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Measuring Glutathione Redox Potential of HIV-1-infected Macrophages
Redox signaling plays a crucial role in the pathogenesis of human immunodeficiency virus type-1 (HIV-1). The majority of HIV redox research relies on measuring redox stress using invasive technologies, which are unreliable and do not provide information about the contributions of subcellular compart...
Autores principales: | , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
American Society for Biochemistry and Molecular Biology
2015
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4294471/ https://www.ncbi.nlm.nih.gov/pubmed/25406321 http://dx.doi.org/10.1074/jbc.M114.588913 |
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author | Bhaskar, Ashima Munshi, MohamedHusen Khan, Sohrab Zafar Fatima, Sadaf Arya, Rahul Jameel, Shahid Singh, Amit |
author_facet | Bhaskar, Ashima Munshi, MohamedHusen Khan, Sohrab Zafar Fatima, Sadaf Arya, Rahul Jameel, Shahid Singh, Amit |
author_sort | Bhaskar, Ashima |
collection | PubMed |
description | Redox signaling plays a crucial role in the pathogenesis of human immunodeficiency virus type-1 (HIV-1). The majority of HIV redox research relies on measuring redox stress using invasive technologies, which are unreliable and do not provide information about the contributions of subcellular compartments. A major technological leap emerges from the development of genetically encoded redox-sensitive green fluorescent proteins (roGFPs), which provide sensitive and compartment-specific insights into redox homeostasis. Here, we exploited a roGFP-based specific bioprobe of glutathione redox potential (E(GSH); Grx1-roGFP2) and measured subcellular changes in E(GSH) during various phases of HIV-1 infection using U1 monocytic cells (latently infected U937 cells with HIV-1). We show that although U937 and U1 cells demonstrate significantly reduced cytosolic and mitochondrial E(GSH) (approximately −310 mV), active viral replication induces substantial oxidative stress (E(GSH) more than −240 mV). Furthermore, exposure to a physiologically relevant oxidant, hydrogen peroxide (H(2)O(2)), induces significant deviations in subcellular E(GSH) between U937 and U1, which distinctly modulates susceptibility to apoptosis. Using Grx1-roGFP2, we demonstrate that a marginal increase of about ∼25 mV in E(GSH) is sufficient to switch HIV-1 from latency to reactivation, raising the possibility of purging HIV-1 by redox modulators without triggering detrimental changes in cellular physiology. Importantly, we show that bioactive lipids synthesized by clinical drug-resistant isolates of Mycobacterium tuberculosis reactivate HIV-1 through modulation of intracellular E(GSH). Finally, the expression analysis of U1 and patient peripheral blood mononuclear cells demonstrated a major recalibration of cellular redox homeostatic pathways during persistence and active replication of HIV. |
format | Online Article Text |
id | pubmed-4294471 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2015 |
publisher | American Society for Biochemistry and Molecular Biology |
record_format | MEDLINE/PubMed |
spelling | pubmed-42944712015-01-21 Measuring Glutathione Redox Potential of HIV-1-infected Macrophages Bhaskar, Ashima Munshi, MohamedHusen Khan, Sohrab Zafar Fatima, Sadaf Arya, Rahul Jameel, Shahid Singh, Amit J Biol Chem Microbiology Redox signaling plays a crucial role in the pathogenesis of human immunodeficiency virus type-1 (HIV-1). The majority of HIV redox research relies on measuring redox stress using invasive technologies, which are unreliable and do not provide information about the contributions of subcellular compartments. A major technological leap emerges from the development of genetically encoded redox-sensitive green fluorescent proteins (roGFPs), which provide sensitive and compartment-specific insights into redox homeostasis. Here, we exploited a roGFP-based specific bioprobe of glutathione redox potential (E(GSH); Grx1-roGFP2) and measured subcellular changes in E(GSH) during various phases of HIV-1 infection using U1 monocytic cells (latently infected U937 cells with HIV-1). We show that although U937 and U1 cells demonstrate significantly reduced cytosolic and mitochondrial E(GSH) (approximately −310 mV), active viral replication induces substantial oxidative stress (E(GSH) more than −240 mV). Furthermore, exposure to a physiologically relevant oxidant, hydrogen peroxide (H(2)O(2)), induces significant deviations in subcellular E(GSH) between U937 and U1, which distinctly modulates susceptibility to apoptosis. Using Grx1-roGFP2, we demonstrate that a marginal increase of about ∼25 mV in E(GSH) is sufficient to switch HIV-1 from latency to reactivation, raising the possibility of purging HIV-1 by redox modulators without triggering detrimental changes in cellular physiology. Importantly, we show that bioactive lipids synthesized by clinical drug-resistant isolates of Mycobacterium tuberculosis reactivate HIV-1 through modulation of intracellular E(GSH). Finally, the expression analysis of U1 and patient peripheral blood mononuclear cells demonstrated a major recalibration of cellular redox homeostatic pathways during persistence and active replication of HIV. American Society for Biochemistry and Molecular Biology 2015-01-09 2014-11-18 /pmc/articles/PMC4294471/ /pubmed/25406321 http://dx.doi.org/10.1074/jbc.M114.588913 Text en © 2015 by The American Society for Biochemistry and Molecular Biology, Inc. Author's Choice—Final version full access. Creative Commons Attribution Unported License (http://creativecommons.org/licenses/by/3.0/) applies to Author Choice Articles |
spellingShingle | Microbiology Bhaskar, Ashima Munshi, MohamedHusen Khan, Sohrab Zafar Fatima, Sadaf Arya, Rahul Jameel, Shahid Singh, Amit Measuring Glutathione Redox Potential of HIV-1-infected Macrophages |
title | Measuring Glutathione Redox Potential of HIV-1-infected Macrophages |
title_full | Measuring Glutathione Redox Potential of HIV-1-infected Macrophages |
title_fullStr | Measuring Glutathione Redox Potential of HIV-1-infected Macrophages |
title_full_unstemmed | Measuring Glutathione Redox Potential of HIV-1-infected Macrophages |
title_short | Measuring Glutathione Redox Potential of HIV-1-infected Macrophages |
title_sort | measuring glutathione redox potential of hiv-1-infected macrophages |
topic | Microbiology |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4294471/ https://www.ncbi.nlm.nih.gov/pubmed/25406321 http://dx.doi.org/10.1074/jbc.M114.588913 |
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