Activity of the dietary antioxidant ergothioneine in a virus gene‐based assay for inhibitors of HIV transcription

Abstract The “Long Terminal Repeat” (LTR) of HIV‐1 is the target of cellular transcription factors such as NF‐κB, and serves as the promoter‐enhancer for the viral genome when integrated in host DNA. Various LTR‐reporter gene constructs have been used for in vitro studies of activators or inhibitors of HIV‐1 transcription, e.g., to show that antioxidants such as lipoic acid and selenium inhibit NF‐κB‐dependent HIV‐1 LTR activation. One such construct is the pHIVlacZ plasmid, with the HIV‐1 LTR driving expression of the lacZ gene (encoding β‐galactosidase, β‐gal). Typically, for inhibitor screening, cells transfected with pHIVlacZ are activated using tumor necrosis factor‐α (TNF‐α), and the colorimetric o‐nitrophenol assay is used to assess changes in β‐gal activity. A variant of this assay was developed as described here, in which LTR activation was induced by pro‐fs, a novel HIV‐1 gene product encoded via a −1 frameshift from the protease gene. Cotransfection of cells with pHIVlacZ along with a pro‐fs construct produced a signifcant increase in β‐gal activity over controls. L‐ergothioneine dose dependently inhibited both TNF‐α‐mediated and pro‐fs‐mediated increases in β‐gal activity, with an IC 50 of about 6 mM. Thus antioxidant strategy involving ergothioneine derived from food plants might be of benefit in chronic immunodeficiency diseases.