Affiliations: Pharmaceutical and Biomedical Sciences, College of
Pharmacy, University of Georgia, Athens, GA 30602 USA | Faculty of Health and Social Care, London South Bank
University, 103 Borough Road, London SE1 0AA, UK | Office of Research, PO Box 26170, University of North
Carolina at Greensboro, Greensboro, NC 27402-6170, USA
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.
Journal: BioFactors, vol. 27, no. 1-4, pp. 157-165, 2006
