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Article ; Online: Plastid-encoded RNA polymerase variation in Pelargonium sect Ciconium

Breman, F.C. / Korver, J.W. / Snijder, R.C. / Villard, C. / Schranz, M.E. / Bakker, F.T.

2024 Volume 2

Abstract: Cyto-Nuclear Incompatibility (CNI), in which there is a mismatch in the interaction between organelles and nucleus, impacts plant species evolution as it has a direct effect on the fitness of plants. It can reduce fertility and/or result in bleached ... ...

Abstract	Cyto-Nuclear Incompatibility (CNI), in which there is a mismatch in the interaction between organelles and nucleus, impacts plant species evolution as it has a direct effect on the fitness of plants. It can reduce fertility and/or result in bleached plants devoid of functional chloroplasts. Understanding the processes leading to CNI could help to improve breeding efforts, especially in cases where species with desirable traits need to be crossed into existing cultivars. To better understand the occurrence of CNI and its effects on plant phenotype, we combined near comprehensive crossing series across a clade of species from Pelargonium section Ciconium with comparative genomics and protein modelling for plastid-encoded RNA polymerase (PEP), as the rpo genes encoding PEP subunits were found to be unusually highly divergent, especially in two length-variable regions. Of all plastome-encoded genes, we found these genes to contain more variation than observed across angiosperms and that this underlies structural variation inferred for PEP in P. sect. Ciconium. This variation, resulting in differing physico-chemical properties of the rpo-encoded peptides, provides a possible explanation for the observed CNI, but we cannot directly correlate plastid related CNI phenotypes to rpo genotypes. This suggests that more than one interaction between the nuclear genome and the plastome genes are needed to fully explain the observed patterns.
Keywords	Life Science
Subject code	580
Language	English
Publishing country	nl
Document type	Article ; Online
ZDB-ID	743412-1
ISSN	1592-1573 ; 0394-6169
ISSN (online)	1592-1573
ISSN	0394-6169
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Article ; Online: Double-Membrane Vesicles as Platforms for Viral Replication.

Wolff, Georg / Melia, Charlotte E / Snijder, Eric J / Bárcena, Montserrat

Trends in microbiology

2020 Volume 28, Issue 12, Page(s) 1022–1033

Abstract: Viruses, as obligate intracellular parasites, exploit cellular pathways and resources in a variety of fascinating ways. A striking example of this is the remodelling of intracellular membranes into specialized structures that support the replication of ... ...

Abstract	Viruses, as obligate intracellular parasites, exploit cellular pathways and resources in a variety of fascinating ways. A striking example of this is the remodelling of intracellular membranes into specialized structures that support the replication of positive-sense ssRNA (+RNA) viruses infecting eukaryotes. These distinct forms of virus-induced structures include double-membrane vesicles (DMVs), found during viral infections as diverse and notorious as those of coronaviruses, enteroviruses, noroviruses, or hepatitis C virus. Our understanding of these DMVs has evolved over the past 15 years thanks to advances in imaging techniques and modern molecular biology tools. In this article, we review contemporary understanding of the biogenesis, structure, and function of virus-induced DMVs as well as the open questions posed by these intriguing structures.
MeSH term(s)	Animals ; Coronavirus/physiology ; Enterovirus/physiology ; Hepacivirus/physiology ; Hepatitis C/virology ; Host Microbial Interactions/physiology ; Humans ; Intracellular Membranes/virology ; Norovirus/physiology ; Organelle Biogenesis ; RNA, Viral ; Viral Proteins ; Virus Replication/physiology
Chemical Substances	RNA, Viral ; Viral Proteins
Keywords	covid19
Language	English
Publishing date	2020-06-11
Publishing country	England
Document type	Journal Article ; Review
ZDB-ID	1158963-2
ISSN	1878-4380 ; 0966-842X
ISSN (online)	1878-4380
ISSN	0966-842X
DOI	10.1016/j.tim.2020.05.009
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Article ; Online: Pulmonary Vascular Complications in Hereditary Hemorrhagic Telangiectasia and the Underlying Pathophysiology.

Bofarid, Sala / Hosman, Anna E / Mager, Johannes J / Snijder, Repke J / Post, Marco C

International journal of molecular sciences

2021 Volume 22, Issue 7

Abstract: In this review, we discuss the role of transforming growth factor-beta (TGF-β) in the development of pulmonary vascular disease (PVD), both pulmonary arteriovenous malformations (AVM) and pulmonary hypertension (PH), in hereditary hemorrhagic ... ...

Abstract	In this review, we discuss the role of transforming growth factor-beta (TGF-β) in the development of pulmonary vascular disease (PVD), both pulmonary arteriovenous malformations (AVM) and pulmonary hypertension (PH), in hereditary hemorrhagic telangiectasia (HHT). HHT or Rendu-Osler-Weber disease is an autosomal dominant genetic disorder with an estimated prevalence of 1 in 5000 persons and characterized by epistaxis, telangiectasia and AVMs in more than 80% of cases, HHT is caused by a mutation in the ENG gene on chromosome 9 encoding for the protein endoglin or activin receptor-like kinase 1 (ACVRL1) gene on chromosome 12 encoding for the protein ALK-1, resulting in HHT type 1 or HHT type 2, respectively. A third disease-causing mutation has been found in the SMAD-4 gene, causing a combination of HHT and juvenile polyposis coli. All three genes play a role in the TGF-β signaling pathway that is essential in angiogenesis where it plays a pivotal role in neoangiogenesis, vessel maturation and stabilization. PH is characterized by elevated mean pulmonary arterial pressure caused by a variety of different underlying pathologies. HHT carries an additional increased risk of PH because of high cardiac output as a result of anemia and shunting through hepatic AVMs, or development of pulmonary arterial hypertension due to interference of the TGF-β pathway. HHT in combination with PH is associated with a worse prognosis due to right-sided cardiac failure. The treatment of PVD in HHT includes medical or interventional therapy.
MeSH term(s)	Activin Receptors, Type II/metabolism ; Animals ; Arteriovenous Malformations/complications ; Arteriovenous Malformations/genetics ; Endoglin/metabolism ; Humans ; Hypertension, Pulmonary/complications ; Hypertension, Pulmonary/genetics ; Lung Diseases/complications ; Lung Diseases/genetics ; Mutation ; Risk ; Signal Transduction ; Telangiectasia, Hereditary Hemorrhagic/complications ; Telangiectasia, Hereditary Hemorrhagic/genetics ; Transforming Growth Factor beta/metabolism ; Vascular Diseases/complications ; Vascular Diseases/genetics
Chemical Substances	ENG protein, human ; Endoglin ; Transforming Growth Factor beta ; ACVRL1 protein, human (EC 2.7.11.30) ; Activin Receptors, Type II (EC 2.7.11.30)
Language	English
Publishing date	2021-03-27
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/ijms22073471
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Article ; Online: The alphavirus nonstructural protein 2 NTPase induces a host translational shut-off through phosphorylation of eEF2 via cAMP-PKA-eEF2K signaling.

Treffers, Emmely E / Tas, Ali / Scholte, Florine E M / de Ru, Arnoud H / Snijder, Eric J / van Veelen, Peter A / van Hemert, Martijn J

PLoS pathogens

2023 Volume 19, Issue 2, Page(s) e1011179

Abstract: Chikungunya virus (CHIKV) is a reemerging alphavirus. Since 2005, it has infected millions of people during outbreaks in Africa, Asia, and South/Central America. CHIKV replication depends on host cell factors at many levels and is expected to have a ... ...

Abstract	Chikungunya virus (CHIKV) is a reemerging alphavirus. Since 2005, it has infected millions of people during outbreaks in Africa, Asia, and South/Central America. CHIKV replication depends on host cell factors at many levels and is expected to have a profound effect on cellular physiology. To obtain more insight into host responses to infection, stable isotope labeling with amino acids in cell culture and liquid chromatography-tandem mass spectrometry were used to assess temporal changes in the cellular phosphoproteome during CHIKV infection. Among the ~3,000 unique phosphorylation sites analyzed, the largest change in phosphorylation status was measured on residue T56 of eukaryotic elongation factor 2 (eEF2), which showed a >50-fold increase at 8 and 12 h p.i. Infection with other alphaviruses (Semliki Forest, Sindbis and Venezuelan equine encephalitis virus (VEEV)) triggered a similarly strong eEF2 phosphorylation. Expression of a truncated form of CHIKV or VEEV nsP2, containing only the N-terminal and NTPase/helicase domains (nsP2-NTD-Hel), sufficed to induce eEF2 phosphorylation, which could be prevented by mutating key residues in the Walker A and B motifs of the NTPase domain. Alphavirus infection or expression of nsP2-NTD-Hel resulted in decreased cellular ATP levels and increased cAMP levels. This did not occur when catalytically inactive NTPase mutants were expressed. The wild-type nsP2-NTD-Hel inhibited cellular translation independent of the C-terminal nsP2 domain, which was previously implicated in directing the virus-induced host shut-off for Old World alphaviruses. We hypothesize that the alphavirus NTPase activates a cellular adenylyl cyclase resulting in increased cAMP levels, thus activating PKA and subsequently eukaryotic elongation factor 2 kinase. This in turn triggers eEF2 phosphorylation and translational inhibition. We conclude that the nsP2-driven increase of cAMP levels contributes to the alphavirus-induced shut-off of cellular protein synthesis that is shared between Old and New World alphaviruses. MS Data are available via ProteomeXchange with identifier PXD009381.
MeSH term(s)	Humans ; Alphavirus/metabolism ; Nucleoside-Triphosphatase/metabolism ; Peptide Elongation Factor 2/metabolism ; Eukaryota ; Phosphorylation ; Chikungunya virus/physiology ; Chikungunya Fever ; Viral Nonstructural Proteins/metabolism ; Virus Replication ; Elongation Factor 2 Kinase/metabolism
Chemical Substances	Nucleoside-Triphosphatase (EC 3.6.1.15) ; Peptide Elongation Factor 2 ; Viral Nonstructural Proteins ; EEF2K protein, human (EC 2.7.1.17) ; Elongation Factor 2 Kinase (EC 2.7.11.20)
Language	English
Publishing date	2023-02-27
Publishing country	United States
Document type	Journal Article
ZDB-ID	2205412-1
ISSN	1553-7374 ; 1553-7374
ISSN (online)	1553-7374
ISSN	1553-7374
DOI	10.1371/journal.ppat.1011179
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Article ; Online: R-Propranolol Has Broad-Spectrum Anti-Coronavirus Activity and Suppresses Factors Involved in Pathogenic Angiogenesis.

Thaler, Melissa / Salgado-Benvindo, Clarisse / Leijs, Anouk / Tas, Ali / Ninaber, Dennis K / Arbiser, Jack L / Snijder, Eric J / van Hemert, Martijn J

International journal of molecular sciences

2023 Volume 24, Issue 5

Abstract: ... Therefore, therapy should not only aim to inhibit the virus, but also to suppress pathogenic host responses, e.g ...

Abstract	The SARS-CoV-2 pandemic highlighted the need for broad-spectrum antivirals to increase our preparedness. Patients often require treatment by the time that blocking virus replication is less effective. Therefore, therapy should not only aim to inhibit the virus, but also to suppress pathogenic host responses, e.g., leading to microvascular changes and pulmonary damage. Clinical studies have previously linked SARS-CoV-2 infection to pathogenic intussusceptive angiogenesis in the lungs, involving the upregulation of angiogenic factors such as ANGPTL4. The β-blocker propranolol is used to suppress aberrant ANGPTL4 expression in the treatment of hemangiomas. Therefore, we investigated the effect of propranolol on SARS-CoV-2 infection and the expression of ANGPTL4. SARS-CoV-2 upregulated ANGPTL4 in endothelial and other cells, which could be suppressed with R-propranolol. The compound also inhibited the replication of SARS-CoV-2 in Vero-E6 cells and reduced the viral load by up to ~2 logs in various cell lines and primary human airway epithelial cultures. R-propranolol was as effective as S-propranolol but lacks the latter's undesired β-blocker activity. R-propranolol also inhibited SARS-CoV and MERS-CoV. It inhibited a post-entry step of the replication cycle, likely via host factors. The broad-spectrum antiviral effect and suppression of factors involved in pathogenic angiogenesis make R-propranolol an interesting molecule to further explore for the treatment of coronavirus infections.
MeSH term(s)	Animals ; Chlorocebus aethiops ; Humans ; COVID-19 ; Propranolol/pharmacology ; SARS-CoV-2 ; Vero Cells ; Cell Line ; Antiviral Agents/pharmacology ; Virus Replication
Chemical Substances	Propranolol (9Y8NXQ24VQ) ; Antiviral Agents
Language	English
Publishing date	2023-02-27
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/ijms24054588
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Article ; Online: The Nonstructural Proteins Directing Coronavirus RNA Synthesis and Processing.

Snijder, E J / Decroly, E / Ziebuhr, J

Advances in virus research

2016 Volume 96, Page(s) 59–126

Abstract: Coronaviruses are animal and human pathogens that can cause lethal zoonotic infections like SARS and MERS. They have polycistronic plus-stranded RNA genomes and belong to the order Nidovirales, a diverse group of viruses for which common ancestry was ... ...

Abstract	Coronaviruses are animal and human pathogens that can cause lethal zoonotic infections like SARS and MERS. They have polycistronic plus-stranded RNA genomes and belong to the order Nidovirales, a diverse group of viruses for which common ancestry was inferred from the common principles underlying their genome organization and expression, and from the conservation of an array of core replicase domains, including key RNA-synthesizing enzymes. Coronavirus genomes (~26-32 kilobases) are the largest RNA genomes known to date and their expansion was likely enabled by acquiring enzyme functions that counter the commonly high error frequency of viral RNA polymerases. The primary functions that direct coronavirus RNA synthesis and processing reside in nonstructural protein (nsp) 7 to nsp16, which are cleavage products of two large replicase polyproteins translated from the coronavirus genome. Significant progress has now been made regarding their structural and functional characterization, stimulated by technical advances like improved methods for bioinformatics and structural biology, in vitro enzyme characterization, and site-directed mutagenesis of coronavirus genomes. Coronavirus replicase functions include more or less universal activities of plus-stranded RNA viruses, like an RNA polymerase (nsp12) and helicase (nsp13), but also a number of rare or even unique domains involved in mRNA capping (nsp14, nsp16) and fidelity control (nsp14). Several smaller subunits (nsp7-nsp10) act as crucial cofactors of these enzymes and contribute to the emerging "nsp interactome." Understanding the structure, function, and interactions of the RNA-synthesizing machinery of coronaviruses will be key to rationalizing their evolutionary success and the development of improved control strategies.
MeSH term(s)	Acid Anhydride Hydrolases/genetics ; Acid Anhydride Hydrolases/metabolism ; Animals ; Gene Expression Regulation, Viral ; Humans ; Methyltransferases/genetics ; Methyltransferases/metabolism ; Protein Domains ; RNA Helicases/genetics ; RNA Helicases/metabolism ; RNA, Viral/biosynthesis ; RNA, Viral/genetics ; SARS Virus/genetics ; SARS Virus/metabolism ; Severe Acute Respiratory Syndrome/pathology ; Severe Acute Respiratory Syndrome/virology ; Viral Nonstructural Proteins/genetics ; Viral Nonstructural Proteins/metabolism ; Virus Replication
Chemical Substances	RNA, Viral ; Viral Nonstructural Proteins ; Methyltransferases (EC 2.1.1.-) ; Acid Anhydride Hydrolases (EC 3.6.-) ; RNA triphosphatase (EC 3.6.1.-) ; RNA Helicases (EC 3.6.4.13)
Keywords	covid19
Language	English
Publishing date	2016-09-14
Publishing country	United States
Document type	Journal Article ; Review
ZDB-ID	195-8
ISSN	1557-8399 ; 0065-3527
ISSN (online)	1557-8399
ISSN	0065-3527
DOI	10.1016/bs.aivir.2016.08.008
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Article ; Online: The contribution of obesity to the population burden of high metabolic cardiovascular risk among different ethnic groups. The HELIUS study.

Perini, Wilco / van Valkengoed, Irene G M / Snijder, Marieke B / Peters, Ron J G / Kunst, Anton E

European journal of public health

2020 Volume 30, Issue 2, Page(s) 322–327

Abstract: Background: The burden of cardiovascular risk is distributed unequally between ethnic groups. It is uncertain to what extent this is attributable to ethnic differences in general and abdominal obesity. Therefore, we studied the contribution of general ... ...

Abstract	Background: The burden of cardiovascular risk is distributed unequally between ethnic groups. It is uncertain to what extent this is attributable to ethnic differences in general and abdominal obesity. Therefore, we studied the contribution of general and abdominal obesity to metabolic cardiovascular risk among different ethnic groups. Methods: We used data of 21 411 participants of Dutch, South-Asian Surinamese, African-Surinamese, Ghanaian, Turkish or Moroccan origin in Healthy Life in an Urban Setting (Amsterdam, the Netherlands). Obesity was defined using body-mass-index (general) or waist-to-height-ratio (abdominal). High metabolic risk was defined as having at least two of the following: triglycerides ≥1.7 mmol/l, fasting glucose ≥5.6 mmol/l, blood pressure ≥130 mmHg systolic and/or ≥85 mmHg diastolic and high-density lipoprotein cholesterol <1.03 mmol/l (men) or <1.29 mmol/l (women). Results: Among ethnic minority men, age-adjusted prevalence rates of high metabolic risk ranged from 32 to 59% vs. 33% among Dutch men. Contributions of general obesity to high metabolic risk ranged from 7.1 to 17.8%, vs. 10.1% among Dutch men, whereas contributions of abdominal obesity ranged from 52.1 to 92.3%, vs. 53.9% among Dutch men. Among ethnic minority women, age-adjusted prevalence rates of high metabolic risk ranged from 24 to 35% vs. 12% among Dutch women. Contributions of general obesity ranged from 14.6 to 41.8%, vs. 20% among Dutch women, whereas contributions of abdominal obesity ranged from 68.0 to 92.8%, vs. 72.1% among Dutch women. Conclusions: Obesity, especially abdominal obesity, contributes significantly to the prevalence of high metabolic cardiovascular risk. Results suggest that this contribution varies substantially between ethnic groups, which helps explain ethnic differences in cardiovascular risk.
MeSH term(s)	Cardiovascular Diseases/epidemiology ; Cross-Sectional Studies ; Ethnic Groups ; Female ; Ghana ; Heart Disease Risk Factors ; Humans ; Male ; Minority Groups ; Netherlands/epidemiology ; Obesity/epidemiology ; Prevalence ; Risk Factors
Language	English
Publishing date	2020-03-09
Publishing country	England
Document type	Journal Article ; Research Support, Non-U.S. Gov't
ZDB-ID	1129243-x
ISSN	1464-360X ; 1101-1262
ISSN (online)	1464-360X
ISSN	1101-1262
DOI	10.1093/eurpub/ckz190
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Zs.A 3499: Show issues

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Article ; Online: Pulmonary Vascular Complications in Hereditary Hemorrhagic Telangiectasia and the Underlying Pathophysiology

Sala Bofarid / Anna E. Hosman / J.J. Mager / R.J. Snijder / Marco C. Post

International Journal of Molecular Sciences, Vol 22, Iss 3471, p

2021 Volume 3471

Abstract	In this review, we discuss the role of transforming growth factor-beta (TGF-β) in the development of pulmonary vascular disease (PVD), both pulmonary arteriovenous malformations (AVM) and pulmonary hypertension (PH), in hereditary hemorrhagic telangiectasia (HHT). HHT or Rendu-Osler-Weber disease is an autosomal dominant genetic disorder with an estimated prevalence of 1 in 5000 persons and characterized by epistaxis, telangiectasia and AVMs in more than 80% of cases, HHT is caused by a mutation in the ENG gene on chromosome 9 encoding for the protein endoglin or activin receptor-like kinase 1 (ACVRL1) gene on chromosome 12 encoding for the protein ALK-1, resulting in HHT type 1 or HHT type 2, respectively. A third disease-causing mutation has been found in the SMAD-4 gene, causing a combination of HHT and juvenile polyposis coli. All three genes play a role in the TGF-β signaling pathway that is essential in angiogenesis where it plays a pivotal role in neoangiogenesis, vessel maturation and stabilization. PH is characterized by elevated mean pulmonary arterial pressure caused by a variety of different underlying pathologies. HHT carries an additional increased risk of PH because of high cardiac output as a result of anemia and shunting through hepatic AVMs, or development of pulmonary arterial hypertension due to interference of the TGF-β pathway. HHT in combination with PH is associated with a worse prognosis due to right-sided cardiac failure. The treatment of PVD in HHT includes medical or interventional therapy.
Keywords	HHT ; endoglin ; Pulmonary Vascular Disease ; Biology (General) ; QH301-705.5 ; Chemistry ; QD1-999
Subject code	610
Language	English
Publishing date	2021-03-01T00:00:00Z
Publisher	MDPI AG
Document type	Article ; Online
Database	BASE - Bielefeld Academic Search Engine (life sciences selection)

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Article ; Online: Spatially Addressable Multiplex Biodetection by Calibrated Micro/Nanostructured Surfaces.

Dobroiu, Serban / van Delft, Falco C M J M / Sudalaiyadum Perumal, Ayyappasamy / Dash, Shantoshini / Aveyard, Jenny / van Zijl, Jeroen / Snijder, Jaap / van den Heuvel, Eric / van Berkum, Jurgen / Blanchard, Marie Pierre / Favard, Cyril / Nicolau, Dan V

ACS sensors

2023 Volume 8, Issue 5, Page(s) 1882–1890

Abstract: ... FLIC extreme sensitivity to wavelength is also its main problem: small, e.g., 10 nm range, variations ... technology was further validated on a diagnostically important target, i.e., the receptor-binding domain (RBD ...

Abstract	A challenge of any biosensing technology is the detection of very low concentrations of analytes. The fluorescence interference contrast (FLIC) technique improves the fluorescence-based sensitivity by selectively amplifying, or suppressing, the emission of a fluorophore-labeled biomolecule immobilized on a transparent layer placed on top of a mirror basal surface. The standing wave of the reflected emission light means that the height of the transparent layer operates as a surface-embedded optical filter for the fluorescence signal. FLIC extreme sensitivity to wavelength is also its main problem: small, e.g., 10 nm range, variations of the vertical position of the fluorophore can translate in unwanted suppression of the detection signal. Herein, we introduce the concept of quasi-circular lenticular microstructured domes operating as continuous-mode optical filters, generating fluorescent concentric rings, with diameters determined by the wavelengths of the fluorescence light, in turn modulated by FLIC. The critical component of the lenticular structures was the shallow sloping side wall, which allowed the simultaneous separation of fluorescent patterns for virtually any fluorophore wavelength. Purposefully designed microstructures with either stepwise or continuous-slope dome geometries were fabricated to modulate the intensity and the lateral position of a fluorescence signal. The simulation of FLIC effects induced by the lenticular microstructures was confirmed by the measurement of the fluorescence profile for three fluorescent dyes, as well as high-resolution fluorescence scanning using stimulated emission depletion (STED) microscopy. The high sensitivity of the spatially addressable FLIC technology was further validated on a diagnostically important target, i.e., the receptor-binding domain (RBD) of the SARS-Cov2 via the detection of RBD:anti-S1-antibody.
MeSH term(s)	Humans ; Microscopy, Fluorescence/methods ; RNA, Viral ; COVID-19 ; SARS-CoV-2 ; Fluorescent Dyes/chemistry
Chemical Substances	RNA, Viral ; Fluorescent Dyes
Language	English
Publishing date	2023-04-26
Publishing country	United States
Document type	Journal Article ; Research Support, Non-U.S. Gov't
ISSN	2379-3694
ISSN (online)	2379-3694
DOI	10.1021/acssensors.2c01939
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Article ; Online: A swine arterivirus deubiquitinase stabilizes two major envelope proteins and promotes production of viral progeny.

Guo, Rui / Yan, Xingyu / Li, Yanhua / Cui, Jin / Misra, Saurav / Firth, Andrew E / Snijder, Eric J / Fang, Ying

PLoS pathogens

2021 Volume 17, Issue 3, Page(s) e1009403

Abstract: Arteriviruses are enveloped positive-strand RNA viruses that assemble and egress using the host cell's exocytic pathway. In previous studies, we demonstrated that most arteriviruses use a unique -2 ribosomal frameshifting mechanism to produce a C- ... ...

Abstract	Arteriviruses are enveloped positive-strand RNA viruses that assemble and egress using the host cell's exocytic pathway. In previous studies, we demonstrated that most arteriviruses use a unique -2 ribosomal frameshifting mechanism to produce a C-terminally modified variant of their nonstructural protein 2 (nsp2). Like full-length nsp2, the N-terminal domain of this frameshift product, nsp2TF, contains a papain-like protease (PLP2) that has deubiquitinating (DUB) activity, in addition to its role in proteolytic processing of replicase polyproteins. In cells infected with porcine reproductive and respiratory syndrome virus (PRRSV), nsp2TF localizes to compartments of the exocytic pathway, specifically endoplasmic reticulum-Golgi intermediate compartment (ERGIC) and Golgi complex. Here, we show that nsp2TF interacts with the two major viral envelope proteins, the GP5 glycoprotein and membrane (M) protein, which drive the key process of arterivirus assembly and budding. The PRRSV GP5 and M proteins were found to be poly-ubiquitinated, both in an expression system and in cells infected with an nsp2TF-deficient mutant virus. In contrast, ubiquitinated GP5 and M proteins did not accumulate in cells infected with the wild-type, nsp2TF-expressing virus. Further analysis implicated the DUB activity of the nsp2TF PLP2 domain in deconjugation of ubiquitin from GP5/M proteins, thus antagonizing proteasomal degradation of these key viral structural proteins. Our findings suggest that nsp2TF is targeted to the exocytic pathway to reduce proteasome-driven turnover of GP5/M proteins, thus promoting the formation of GP5-M dimers that are critical for arterivirus assembly.
MeSH term(s)	Animals ; Cell Line ; Deubiquitinating Enzymes/metabolism ; Gene Expression Regulation, Viral/physiology ; Humans ; Porcine Reproductive and Respiratory Syndrome/virology ; Porcine respiratory and reproductive syndrome virus/metabolism ; Swine ; Viral Envelope Proteins/metabolism ; Viral Proteins/metabolism ; Virus Assembly/physiology ; Virus Replication/physiology
Chemical Substances	Viral Envelope Proteins ; Viral Proteins ; Deubiquitinating Enzymes (EC 3.4.19.12)
Language	English
Publishing date	2021-03-18
Publishing country	United States
Document type	Journal Article ; Research Support, U.S. Gov't, Non-P.H.S.
ZDB-ID	2205412-1
ISSN	1553-7374 ; 1553-7366
ISSN (online)	1553-7374
ISSN	1553-7366
DOI	10.1371/journal.ppat.1009403
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