20 results on '"Stoy H"'
Search Results
2. Feiba And Autoplex: A Comparison Of These Two Activated Prothrombin Complex Concentrates
- Author
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Lechler, E, Eggeling, B, Meyer-Börnecke, D, and Stoy, H
- Published
- 1981
- Full Text
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3. Nuclear actin polymerization rapidly mediates replication fork remodeling upon stress by limiting PrimPol activity.
- Author
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Palumbieri MD, Merigliano C, González-Acosta D, Kuster D, Krietsch J, Stoy H, von Känel T, Ulferts S, Welter B, Frey J, Doerdelmann C, Sanchi A, Grosse R, Chiolo I, and Lopes M
- Subjects
- Polymerization, Cell Line, Tumor, DNA genetics, Actins genetics, DNA Replication
- Abstract
Cells rapidly respond to replication stress actively slowing fork progression and inducing fork reversal. How replication fork plasticity is achieved in the context of nuclear organization is currently unknown. Using nuclear actin probes in living and fixed cells, we visualized nuclear actin filaments in unperturbed S phase and observed their rapid extension in number and length upon genotoxic treatments, frequently taking contact with replication factories. Chemically or genetically impairing nuclear actin polymerization shortly before these treatments prevents active fork slowing and abolishes fork reversal. Defective fork remodeling is linked to deregulated chromatin loading of PrimPol, which promotes unrestrained and discontinuous DNA synthesis and limits the recruitment of RAD51 and SMARCAL1 to nascent DNA. Moreover, defective nuclear actin polymerization upon mild replication interference induces chromosomal instability in a PRIMPOL-dependent manner. Hence, by limiting PrimPol activity, nuclear F-actin orchestrates replication fork plasticity and is a key molecular determinant in the rapid cellular response to genotoxic treatments., (© 2023. The Author(s).)
- Published
- 2023
- Full Text
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4. Excessive reactive oxygen species induce transcription-dependent replication stress.
- Author
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Andrs M, Stoy H, Boleslavska B, Chappidi N, Kanagaraj R, Nascakova Z, Menon S, Rao S, Oravetzova A, Dobrovolna J, Surendranath K, Lopes M, and Janscak P
- Subjects
- Humans, Reactive Oxygen Species, S Phase genetics, Hydroxyurea pharmacology, DNA, DNA Replication, DNA-Binding Proteins metabolism
- Abstract
Elevated levels of reactive oxygen species (ROS) reduce replication fork velocity by causing dissociation of the TIMELESS-TIPIN complex from the replisome. Here, we show that ROS generated by exposure of human cells to the ribonucleotide reductase inhibitor hydroxyurea (HU) promote replication fork reversal in a manner dependent on active transcription and formation of co-transcriptional RNA:DNA hybrids (R-loops). The frequency of R-loop-dependent fork stalling events is also increased after TIMELESS depletion or a partial inhibition of replicative DNA polymerases by aphidicolin, suggesting that this phenomenon is due to a global replication slowdown. In contrast, replication arrest caused by HU-induced depletion of deoxynucleotides does not induce fork reversal but, if allowed to persist, leads to extensive R-loop-independent DNA breakage during S-phase. Our work reveals a link between oxidative stress and transcription-replication interference that causes genomic alterations recurrently found in human cancer., (© 2023. The Author(s).)
- Published
- 2023
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5. Replication fork plasticity upon replication stress requires rapid nuclear actin polymerization.
- Author
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Palumbieri MD, Merigliano C, Acosta DG, von Känel T, Welter B, Stoy H, Krietsch J, Ulferts S, Sanchi A, Grosse R, Chiolo I, and Lopes M
- Abstract
Cells rapidly respond to replication stress actively slowing fork progression and inducing fork reversal. How replication fork plasticity is achieved in the context of nuclear organization is currently unknown. Using nuclear actin probes in living and fixed cells, we visualized nuclear actin filaments in unperturbed S phase, rapidly extending in number and thickness upon genotoxic treatments, and taking frequent contact with replication factories. Chemically or genetically impairing nuclear actin polymerization shortly before these treatments prevents active fork slowing and abolishes fork reversal. Defective fork plasticity is linked to reduced recruitment of RAD51 and SMARCAL1 to nascent DNA. Conversely, PRIMPOL gains access to replicating chromatin, promoting unrestrained and discontinuous DNA synthesis, which is associated with increased chromosomal instability and decreased cellular resistance to replication stress. Hence, nuclear F-actin orchestrates replication fork plasticity and is a key molecular determinant in the rapid cellular response to genotoxic treatments.
- Published
- 2023
- Full Text
- View/download PDF
6. Direct visualization of transcription-replication conflicts reveals post-replicative DNA:RNA hybrids.
- Author
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Stoy H, Zwicky K, Kuster D, Lang KS, Krietsch J, Crossley MP, Schmid JA, Cimprich KA, Merrikh H, and Lopes M
- Subjects
- Humans, DNA chemistry, DNA-Binding Proteins metabolism, Chromosomes metabolism, Genomic Instability, RNA, DNA Replication
- Abstract
Transcription-replication collisions (TRCs) are crucial determinants of genome instability. R-loops were linked to head-on TRCs and proposed to obstruct replication fork progression. The underlying mechanisms, however, remained elusive due to the lack of direct visualization and of non-ambiguous research tools. Here, we ascertained the stability of estrogen-induced R-loops on the human genome, visualized them directly by electron microscopy (EM), and measured R-loop frequency and size at the single-molecule level. Combining EM and immuno-labeling on locus-specific head-on TRCs in bacteria, we observed the frequent accumulation of DNA:RNA hybrids behind replication forks. These post-replicative structures are linked to fork slowing and reversal across conflict regions and are distinct from physiological DNA:RNA hybrids at Okazaki fragments. Comet assays on nascent DNA revealed a marked delay in nascent DNA maturation in multiple conditions previously linked to R-loop accumulation. Altogether, our findings suggest that TRC-associated replication interference entails transactions that follow initial R-loop bypass by the replication fork., (© 2023. The Author(s).)
- Published
- 2023
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7. Short Arrestin-3-Derived Peptides Activate JNK3 in Cells.
- Author
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Perry-Hauser NA, Kaoud TS, Stoy H, Zhan X, Chen Q, Dalby KN, Iverson TM, Gurevich VV, and Gurevich EV
- Subjects
- Arrestins metabolism, Peptides metabolism, Peptides pharmacology, Phosphorylation physiology, Protein Binding physiology, beta-Arrestin 2 metabolism, beta-Arrestins metabolism, Arrestin metabolism, Mitogen-Activated Protein Kinase 10 metabolism
- Abstract
Arrestins were first discovered as suppressors of G protein-mediated signaling by G protein-coupled receptors. It was later demonstrated that arrestins also initiate several signaling branches, including mitogen-activated protein kinase cascades. Arrestin-3-dependent activation of the JNK family can be recapitulated with peptide fragments, which are monofunctional elements distilled from this multi-functional arrestin protein. Here, we use maltose-binding protein fusions of arrestin-3-derived peptides to identify arrestin elements that bind kinases of the ASK1-MKK4/7-JNK3 cascade and the shortest peptide facilitating JNK signaling. We identified a 16-residue arrestin-3-derived peptide expressed as a Venus fusion that leads to activation of JNK3α2 in cells. The strength of the binding to the kinases does not correlate with peptide activity. The ASK1-MKK4/7-JNK3 cascade has been implicated in neuronal apoptosis. While inhibitors of MAP kinases exist, short peptides are the first small molecule tools that can activate MAP kinases.
- Published
- 2022
- Full Text
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8. Identification of a novel compound that simultaneously impairs the ubiquitin-proteasome system and autophagy.
- Author
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Giovannucci TA, Salomons FA, Stoy H, Herzog LK, Xu S, Qian W, Merino LG, Gierisch ME, Haraldsson M, Lystad AH, Uvell H, Simonsen A, Gustavsson AL, Vallin M, and Dantuma NP
- Subjects
- Autophagy, Proteolysis, Proteasome Endopeptidase Complex metabolism, Ubiquitin metabolism
- Abstract
The ubiquitin-proteasome system (UPS) and macroautophagy/autophagy are the main proteolytic systems in eukaryotic cells for preserving protein homeostasis, i.e., proteostasis. By facilitating the timely destruction of aberrant proteins, these complementary pathways keep the intracellular environment free of inherently toxic protein aggregates. Chemical interference with the UPS or autophagy has emerged as a viable strategy for therapeutically targeting malignant cells which, owing to their hyperactive state, heavily rely on the sanitizing activity of these proteolytic systems. Here, we report on the discovery of CBK79, a novel compound that impairs both protein degradation by the UPS and autophagy. While CBK79 was identified in a high-content screen for drug-like molecules that inhibit the UPS, subsequent analysis revealed that this compound also compromises autophagic degradation of long-lived proteins. We show that CBK79 induces non-canonical lipidation of MAP1LC3B/LC3B (microtubule-associated protein 1 light chain 3 beta) that requires ATG16L1 but is independent of the ULK1 (unc-51 like autophagy activating kinase 1) and class III phosphatidylinositol 3-kinase (PtdIns3K) complexes. Thermal preconditioning of cells prevented CBK79-induced UPS impairment but failed to restore autophagy, indicating that activation of stress responses does not allow cells to bypass the inhibitory effect of CBK79 on autophagy. The identification of a small molecule that simultaneously impairs the two main proteolytic systems for protein quality control provides a starting point for the development of a novel class of proteostasis-targeting drugs.
- Published
- 2022
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9. Locus-Specific Analysis of Replication Dynamics and Detection of DNA-RNA Hybrids by Immuno Electron Microscopy.
- Author
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Stoy H, S Lang K, Merrikh H, and Lopes M
- Subjects
- DNA genetics, Microscopy, Electron, Microscopy, Immunoelectron, DNA Replication, RNA genetics
- Abstract
DNA-RNA hybrids can interfere with DNA replication, but the underlying intermediates and molecular mechanisms have remained elusive. Here, we describe a single molecule approach that allows to monitor DNA-RNA hybrids locus-specifically in the context of ongoing replication. Using restriction digestion, gel electrophoresis and gel elution, this workflow allows to efficiently isolate replication intermediates and to study replication dynamics across a specific genomic locus. Here, we applied this procedure to isolate a bacterial genomic locus carrying an inducible transcription-replication conflict. Moreover, we combined electron microscopy with S9.6-Gold immuno-labeling to detect DNA-RNA hybrids on the isolated replication intermediates. With some limitations, this approach may be adapted to locus-specific replication analyses in different organisms., (© 2022. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.)
- Published
- 2022
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10. Direct R-Loop Visualization on Genomic DNA by Native Automated Electron Microscopy.
- Author
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Stoy H, Luethi J, Roessler FK, Riemann J, Kaech A, and Lopes M
- Subjects
- DNA chemistry, DNA genetics, DNA Damage, DNA Replication, Genomics, Humans, Microscopy, Electron, RNA genetics, Genomic Instability, R-Loop Structures
- Abstract
R-loop are physiologically present on genomic DNA of different organisms and play important roles in genome regulation. However, an increase in their abundance and/or size has been suggested to interfere with the DNA replication process, contributing to genome instability. Most available approaches to monitor R-loops are based on antibodies/enzymes that cannot effectively distinguish R-loops from DNA-RNA hybrids and assess R-loop size and frequency in a population of molecules. Electron microscopy has successfully allowed single-molecule visualization of DNA replication and repair intermediates, uncovering key architectural modifications in DNA, induced by genotoxic stress or by the associated cellular response. Here, we describe recent modifications of this visualization workflow to implement partial automation of image acquisition and analysis. Coupling this refined workflow with sample preparation procedures that protect R-loop stability allows for direct visualization of R-loop structures on genomic DNA, independently from probes. Combining single-molecule information and DNA content assessment, this approach provides direct estimations of R-loop frequency, size, and burden on genomic DNA., (© 2022. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.)
- Published
- 2022
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11. Poly(ADP-ribosyl)ation temporally confines SUMO-dependent ataxin-3 recruitment to control DNA double-strand break repair.
- Author
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Pfeiffer A, Herzog LK, Luijsterburg MS, Shah RG, Rother MB, Stoy H, Kühbacher U, van Attikum H, Shah GM, and Dantuma NP
- Subjects
- Cell Line, Tumor, DNA, DNA Damage, DNA Repair genetics, Humans, Poly (ADP-Ribose) Polymerase-1 genetics, Ataxin-3 genetics, DNA Breaks, Double-Stranded, Poly ADP Ribosylation
- Abstract
DNA damage-induced SUMOylation serves as a signal for two antagonizing proteins that both stimulate repair of DNA double-strand breaks (DSBs). Here, we demonstrate that the SUMO-dependent recruitment of the deubiquitylating enzyme ataxin-3 to DSBs, unlike recruitment of the ubiquitin ligase RNF4, additionally depends on poly [ADP-ribose] polymerase 1 (PARP1)-mediated poly(ADP-ribosyl)ation (PARylation). The co-dependence of ataxin-3 recruitment on PARylation and SUMOylation temporally confines ataxin-3 to DSBs immediately after occurrence of DNA damage. We propose that this mechanism ensures that ataxin-3 prevents the premature removal of DNA repair proteins only during the early phase of the DSB response and does not interfere with the subsequent timely displacement of DNA repair proteins by RNF4. Thus, our data show that PARylation differentially regulates SUMO-dependent recruitment of ataxin-3 and RNF4 to DSBs, explaining how both proteins can play a stimulatory role at DSBs despite their opposing activities., Competing Interests: Competing interestsThe authors declare no competing or financial interests., (© 2021. Published by The Company of Biologists Ltd.)
- Published
- 2021
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12. Cyclin A2 localises in the cytoplasm at the S/G2 transition to activate PLK1.
- Author
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Silva Cascales H, Burdova K, Middleton A, Kuzin V, Müllers E, Stoy H, Baranello L, Macurek L, and Lindqvist A
- Subjects
- CDC2 Protein Kinase deficiency, CDC2 Protein Kinase genetics, Cell Nucleus metabolism, Chromatin metabolism, Cyclin A2 genetics, Cyclin-Dependent Kinase 2 deficiency, Cyclin-Dependent Kinase 2 genetics, DNA Damage genetics, Enzyme Activation genetics, HeLa Cells, Humans, Mitosis genetics, Phosphorylation genetics, Protein Binding, Transfection, Polo-Like Kinase 1, Cell Cycle Proteins metabolism, Cyclin A2 metabolism, Cytoplasm metabolism, G2 Phase genetics, Protein Serine-Threonine Kinases metabolism, Proto-Oncogene Proteins metabolism, S Phase genetics, Signal Transduction genetics
- Abstract
Cyclin A2 is a key regulator of the cell cycle, implicated both in DNA replication and mitotic entry. Cyclin A2 participates in feedback loops that activate mitotic kinases in G2 phase, but why active Cyclin A2-CDK2 during the S phase does not trigger mitotic kinase activation remains unclear. Here, we describe a change in localisation of Cyclin A2 from being only nuclear to both nuclear and cytoplasmic at the S/G2 border. We find that Cyclin A2-CDK2 can activate the mitotic kinase PLK1 through phosphorylation of Bora, and that only cytoplasmic Cyclin A2 interacts with Bora and PLK1. Expression of predominately cytoplasmic Cyclin A2 or phospho-mimicking PLK1 T210D can partially rescue a G2 arrest caused by Cyclin A2 depletion. Cytoplasmic presence of Cyclin A2 is restricted by p21, in particular after DNA damage. Cyclin A2 chromatin association during DNA replication and additional mechanisms contribute to Cyclin A2 localisation change in the G2 phase. We find no evidence that such mechanisms involve G2 feedback loops and suggest that cytoplasmic appearance of Cyclin A2 at the S/G2 transition functions as a trigger for mitotic kinase activation., (© 2021 Silva Cascales et al.)
- Published
- 2021
- Full Text
- View/download PDF
13. HLTF Promotes Fork Reversal, Limiting Replication Stress Resistance and Preventing Multiple Mechanisms of Unrestrained DNA Synthesis.
- Author
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Bai G, Kermi C, Stoy H, Schiltz CJ, Bacal J, Zaino AM, Hadden MK, Eichman BF, Lopes M, and Cimprich KA
- Subjects
- Cell Line, Tumor, DNA genetics, DNA Damage genetics, DNA Primase metabolism, DNA Primase physiology, DNA Repair genetics, DNA Replication genetics, DNA-Binding Proteins genetics, DNA-Directed DNA Polymerase metabolism, DNA-Directed DNA Polymerase physiology, HEK293 Cells, Humans, K562 Cells, Multifunctional Enzymes metabolism, Multifunctional Enzymes physiology, Nucleotidyltransferases metabolism, Nucleotidyltransferases physiology, Transcription Factors genetics, DNA biosynthesis, DNA Replication physiology, DNA-Binding Proteins metabolism, Transcription Factors metabolism
- Abstract
DNA replication stress can stall replication forks, leading to genome instability. DNA damage tolerance pathways assist fork progression, promoting replication fork reversal, translesion DNA synthesis (TLS), and repriming. In the absence of the fork remodeler HLTF, forks fail to slow following replication stress, but underlying mechanisms and cellular consequences remain elusive. Here, we demonstrate that HLTF-deficient cells fail to undergo fork reversal in vivo and rely on the primase-polymerase PRIMPOL for repriming, unrestrained replication, and S phase progression upon limiting nucleotide levels. By contrast, in an HLTF-HIRAN mutant, unrestrained replication relies on the TLS protein REV1. Importantly, HLTF-deficient cells also exhibit reduced double-strand break (DSB) formation and increased survival upon replication stress. Our findings suggest that HLTF promotes fork remodeling, preventing other mechanisms of replication stress tolerance in cancer cells. This remarkable plasticity of the replication fork may determine the outcome of replication stress in terms of genome integrity, tumorigenesis, and response to chemotherapy., Competing Interests: Declaration of Interests The authors declare no competing interests., (Published by Elsevier Inc.)
- Published
- 2020
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14. Histone Ubiquitination by the DNA Damage Response Is Required for Efficient DNA Replication in Unperturbed S Phase.
- Author
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Schmid JA, Berti M, Walser F, Raso MC, Schmid F, Krietsch J, Stoy H, Zwicky K, Ursich S, Freire R, Lopes M, and Penengo L
- Subjects
- Cell Line, DNA Breaks, Double-Stranded, DNA Replication physiology, DNA-Binding Proteins metabolism, Humans, S Phase physiology, Signal Transduction, Tumor Suppressor p53-Binding Protein 1 metabolism, Ubiquitin-Protein Ligases metabolism, Ubiquitination physiology, DNA Damage physiology, DNA Repair physiology, Histones metabolism
- Abstract
Chromatin ubiquitination by the ubiquitin ligase RNF168 is critical to regulate the DNA damage response (DDR). DDR deficiencies lead to cancer-prone syndromes, but whether this reflects DNA repair defects is still elusive. We identified key factors of the RNF168 pathway as essential mediators of efficient DNA replication in unperturbed S phase. We found that loss of RNF168 leads to reduced replication fork progression and to reversed fork accumulation, particularly evident at repetitive sequences stalling replication. Slow fork progression depends on MRE11-dependent degradation of reversed forks, implicating RNF168 in reversed fork protection and restart. Consistent with regular nucleosomal organization of reversed forks, the replication function of RNF168 requires H2A ubiquitination. As this novel function is shared with the key DDR players ATM, γH2A.X, RNF8, and 53BP1, we propose that double-stranded ends at reversed forks engage classical DDR factors, suggesting an alternative function of this pathway in preventing genome instability and human disease., (Copyright © 2018 Elsevier Inc. All rights reserved.)
- Published
- 2018
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15. DNA damage during S-phase mediates the proliferation-quiescence decision in the subsequent G1 via p21 expression.
- Author
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Barr AR, Cooper S, Heldt FS, Butera F, Stoy H, Mansfeld J, Novák B, and Bakal C
- Subjects
- Cell Cycle Checkpoints genetics, Cell Division genetics, Cell Line, Cell Tracking methods, Cyclin-Dependent Kinase Inhibitor p21 metabolism, Gene Knockout Techniques, Genomic Instability, Green Fluorescent Proteins genetics, Green Fluorescent Proteins metabolism, Humans, Microscopy, Confocal, Tumor Suppressor Protein p53 genetics, Tumor Suppressor Protein p53 metabolism, Cell Proliferation genetics, Cyclin-Dependent Kinase Inhibitor p21 genetics, DNA Damage, G1 Phase genetics, S Phase genetics
- Abstract
Following DNA damage caused by exogenous sources, such as ionizing radiation, the tumour suppressor p53 mediates cell cycle arrest via expression of the CDK inhibitor, p21. However, the role of p21 in maintaining genomic stability in the absence of exogenous DNA-damaging agents is unclear. Here, using live single-cell measurements of p21 protein in proliferating cultures, we show that naturally occurring DNA damage incurred over S-phase causes p53-dependent accumulation of p21 during mother G2- and daughter G1-phases. High p21 levels mediate G1 arrest via CDK inhibition, yet lower levels have no impact on G1 progression, and the ubiquitin ligases CRL4
Cdt2 and SCFSkp2 couple to degrade p21 prior to the G1/S transition. Mathematical modelling reveals that a bistable switch, created by CRL4Cdt2 , promotes irreversible S-phase entry by keeping p21 levels low, preventing premature S-phase exit upon DNA damage. Thus, we characterize how p21 regulates the proliferation-quiescence decision to maintain genomic stability.- Published
- 2017
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16. Corrigendum: Wall teichoic acids mediate increased virulence in Staphylococcus aureus.
- Author
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Wanner S, Schade J, Keinhörster D, Weller N, George SE, Kull L, Bauer J, Grau T, Winstel V, Stoy H, Kretschmer D, Kolata J, Wolz C, Bröker BM, and Weidenmaier C
- Published
- 2017
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17. Wall teichoic acids mediate increased virulence in Staphylococcus aureus.
- Author
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Wanner S, Schade J, Keinhörster D, Weller N, George SE, Kull L, Bauer J, Grau T, Winstel V, Stoy H, Kretschmer D, Kolata J, Wolz C, Bröker BM, and Weidenmaier C
- Subjects
- Animals, Anti-Bacterial Agents pharmacology, Community-Acquired Infections microbiology, Male, Methicillin-Resistant Staphylococcus aureus chemistry, Methicillin-Resistant Staphylococcus aureus drug effects, Methicillin-Resistant Staphylococcus aureus genetics, Mice, Skin microbiology, Skin pathology, Teichoic Acids analysis, Virulence, Virulence Factors biosynthesis, Abscess microbiology, Cell Wall chemistry, Methicillin-Resistant Staphylococcus aureus pathogenicity, Staphylococcal Skin Infections microbiology, Teichoic Acids biosynthesis
- Abstract
Community-associated methicillin-resistant Staphylococcus aureus (CA-MRSA) are the cause of a severe pandemic consisting primarily of skin and soft tissue infections. The underlying pathomechanisms have not been fully understood and we report here a mechanism that plays an important role for the elevated virulence of CA-MRSA. Surprisingly, skin abscess induction in an animal model was correlated with the amount of a major cell wall component of S. aureus, termed wall teichoic acid (WTA). CA-MRSA exhibited increased cell-wall-associated WTA content (WTA
high ) and thus were more active in inducing abscess formation via a WTA-dependent and T-cell-mediated mechanism than S. aureus strains with a WTAlow phenotype. We show here that WTA is directly involved in S. aureus strain-specific virulence and provide insight into the underlying molecular mechanisms that could guide the development of novel anti-infective strategies.- Published
- 2017
- Full Text
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18. Peptide mini-scaffold facilitates JNK3 activation in cells.
- Author
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Zhan X, Stoy H, Kaoud TS, Perry NA, Chen Q, Perez A, Els-Heindl S, Slagis JV, Iverson TM, Beck-Sickinger AG, Gurevich EV, Dalby KN, and Gurevich VV
- Subjects
- Animals, COS Cells, Chlorocebus aethiops, Enzyme Activation drug effects, Humans, Mitogen-Activated Protein Kinase 10 genetics, Enzyme Activators chemical synthesis, Enzyme Activators chemistry, Enzyme Activators pharmacology, Mitogen-Activated Protein Kinase 10 metabolism, Peptides chemical synthesis, Peptides chemistry, Peptides pharmacology, beta-Arrestin 1 chemistry, beta-Arrestin 2 chemistry
- Abstract
Three-kinase mitogen-activated protein kinase (MAPK) signaling cascades are present in virtually all eukaryotic cells. MAPK cascades are organized by scaffold proteins, which assemble cognate kinases into productive signaling complexes. Arrestin-3 facilitates JNK activation in cells, and a short 25-residue arrestin-3 peptide was identified as the critical JNK3-binding element. Here we demonstrate that this peptide also binds MKK4, MKK7, and ASK1, which are upstream JNK3-activating kinases. This peptide is sufficient to enhance JNK3 activity in cells. A homologous arrestin-2 peptide, which differs only in four positions, binds MKK4, but not MKK7 or JNK3, and is ineffective in cells at enhancing activation of JNK3. The arrestin-3 peptide is the smallest MAPK scaffold known. This peptide or its mimics can regulate MAPKs, affecting cellular decisions to live or die.
- Published
- 2016
- Full Text
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19. How genetic errors in GPCRs affect their function: Possible therapeutic strategies.
- Author
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Stoy H and Gurevich VV
- Abstract
Activating and inactivating mutations in numerous human G protein-coupled receptors (GPCRs) are associated with a wide range of disease phenotypes. Here we use several class A GPCRs with a particularly large set of identified disease-associated mutations, many of which were biochemically characterized, along with known GPCR structures and current models of GPCR activation, to understand the molecular mechanisms yielding pathological phenotypes. Based on this mechanistic understanding we also propose different therapeutic approaches, both conventional, using small molecule ligands, and novel, involving gene therapy.
- Published
- 2015
- Full Text
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20. Fluorescence lifetime imaging microscopy: in vivo application to diagnosis of oral carcinoma.
- Author
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Sun Y, Phipps J, Elson DS, Stoy H, Tinling S, Meier J, Poirier B, Chuang FS, Farwell DG, and Marcu L
- Subjects
- Animals, Cheek pathology, Cricetinae, Microscopy, Fluorescence methods, Mouth Neoplasms pathology, Microscopy, Fluorescence instrumentation, Mouth Neoplasms diagnosis
- Abstract
A compact clinically compatible fluorescence lifetime imaging microscopy (FLIM) system was designed and built for intraoperative disease diagnosis and validated in vivo in a hamster oral carcinogenesis model. This apparatus allows for the remote image collection via a flexible imaging probe consisting of a gradient index objective lens and a fiber bundle. Tissue autofluorescence (337 nm excitation) was imaged using an intensified CCD with a gate width down to 0.2 ns. We demonstrate a significant contrast in fluorescence lifetime between tumor (1.77+/-0.26 ns) and normal (2.50+/-0.36 ns) tissues at 450 nm and an over 80% intensity decrease at 390 nm emission in tumor versus normal areas. The time-resolved images were minimally affected by tissue morphology, endogenous absorbers, and illumination. These results demonstrate the potential of FLIM as an intraoperative diagnostic technique.
- Published
- 2009
- Full Text
- View/download PDF
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