ABSTRACT

Reactive oxygen intermediates like hydrogen peroxide (HO) have been shown to serve as messengers in the induction of NF-B and, then, in the activation and replication of human immunodeficiency virus (HIV)-1 in human cells. Because HO can be converted into the highly reactive OH at various locations inside the cells, we started to investigate the generation of Reactive oxygen intermediates by photosensitization. This technique is based on the use of a photosensitizer which is a molecule absorbing visible light and which can be located at various sites inside the cell depending on its physicochemical properties. In this work, we used proflavine (PF), a cationic molecule having a high affinity for DNA, capable of intercalating between DNA base pairs. Upon visible light irradiation, intercalated PF molecules oxidize guanine residues and generate DNA single-strand breaks. In lymphocytes or monocytes latently infected with HIV-1 (ACH-2 or U1, respectively), this photosensitizing treatment induced a cytotoxicity, an induction of NF-B, and a reactivation of HIV-1 in cells surviving the treatment. NF-B induction by PF-mediated photosensitization was not affected by the presence of N-acetyl-L-cysteine while strong inhibition was recorded when the induction was triggered by HO or by phorbol 12-myristate 13-acetate. Another transcription factor like AP-1 is less activated by this photosensitizing treatment. In comparison with other inducing treatments, such as phorbol 12-myristate 13-acetate or tumor necrosis factor , the activation of NF-B is slow, being optimal 120 min after treatment. These kinetic data were obtained by following, on the same samples, both the appearance of NF-B in the nucleus and the disappearance of IB- in cytoplasmic extracts. These data allow us to postulate that signaling events, initiated by DNA oxidative damages, are transmitted into the cytoplasm where the inactive NF-B factor is resident and allow the translocation of p50/p65 subunits of NF-B to the nucleus leading to HIV-1 gene expression.

FOOTNOTES

*

This work has been supported in part by grants from the Belgian National Fund for Scientific Research (NFSR), the Belgian National Lottery, and Pasteur-Merieux (Lyon, France). The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore by hereby marked ``advertisement'' in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

§

Supported by a scholarship grant on oxidative stress and HIV infection from Pasteur-Merieux Sérum et Vaccins (France).

¶

Research Associate from the Belgian NFSR (Brussels, Belgium).

**

Research Director from the Belgian NFSR.

()

The abbreviations used are: HIV, human immunodeficiency virus; PF, proflavine; LTR, long terminal repeat; IL, interleukin; TNF, tumor necrosis factor; ROI, reactive oxygen intermediate; EMSA, electrophoretic mobility shift assay; PMA, phorbol 12-myristate 13-acetate; CHX, cycloheximide; NAC, N-acetyl-L-cysteine.

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