Measuring Glutathione Redox Potential of HIV-1-infected Macrophages

Bhaskar, Ashima and Munshi, MohamedHusen and Khan, Sohrab Zafar and Fatima, Sadaf and Arya, Rahul and Jameel, Shahid and Singh, Amit (2015) Measuring Glutathione Redox Potential of HIV-1-infected Macrophages. In: JOURNAL OF BIOLOGICAL CHEMISTRY, 290 (2). pp. 1020-1038.

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Abstract

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 (H2O2), 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 similar to 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.

Item Type:	Journal Article
Publication:	JOURNAL OF BIOLOGICAL CHEMISTRY
Publisher:	AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC
Additional Information:	Copy right for this article belongs to the AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC, 9650 ROCKVILLE PIKE, BETHESDA, MD 20814-3996 USA
Department/Centre:	Division of Biological Sciences > Microbiology & Cell Biology
Date Deposited:	04 Mar 2015 12:06
Last Modified:	04 Mar 2015 12:06
URI:	http://eprints.iisc.ac.in/id/eprint/50948

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