5-Ethynyl-2'-deoxycytidine and 5-ethynyl-2'-deoxyuridine are differentially incorporated in cells infected with HSV-1, HCMV, and KSHV viruses Microbiology

Nucleoside analogues are a valuable experimental tool. Incorporation of these molecules into newly synthesized DNA (i.e. pulse-labeling) is used to monitor cell proliferation or to isolate nascent DNA. Some of the most common nucleoside analogues used for pulse-labeling of DNA in cells are the deoxypyrimidine analogues 5-ethynyl-2′-deoxyuridine (EdU) and 5-ethynyl-2′-deoxycytidine (EdC). Click chemistry enables conjugation of an azide molecule tagged with a fluorescent dye or biotin to the alkyne of the analog, which can then be used to detect incorporation of EdU and EdC into DNA. The use of EdC is often recommended because of the potential cytotoxicity associated with EdU during longer incubations. Here, by comparing the relative incorporation efficiencies of EdU and EdC during short 30-min pulses, we demonstrate significantly lower incorporation of EdC than of EdU in noninfected human fibroblast cells or in cells infected with either human cytomegalovirus or Kaposi's sarcoma-associated herpesvirus. Interestingly, cells infected with herpes simplex virus type-1 (HSV-1) incorporated EdC and EdU at similar levels during short pulses. Of note, exogenous expression of HSV-1 thymidine kinase increased the incorporation efficiency of EdC. These results highlight the limitations when using substituted pyrimidine analogues in pulse-labeling and suggest that EdU is the preferable nucleoside analogue for short pulse-labeling experiments, resulting in increased recovery an...
Source: Journal of Biological Chemistry - Category: Chemistry Authors: Tags: Methods and Resources Source Type: research

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The hijacking of cellular function through expression of proteins that interfere with the activity of cellular enzymes and regulatory complexes is a common strategy used by viruses to remodel the cell environment in favor of their own replication and spread. Here we report that the ubiquitin deconjugases encoded in the N-terminal domain of the large tegument proteins of Epstein-Barr virus (EBV), Kaposi Sarcoma herpesvirus (KSHV) and human cytomegalovirus (HCMV), but not herpes simplex virus-1 (HSV-1), target an early step of the IFN signaling cascade that involves the formation of a trimolecular complex with the ubiquitin ...
Source: Frontiers in Immunology - Category: Allergy & Immunology Source Type: research
Conclusion and Future Perspectives This review illustrates our current knowledge of USP7, including its source and characterization, structure, binding partners and substrates in various biological processes. Besides, how USP7 regulates various aspects of a cell under both normal and pathological states are elaborated in detail. As the processes of ubiquitination and deubiquitination are extremely dynamic and context-specific, a series of studies have linked USP7 to different cancers. The biology, particularly the immune oncology mechanisms, reveal that USP7 inhibitors would be useful drugs, thus it is vital to develop hi...
Source: Frontiers in Pharmacology - Category: Drugs & Pharmacology Source Type: research
Reena Goswami1, Gayatri Subramanian2, Liliya Silayeva1, Isabelle Newkirk1, Deborah Doctor1, Karan Chawla2, Saurabh Chattopadhyay2, Dhyan Chandra3, Nageswararao Chilukuri1 and Venkaiah Betapudi1,4* 1Neuroscience Branch, Research Division, United States Army Medical Research Institute of Chemical Defense, Aberdeen, MD, United States 2Department of Medical Microbiology and Immunology, University of Toledo College of Medicine and Life Sciences, Toledo, OH, United States 3Roswell Park Comprehensive Cancer Center, Buffalo, NY, United States 4Department of Physiology and Biophysics, Case Western Reserve University, Clev...
Source: Frontiers in Oncology - Category: Cancer & Oncology Source Type: research
In conclusion, our results showed that CMV infection may play a role in the initiation or amplification of inflammatory responses in AOSD. Introduction Adult-onset Still's disease (AOSD) is a rare but clinically well-known systemic inflammatory disease. It is typically characterized by a high spiking fever, evanescent skin rash, arthralgia, sore throat and neutrophilia (1–3). Even though the etiology of AOSD remains unknown, there is evidence that it's triggered by environmental factors with genetic predisposition (4). It has long been suspected that viral infections might contribute to the on...
Source: Frontiers in Immunology - Category: Allergy & Immunology Source Type: research
In this study, we showed that KSHV-infected cells induce interferon-stimulated genes (ISGs) response but not type I interferon in uninfected bystander cells using EVs. mRNA microarray analysis showed that ISGs and IRF-activating genes were prominently activated in EVs from KSHV-infected cells (KSHV EVs)-treated human endothelial cells, which were validated by RT-qPCR and western blot analysis. We also found that this response was not associated with cell death or apoptosis by virus infection. Mechanistically, the cGAS-STING pathway was linked with these KSHV EVs-mediated ISGs expressions, and mitochondrial DNA on the surfa...
Source: Frontiers in Immunology - Category: Allergy & Immunology Source Type: research
María Maximina B. Moreno-Altamirano1*, Simon E. Kolstoe2 and Francisco Javier Sánchez-García1* 1Laboratorio de Inmunorregulación, Departamento de Inmunología, Escuela Nacional de Ciencias Biológicas, Instituto Politécnico Nacional, Mexico City, Mexico 2School of Health Sciences, University of Portsmouth, Portsmouth, United Kingdom Over the last decade, there has been significant advances in the understanding of the cross-talk between metabolism and immune responses. It is now evident that immune cell effector function strongly depends on the metabolic pathway in w...
Source: Frontiers in cellular and infection microbiology - Category: Microbiology Source Type: research
Christine I. Alston1,2 and Richard D. Dix1,2* 1Department of Biology, Viral Immunology Center, Georgia State University, Atlanta, GA, United States 2Department of Ophthalmology, Emory University School of Medicine, Atlanta, GA, United States Suppressor of cytokine signaling (SOCS) proteins provide selective negative feedback to prevent pathogeneses caused by overstimulation of the immune system. Of the eight known SOCS proteins, SOCS1 and SOCS3 are the best studied, and systemic deletion of either gene causes early lethality in mice. Many viruses, including herpesviruses such as herpes simplex virus and cytomega...
Source: Frontiers in Immunology - Category: Allergy & Immunology Source Type: research
Monica Parodi1, Herman Favoreel2, Giovanni Candiano3, Silvia Gaggero4, Simona Sivori4,5, Maria Cristina Mingari1,4,5, Lorenzo Moretta6, Massimo Vitale1 and Claudia Cantoni4,5,7* 1Immunology Operative Unit, IRCCS Ospedale Policlinico San Martino, Genoa, Italy 2Department of Virology, Parasitology and Immunology, Faculty of Veterinary Medicine, Ghent University, Merelbeke, Belgium 3Laboratory of Molecular Nephrology, IRCCS Istituto Giannina Gaslini, Genoa, Italy 4Department of Experimental Medicine, University of Genoa, Genoa, Italy 5Center of Excellence for Biomedical Research, University of Genoa, Genoa, Italy ...
Source: Frontiers in Immunology - Category: Allergy & Immunology Source Type: research
LITFL • Life in the Fast Lane Medical Blog LITFL • Life in the Fast Lane Medical Blog - Emergency medicine and critical care medical education blog aka Tropical Travel Trouble 009 The diagnosis of HIV is no longer fatal and the term AIDS is becoming less frequent. In many countries, people with HIV are living longer than those with diabetes. This post will hopefully teach the basics of a complex disease and demystify some of the potential diseases you need to consider in those who are severely immunosuppressed. While trying to be comprehensive this post can not be exhaustive (as you can imagine any patient with ...
Source: Life in the Fast Lane - Category: Emergency Medicine Authors: Tags: Clinical Cases Tropical Medicine AIDS art cryptococcoma cryptococcus HIV HIV1 HIV2 PEP PrEP TB toxoplasma tuberculoma Source Type: blogs
ong Liu The CRISPR/Cas9 system has been applied in the genome editing and disruption of latent infections for herpesviruses such as the herpes simplex virus, Epstein–Barr virus, cytomegalovirus, and Kaposi’s sarcoma-associated herpesvirus. CRISPR/Cas9-directed mutagenesis can introduce similar types of mutations to the viral genome as can bacterial artificial chromosome recombination engineering, which maintains and reconstitutes the viral genome successfully. The cleavage mediated by CRISPR/Cas9 enables the manipulation of disease-associated viral strains with unprecedented efficiency and precision...
Source: Viruses - Category: Virology Authors: Tags: Review Source Type: research
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