Molecular Mechanism of Decreased Glutathione Content in Human Immunodeficiency Virus Type 1 Tat-transgenic Mice

Abstract

Human immunodeficiency virus (HIV) progressively depletes GSH content in humans.Although the accumulated evidence suggests a role of decreased GSH in the pathogenesis of HIV, significant controversy remains concerning the mechanism of GSH depletion, especially in regard to envisioning appropriate therapeutic strategies to help compensate for such decreased antioxidant capacity. Tat, a transactivator encoded by HIV, is sufficient to cause GSH depletion in vitro and is implicated in AIDS-associated Kaposi's sarcoma and B cell lymphoma. In this study, we report a decrease in GSH biosynthesis with Tat, using HIV-1 Tat transgenic (Tat؉) mice. A significant decline in the total intracellular GSH content in liver and erythrocytes of Tat؉ mice was accompanied by decreased ␥-glutamylcysteine synthetase regulatory subunit mRNA and protein content, which resulted in an increased sensitivity of ␥-glutamylcysteine synthetase to feedback inhibition by GSH. Further study revealed a significant reduction in the activity of GSH synthetase in liver of Tat؉ mice, which was linearly associated with their GSH content. Therefore, Tat appears to decrease GSH in vivo, at least partially, through modulation of GSH biosynthetic enzymes. Human immunodeficiency virus (HIV)1 infection is associated with a systemic decrease in the GSH content in humans (1, 2). Such biochemical alteration appears to be a direct consequence of retroviral infection rather than a late and indirect consequence of advanced disease state, is progressive in nature, and often occurs in the absence of any symptoms associated with AIDS (2-5). The importance of this decreased GSH is suggested by the fact that decreased GSH can enhance HIV expression via activation of NF-B (4, 6, 7) and contribute to diverse immune dysfunctions found in AIDS, including the loss of CD4ϩ T lymphocytes via apoptosis (8 -11). In fact, such decline in the GSH content may play a role in the increased drug toxicity and oxidative damage, often found in HIV-infected individuals (12-15). Furthermore, increased GSH or other sulfhydryl compounds can inhibit viral replication (7, 16), HIV-1 reverse transcriptase activity (16,17), and the late stage of viral expression (18). GSH content may even predict the survival of HIV-seropositive (HIVϩ) individuals (19,20). Therefore, it is perhaps crucial to determine whether this decrease in GSH occurs through an accelerated loss of GSH or specific modulation of its synthesis, especially as it relates to potential therapeutic strategies to compensate for such decreased antioxidant capacity. However, despite over a decade of intensive research on this subject, the exact mechanism(s) causing decreased GSH content with HIV remains unclear.Cells maintain their high GSH content in part by reducing GSSG back to GSH, preventing the loss of GSH as GSSG (12). Also, GSH is synthesized from its amino acid constituents via two consecutive reactions catalyzed by ␥-glutamylcysteine synthetase (GCS; glutamate-cysteine ligase, EC 6.3.2.2) and GSH synthetase (GS; EC6.3.2.3). I...

Regulation of glutathione synthesis

“…Treatment of rats with monocrotaline, a pyrrolizidine alkaloid, was also shown to increase the activity of GS to a comparable level as that of GCL (Yan and Huxtable, 1996). Interestingly, Choi and colleagues described decreased hepatic GSH levels, which correlated with a fall in GS activity in Tat transgenic mice (Choi et al, 2000). In this model, there was no change in the GCLC protein level or GCL activity when measured under saturating substrate conditions.…”

Inducible Alterations of Glutathione Levels in Adult Dopaminergic Midbrain Neurons Result in Nigrostriatal Degeneration

“…The samples were precipitated and sonicated twice in this solution to ensure complete GSH recovery. The samples were further processed and analyzed for their glutathione content using PerkinElmer (Wellesley, MA) HPLC system, equipped with LC-890 UV spectrophotometric detector and an autoinjector (Choi et al, 2000). Glutathione concentrations were normalized by protein assay using BCA protein assay kit, per the recommendations of the manufacturer (Pierce, Rockford, IL).…”

Role of AP-1 in the Coordinate Induction of Rat Glutamate-cysteine Ligase and Glutathione Synthetase bytert-Butylhydroquinone

“…Mutation inactivation of GS has been described (22). Recently two reports have shown a selective fall in GS activity which resulted in lowered GSH level (23,24). If GCL is rate-limiting, how can a change in GS activity influence the steady state GSH level?…”