Interferon-α Regulates Glutaminase 1 Promoter through STAT1 Phosphorylation: Relevance to HIV-1 Associated Neurocognitive Disorders

Lixia Zhao,Yunlong Huang,Norman Curthoys,Lynn Taylor,Yi Wang,Changhai Tian,Jialin Zheng,Hamilton Vernon,Roberto F Speck

doi:10.1371/journal.pone.0032995

Abstract

HIV-1 associated neurocognitive disorders (HAND) develop during progressive HIV-1 infection and affect up to 50% of infected individuals. Activated microglia and macrophages are critical cell populations that are involved in the pathogenesis of HAND, which is specifically related to the production and release of various soluble neurotoxic factors including glutamate. In the central nervous system (CNS), glutamate is typically derived from glutamine by mitochondrial enzyme glutaminase. Our previous study has shown that glutaminase is upregulated in HIV-1 infected monocyte-derived-macrophages (MDM) and microglia. However, how HIV-1 leads to glutaminase upregulation, or how glutaminase expression is regulated in general, remains unclear. In this study, using a dual-luciferase reporter assay system, we demonstrated that interferon (IFN) α specifically activated the glutaminase 1 (GLS1) promoter. Furthermore, IFN-α treatment increased signal transducer and activator of transcription 1 (STAT1) phosphorylation and glutaminase mRNA and protein levels. IFN-α stimulation of GLS1 promoter activity correlated to STAT1 phosphorylation and was reduced by fludarabine, a chemical that inhibits STAT1 phosphorylation. Interestingly, STAT1 was found to directly bind to the GLS1 promoter in MDM, an effect that was dependent on STAT1 phosphorylation and significantly enhanced by IFN-α treatment. More importantly, HIV-1 infection increased STAT1 phosphorylation and STAT1 binding to the GLS1 promoter, which was associated with increased glutamate levels. The clinical relevance of these findings was further corroborated with investigation of post-mortem brain tissues. The glutaminase C (GAC, one isoform of GLS1) mRNA levels in HIV associated-dementia (HAD) individuals correlate with STAT1 (p<0.01), IFN-α (p<0.05) and IFN-β (p<0.01). Together, these data indicate that both HIV-1 infection and IFN-α treatment increase glutaminase expression through STAT1 phosphorylation and by binding to the GLS1 promoter. Since glutaminase is a potential component of elevated glutamate production during the pathogenesis of HAND, our data will help to identify additional therapeutic targets for the treatment of HAND.

Highlights

HIV-1 associated neurocognitive disorders (HAND) develop during progressive HIV-1 infection and are characterized by cognitive impairments, behavioral disorders and potential progressive motor abnormalities as a consequence of neuronal damage during prolonged inflammation in the central nervous system (CNS) [1]
Other transcription factors that have the potential to bind to the promoter include activator protein 1 (AP-1), nuclear factor 1 (NF1) and specificity protein 1 (SP-1), which are typically constitutively active in cells
When we looked for the inducible transcription factors, we found up to 8 sequences predicted to be STAT binding sites, indicating the factors that regulate STAT activation might have the potential to regulate the glutaminase 1 (GLS1) promoter

Summary

Introduction

HAND develop during progressive HIV-1 infection and are characterized by cognitive impairments, behavioral disorders and potential progressive motor abnormalities as a consequence of neuronal damage during prolonged inflammation in the CNS [1]. Our previous work has demonstrated that HIV-1-infected MDM and microglia produce more extracellular glutamate than uninfected cells [8,9], an effect that is dependent on mitochondrial glutaminase [10]. In the CNS, mitochondrial glutaminase is a key enzyme that converts glutamine to glutamate, an enzymatic process that has the potential to cause neuronal toxicity [8,11]. Kidney-type glutaminase (KGA) is highly expressed in the brain [12]; it has various isoforms generated through tissue-specific alternative splicing, including GAC, a KGA variant [13]. The human GAC isoform is upregulated in MDM and microglia during HIV infection [9,15]. Despite extensive studies of upstream molecular stimuli, those that are responsible for HIV-1-mediated GAC upregulation and glutamate overproduction, are still obscure

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