WT DNAJC3 with a CADA-resistant DNAJC3 mutant, we observed the enhanced sensitivity of human CD4, PTK7 and ERLEC1 for CADA when DNAJC3 was expressed at high levels. Combined treatment of CADA with a proteasome inhibitor resulted in synergistic inhibition of protein translocation and in the rescue of a small preprotein fraction, which presumably corresponds to the CADA affected protein fraction that is stalled at the Sec61 translocon. We demonstrate that DNAJC3 enhances the protein translation of a reporter protein that is expressed downstream of the CADA-stalled substrate, suggesting that DNAJC3 promotes the clearance of the clogged translocon. We propose a model in which a reduced DNAJC3 level by CADA slows down the clearance of CADA-stalled substrates. This results in higher residual translocation into the ER lumen due to the longer dwelling time of the temporarily stalled substrates in the translocon. Thus, by directly reducing DNAJC3 protein levels, CADA attenuates its net down-modulating effect on its substrates.
MeSH term(s)	Endoplasmic Reticulum/metabolism ; Endoplasmic Reticulum Stress ; HEK293 Cells ; HSP40 Heat-Shock Proteins/metabolism ; Humans ; Protein Transport ; SEC Translocation Channels/metabolism ; Unfolded Protein Response
Chemical Substances	DNAJC3 protein, human ; HSP40 Heat-Shock Proteins ; SEC Translocation Channels
Language	English
Publishing date	2022-01-06
Publishing country	Switzerland
Document type	Journal Article
ZDB-ID	2019364-6
ISSN	1422-0067 ; 1422-0067 ; 1661-6596
ISSN (online)	1422-0067
ISSN	1422-0067 ; 1661-6596
DOI	10.3390/ijms23020584
Database	MEDical Literature Analysis and Retrieval System OnLINE

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Article ; Online: Inhibitors of the Sec61 Complex and Novel High Throughput Screening Strategies to Target the Protein Translocation Pathway.

Pauwels, Eva / Schülein, Ralf / Vermeire, Kurt

International journal of molecular sciences

2021 Volume 22, Issue 21

Abstract: Proteins targeted to the secretory pathway start their intracellular journey by being transported across biological membranes such as the endoplasmic reticulum (ER). A central component in this protein translocation process across the ER is the Sec61 ... ...

Abstract	Proteins targeted to the secretory pathway start their intracellular journey by being transported across biological membranes such as the endoplasmic reticulum (ER). A central component in this protein translocation process across the ER is the Sec61 translocon complex, which is only intracellularly expressed and does not have any enzymatic activity. In addition, Sec61 translocon complexes are difficult to purify and to reconstitute. Screening for small molecule inhibitors impairing its function has thus been notoriously difficult. However, such translocation inhibitors may not only be valuable tools for cell biology, but may also represent novel anticancer drugs, given that cancer cells heavily depend on efficient protein translocation into the ER to support their fast growth. In this review, different inhibitors of protein translocation will be discussed, and their specific mode of action will be compared. In addition, recently published screening strategies for small molecule inhibitors targeting the whole SRP-Sec61 targeting/translocation pathway will be summarized. Of note, slightly modified assays may be used in the future to screen for substances affecting SecYEG, the bacterial ortholog of the Sec61 complex, in order to identify novel antibiotic drugs.
MeSH term(s)	Animals ; Anti-Bacterial Agents/pharmacology ; Antineoplastic Agents/pharmacology ; Antiviral Agents/pharmacology ; Endoplasmic Reticulum/drug effects ; Endoplasmic Reticulum/metabolism ; High-Throughput Screening Assays ; Humans ; Protein Transport ; SEC Translocation Channels/antagonists & inhibitors ; SEC Translocation Channels/metabolism
Chemical Substances	Anti-Bacterial Agents ; Antineoplastic Agents ; Antiviral Agents ; SEC Translocation Channels
Language	English
Publishing date	2021-11-05
Publishing country	Switzerland
Document type	Journal Article ; Review
ZDB-ID	2019364-6
ISSN	1422-0067 ; 1422-0067 ; 1661-6596
ISSN (online)	1422-0067
ISSN	1422-0067 ; 1661-6596
DOI	10.3390/ijms222112007
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Article ; Online: Intracellular flow cytometry complements RT-qPCR detection of circulating SARS-CoV-2 variants of concern.

Vanhulle, Emiel / Provinciael, Becky / Stroobants, Joren / Camps, Anita / Maes, Piet / Vermeire, Kurt

BioTechniques

2022 Volume 72, Issue 6, Page(s) 245–254

Abstract: Basic and antiviral research on SARS-CoV-2 rely on cellular assays of virus ... ...

Abstract	Basic and antiviral research on SARS-CoV-2 rely on cellular assays of virus replication
MeSH term(s)	Animals ; COVID-19/diagnosis ; Chlorocebus aethiops ; Flow Cytometry ; Humans ; SARS-CoV-2/genetics ; Vero Cells
Language	English
Publishing date	2022-04-21
Publishing country	England
Document type	Journal Article
ZDB-ID	48453-2
ISSN	1940-9818 ; 0736-6205
ISSN (online)	1940-9818
ISSN	0736-6205
DOI	10.2144/btn-2022-0018
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Article ; Online: Above-ground plant properties are not leading indicators of grazing-induced soil carbon accrual in the Northern Great Plains

Kurt O. Reinhart / Matthew J. Rinella / Richard C. Waterman / Hilaire S. Sanni Worogo / Lance T. Vermeire

Ecological Indicators, Vol 158, Iss , Pp 111509- (2024)

2024

Abstract: A new aim for grassland management is to increase soil organic carbon (SOC) and to offset CO2 emissions by companies. This practice of carbon ranching may be informed by grazing-induced shifts in plant biomass and diversity which may foretell changes in ... ...

Abstract	A new aim for grassland management is to increase soil organic carbon (SOC) and to offset CO2 emissions by companies. This practice of carbon ranching may be informed by grazing-induced shifts in plant biomass and diversity which may foretell changes in SOC. Unfortunately, little is known about how grazing-induced shifts in plant properties correspond with shifts in SOC stocks. To help fill this gap, we used data from a field experiment to test whether above-ground plant properties (i.e. biomass, species richness) act as leading indicators of grazing-induced SOC accrual in the Northern Great Plains. The 5-yr bovine grazing experiment had a randomized complete block design and pre-treatment data. Moderate summer grazing (control) is widely used in the Northern Great Plains, and treatments that may alter grassland vegetation and SOC included: severe summer grazing, moderate fall grazing, and severe fall grazing. Severe fall and summer grazing increased SOC but had no effect on plant species richness and biomass relative to controls. Fall moderate grazing increased above-ground plant biomass but had no effect on SOC relative to controls. Changes to grazing practices can affect SOC without measurably affecting plant properties and can affect plant properties without measurably affecting SOC. While two drivers of SOC are plant carbon inputs and microbial respiration, our study indicates that grazing-induced change in above-ground vegetation is not predictive of change in SOC.
Keywords	Carbon dioxide removal ; Carbon ranching ; Carbon sequestration ; Climate change ; Co-benefits ; Livestock grazing ; Ecology ; QH540-549.5
Subject code	580 ; 333
Language	English
Publishing date	2024-01-01T00:00:00Z
Publisher	Elsevier
Document type	Article ; Online
Database	BASE - Bielefeld Academic Search Engine (life sciences selection)

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Article ; Online: Effects of mowing, spring precipitation, soil nutrients, and enzymes on grassland productivity

Kurt O. Reinhart / Kim J. Komatsu / Lance T. Vermeire

Agrosystems, Geosciences & Environment, Vol 5, Iss 4, Pp n/a-n/a (2022)

2022

Abstract: Abstract Little research has assessed how the timing and intensity of grazing might affect plant biomass, available nutrients, soil extracellular enzyme activity (EEA), and how climate change might influence these responses. We tested the effect of two ... ...

Abstract	Abstract Little research has assessed how the timing and intensity of grazing might affect plant biomass, available nutrients, soil extracellular enzyme activity (EEA), and how climate change might influence these responses. We tested the effect of two spring precipitation schemes (70, 100% of ambient), two mowing intensities (moderate, severe), and two mowing season treatments (June, October) on plant and soil properties. We detected an interactive effect of precipitation, mowing intensity, and mowing season on plant biomass. When plots were mowed at a moderate intensity, water reductions had irregular effects on plant biomass depending on the mowing season. Plant biomass was also 11% greater in plots mowed at moderate than at severe intensities. Most soil nutrients were unaffected by treatments except for calcium. Soil EEA was unaffected by treatments; however, the activity of a phosphorus (P)‐acquisition enzyme was more than four times greater than the activity of nitrogen (N)‐ and carbon‐acquisition enzymes. A substantial amount (adjusted R2 = .51) of plot‐to‐plot variation in plant biomass was explained by three soil properties especially an N‐acquisition enzyme and to a lesser degree by plant available P and soil pH. The grassland had a high degree of natural buffering capacity as most soil properties were resistant to shifts in 6‐yr spring precipitation and 5‐yr simulated grazing intensity and season. Grassland plant biomass varied by treatments and was seemingly limited by biogeochemical constraints especially the prevalent need to mobilize P and a secondary need to acquire N as plant biomass increased.
Keywords	Agriculture ; S ; Environmental sciences ; GE1-350
Language	English
Publishing date	2022-01-01T00:00:00Z
Publisher	Wiley
Document type	Article ; Online
Database	BASE - Bielefeld Academic Search Engine (life sciences selection)

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Article ; Online: Reduced DNAJC3 Expression Affects Protein Translocation across the ER Membrane and Attenuates the Down-Modulating Effect of the Translocation Inhibitor Cyclotriazadisulfonamide

Eva Pauwels / Becky Provinciael / Anita Camps / Enno Hartmann / Kurt Vermeire

International Journal of Molecular Sciences, Vol 23, Iss 584, p

2022 Volume 584

Abstract	One of the reported substrates for the endoplasmic reticulum (ER) translocation inhibitor cyclotriazadisulfonamide (CADA) is DNAJC3, a chaperone of the unfolded protein response during ER stress. In this study, we investigated the impact of altered DNAJC3 protein levels on the inhibitory activity of CADA. By comparing WT DNAJC3 with a CADA-resistant DNAJC3 mutant, we observed the enhanced sensitivity of human CD4, PTK7 and ERLEC1 for CADA when DNAJC3 was expressed at high levels. Combined treatment of CADA with a proteasome inhibitor resulted in synergistic inhibition of protein translocation and in the rescue of a small preprotein fraction, which presumably corresponds to the CADA affected protein fraction that is stalled at the Sec61 translocon. We demonstrate that DNAJC3 enhances the protein translation of a reporter protein that is expressed downstream of the CADA-stalled substrate, suggesting that DNAJC3 promotes the clearance of the clogged translocon. We propose a model in which a reduced DNAJC3 level by CADA slows down the clearance of CADA-stalled substrates. This results in higher residual translocation into the ER lumen due to the longer dwelling time of the temporarily stalled substrates in the translocon. Thus, by directly reducing DNAJC3 protein levels, CADA attenuates its net down-modulating effect on its substrates.
Keywords	co-translational translocation ; endoplasmic reticulum ; cyclotriazadisulfonamide ; ER quality control ; DNAJC3 ; signal peptide ; Biology (General) ; QH301-705.5 ; Chemistry ; QD1-999
Subject code	570
Language	English
Publishing date	2022-01-01T00:00:00Z
Publisher	MDPI AG
Document type	Article ; Online
Database	BASE - Bielefeld Academic Search Engine (life sciences selection)

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Article ; Online: Inhibitors of protein translocation across membranes of the secretory pathway: novel antimicrobial and anticancer agents.

Van Puyenbroeck, Victor / Vermeire, Kurt

Cellular and molecular life sciences : CMLS

2018 Volume 75, Issue 9, Page(s) 1541–1558

Abstract: Proteins routed to the secretory pathway start their journey by being transported across biological membranes, such as the endoplasmic reticulum. The essential nature of this protein translocation process has led to the evolution of several factors that ... ...

Abstract	Proteins routed to the secretory pathway start their journey by being transported across biological membranes, such as the endoplasmic reticulum. The essential nature of this protein translocation process has led to the evolution of several factors that specifically target the translocon and block translocation. In this review, various translocation pathways are discussed together with known inhibitors of translocation. Properties of signal peptide-specific systems are highlighted for the development of new therapeutic and antimicrobial applications, as compounds can target signal peptides from either host cells or pathogens and thereby selectively prevent translocation of those specific proteins. Broad inhibition of translocation is also an interesting target for the development of new anticancer drugs because cancer cells heavily depend on efficient protein translocation into the endoplasmic reticulum to support their fast growth.
MeSH term(s)	Animals ; Anti-Infective Agents/pharmacology ; Antineoplastic Agents/pharmacology ; Drug Discovery/methods ; Endoplasmic Reticulum/drug effects ; Endoplasmic Reticulum/metabolism ; Humans ; Models, Molecular ; Molecular Targeted Therapy/methods ; Neoplasms/drug therapy ; Protein Sorting Signals/drug effects ; Protein Transport/drug effects
Chemical Substances	Anti-Infective Agents ; Antineoplastic Agents ; Protein Sorting Signals
Language	English
Publishing date	2018-01-05
Publishing country	Switzerland
Document type	Journal Article ; Research Support, Non-U.S. Gov't ; Review
ZDB-ID	1358415-7
ISSN	1420-9071 ; 1420-682X
ISSN (online)	1420-9071
ISSN	1420-682X
DOI	10.1007/s00018-017-2743-2
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Zs.A 195: Show issues

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Article ; Online: Inhibitors of the Sec61 Complex and Novel High Throughput Screening Strategies to Target the Protein Translocation Pathway

Eva Pauwels / Ralf Schülein / Kurt Vermeire

International Journal of Molecular Sciences, Vol 22, Iss 12007, p

2021 Volume 12007

Abstract	Proteins targeted to the secretory pathway start their intracellular journey by being transported across biological membranes such as the endoplasmic reticulum (ER). A central component in this protein translocation process across the ER is the Sec61 translocon complex, which is only intracellularly expressed and does not have any enzymatic activity. In addition, Sec61 translocon complexes are difficult to purify and to reconstitute. Screening for small molecule inhibitors impairing its function has thus been notoriously difficult. However, such translocation inhibitors may not only be valuable tools for cell biology, but may also represent novel anticancer drugs, given that cancer cells heavily depend on efficient protein translocation into the ER to support their fast growth. In this review, different inhibitors of protein translocation will be discussed, and their specific mode of action will be compared. In addition, recently published screening strategies for small molecule inhibitors targeting the whole SRP-Sec61 targeting/translocation pathway will be summarized. Of note, slightly modified assays may be used in the future to screen for substances affecting SecYEG, the bacterial ortholog of the Sec61 complex, in order to identify novel antibiotic drugs.
Keywords	signal recognition particle dependent protein targeting ; Sec61 dependent translocation ; co-translational translocation ; endoplasmic reticulum ; inhibitor ; high throughput screening ; Biology (General) ; QH301-705.5 ; Chemistry ; QD1-999
Subject code	570
Language	English
Publishing date	2021-11-01T00:00:00Z
Publisher	MDPI AG
Document type	Article ; Online
Database	BASE - Bielefeld Academic Search Engine (life sciences selection)

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Article ; Online: SARS-CoV-2 Permissive glioblastoma cell line for high throughput antiviral screening.

Vanhulle, Emiel / Stroobants, Joren / Provinciael, Becky / Camps, Anita / Noppen, Sam / Maes, Piet / Vermeire, Kurt

Antiviral research

2022 Volume 203, Page(s) 105342

Abstract: Despite the great success of the administered vaccines against SARS-CoV-2, the virus can still spread, as evidenced by the current circulation of the highly contagious Omicron variant. This emphasizes the additional need to develop effective antiviral ... ...

Abstract	Despite the great success of the administered vaccines against SARS-CoV-2, the virus can still spread, as evidenced by the current circulation of the highly contagious Omicron variant. This emphasizes the additional need to develop effective antiviral countermeasures. In the context of early preclinical studies for antiviral assessment, robust cellular infection systems are required to screen drug libraries. In this study, we reported the implementation of a human glioblastoma cell line, stably expressing ACE2, in a SARS-CoV-2 cytopathic effect (CPE) reduction assay. These glioblastoma cells, designated as U87.ACE2
MeSH term(s)	Angiotensin-Converting Enzyme 2 ; Antiviral Agents/pharmacology ; COVID-19 Vaccines ; Cell Line ; Glioblastoma/drug therapy ; High-Throughput Screening Assays ; Humans ; Peptidyl-Dipeptidase A/metabolism ; SARS-CoV-2 ; Spike Glycoprotein, Coronavirus/metabolism ; COVID-19 Drug Treatment
Chemical Substances	Antiviral Agents ; COVID-19 Vaccines ; Spike Glycoprotein, Coronavirus ; spike protein, SARS-CoV-2 ; Peptidyl-Dipeptidase A (EC 3.4.15.1) ; Angiotensin-Converting Enzyme 2 (EC 3.4.17.23)
Language	English
Publishing date	2022-05-18
Publishing country	Netherlands
Document type	Journal Article
ZDB-ID	306628-9
ISSN	1872-9096 ; 0166-3542
ISSN (online)	1872-9096
ISSN	0166-3542
DOI	10.1016/j.antiviral.2022.105342
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Zs.A 1627: Show issues

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Article ; Online: Advancing Marburg virus antiviral screening: Optimization of a novel T7 polymerase-independent minigenome system.

Vanmechelen, Bert / Stroobants, Joren / Vermeire, Kurt / Maes, Piet

Antiviral research

2020 Volume 185, Page(s) 104977

Abstract: Marburg virus (MARV) is the only known pathogenic filovirus not belonging to the genus Ebolavirus. Minigenomes have proven a useful tool to study MARV, but all existing MARV minigenomes are dependent on the addition of an exogenous T7 RNA polymerase to ... ...

Abstract	Marburg virus (MARV) is the only known pathogenic filovirus not belonging to the genus Ebolavirus. Minigenomes have proven a useful tool to study MARV, but all existing MARV minigenomes are dependent on the addition of an exogenous T7 RNA polymerase to drive minigenome expression. However, exogenous expression of a T7 polymerase is not always feasible and can act as a confounding factor in compound screening assays. We have developed an alternative minigenome that is controlled by the natively expressed RNA polymerase II. We demonstrate here the characteristics of this new system and its applicability in a wide range of cell types. Our system shows a clear concentration-dependent activity and shows comparable activity to the existing T7 polymerase-based system at higher concentrations, also in difficult-to-transfect cell lines. In addition, we show that our system can be used for compound screening in a 96-well format, thereby providing an attractive alternative to previously developed MARV minigenomes.
MeSH term(s)	Animals ; Antiviral Agents/pharmacology ; Cell Line ; Cell Line, Tumor ; Chiroptera ; Chlorocebus aethiops ; Cricetinae ; DNA-Directed RNA Polymerases/genetics ; Genome, Viral ; HEK293 Cells ; High-Throughput Screening Assays/methods ; Humans ; Kidney/cytology ; Marburgvirus/drug effects ; Marburgvirus/genetics ; Promoter Regions, Genetic ; Transcription, Genetic ; Vero Cells ; Viral Proteins/genetics ; Virus Replication/drug effects
Chemical Substances	Antiviral Agents ; Viral Proteins ; bacteriophage T7 RNA polymerase (EC 2.7.7.-) ; DNA-Directed RNA Polymerases (EC 2.7.7.6)
Language	English
Publishing date	2020-11-19
Publishing country	Netherlands
Document type	Journal Article ; Research Support, Non-U.S. Gov't
ZDB-ID	306628-9
ISSN	1872-9096 ; 0166-3542
ISSN (online)	1872-9096
ISSN	0166-3542
DOI	10.1016/j.antiviral.2020.104977
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