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A High-throughput Cre-Lox Activated Viral Membrane Fusion Assay to Identify Inhibitors of HIV-1 Viral Membrane Fusion
Published on: August 14, 2018
xCT/SLC7A11 antiporter function inhibits HIV-1 infection
Jesse Rabinowitz1, Hamayun J Sharifi2, Hunter Martin2
1Department of Immunology and Microbial Disease, Albany Medical College, 47 New Scotland Avenue, Albany, NY, 12208, USA.
Abstract:
Human macrophages are protected by intrinsic antiviral defenses that provide moderate protection against HIV-1 infection. Macrophages that do become infected can serve as long-lived reservoirs, to disseminate HIV-1 to CD4+ T cells. Infection of macrophages with HIV-1 and HIV-2 is inhibited by constitutive mobilization of antioxidant response master transcription regulator Nrf2. The downstream mediator of this restriction was not identified. Among the tens of genes controlled directly by Nrf2 in macrophages, we found that xCT/SLC7A11, a 12-transmembrane, cystine-glutamate antiporter promotes antiretroviral activity. We show here that depletion of xCT mRNA increases HIV-1 infection. Reconstitution of xCT knock out cells with wild-type xCT but not a transport-deficient mutant restores anti-HIV-1 activity. Pharmacological inhibitors of xCT amino acid transport also increase infection. The block is independent of known restriction factors and acts against HIV-1 and HIV-2. Like the block triggered through Nrf2, xCT function impedes infection immediately before 2-LTR circle formation.
Insights
The cystine-glutamate antiporter xCT restricts HIV-1 and HIV-2 infection in macrophages. Depleting xCT or inhibiting its transport function increases viral infection, highlighting its role in intrinsic antiviral defense.
Area of Science:
- Immunology
- Virology
- Cell Biology
Background:
- Human macrophages possess intrinsic antiviral defenses against HIV-1.
- Infected macrophages act as persistent reservoirs, facilitating HIV-1 dissemination to CD4+ T cells.
- The antioxidant response master transcription regulator Nrf2 inhibits HIV-1 and HIV-2 infection in macrophages, but its downstream mediator was unknown.
Purpose of the Study:
- To identify the downstream mediator of Nrf2-induced antiretroviral activity in macrophages.
- To investigate the role of xCT/SLC7A11, a cystine-glutamate antiporter, in macrophage restriction of HIV infection.
Main Methods:
- Depletion of xCT mRNA using siRNA.
- Reconstitution of xCT knockout cells with wild-type or transport-deficient xCT mutants.
- Treatment with pharmacological inhibitors of xCT amino acid transport.
- Assessment of HIV-1 and HIV-2 infection levels.
- Analysis of viral DNA forms, including 2-LTR circles.
Main Results:
- Depletion of xCT mRNA significantly increased HIV-1 infection in macrophages.
- Reconstitution with wild-type xCT restored antiretroviral activity, while a transport-deficient mutant did not.
- Pharmacological inhibition of xCT amino acid transport also led to increased HIV infection.
- The restriction mediated by xCT is effective against both HIV-1 and HIV-2 and acts prior to 2-LTR circle formation.
- This restriction mechanism is independent of known HIV restriction factors.
Conclusions:
- The cystine-glutamate antiporter xCT/SLC7A11 is a key downstream mediator of Nrf2-induced antiretroviral activity in human macrophages.
- xCT function is crucial for restricting HIV-1 and HIV-2 infection, acting early in the viral life cycle.
- Targeting xCT or its transport function represents a potential strategy for enhancing intrinsic macrophage resistance to HIV.
