A quantitative bioassay for HIV-1 gene expression based on UV activation: effect of glycyrrhizic acid.
Previous reports have shown that HIV-LTRcat constructs stably transfected in HeLa cells are inducible after exposure to UV light. We have optimized this system for studying the effect of drugs on HIV-1 gene expression. The maximum UV response was observed in quiescent stationary cells stimulated with fresh medium for 3h. Glycyrrhizic acid suppressed UV-induced HIV gene expression in a concentration-dependent manner. The inhibitory effect was strongest when GL was added immediately after UV exposure; it was still evident when GL was added at 5 h, it was completely lost at 10 h, after UV exposure. The inhibitory effect was even more pronounced if the cells were pretreated with sub-effective dose (0.0012 mM) of GL prior to UV exposure. The IC50 values with and without pretreatment were 0.04 and 0.38 mM, respectively. Cell proliferation and viability were not affected by GL at doses as high as 2.4 mM. The inhibitory effect of GL on UV-induced CAT activity correlated with the complete inhibition of binding activities of NF-kappaB p65, NF-kappaB p50, c-Fos, and c-Rel. Thus, the UV-based bioassay as proposed here can be exploited for the routine screening of the compounds that interfere with HIV-1 gene expression.
Cherng
Jaw-Ming
JM
Department of Internal Medicine, Tzu Chi General Hospital, Hualien, Taiwan, ROC.
Lin
Hsiung-Ju
HJ
Hsu
Yung-Han
YH
Hung
Man-Shan
MS
Lin
Jung-Chung
JC
D019380:Anti-HIV Agents / Q000494:pharmacology*; D001681:Biological Assay / Q000379:methods; D015500:Chloramphenicol O-Acetyltransferase / Q000235:genetics / Q000378:metabolism; D015870:Gene Expression / Q000187:drug effects*; D015967:Gene Expression Regulation, Viral; D017930:Genes, Reporter; D019695:Glycyrrhizic Acid / Q000494:pharmacology*; D016325:HIV Long Terminal Repeat; D015497:HIV-1 / Q000187:drug effects* / Q000235:genetics* / Q000378:metabolism / Q000528:radiation effects; D006367:HeLa Cells; D006801:Humans; D008826:Microbial Sensitivity Tests; D016328:NF-kappa B / Q000096:biosynthesis; D052002:NF-kappa B p50 Subunit; D011401:Promoter Regions, Genetic; D016760:Proto-Oncogene Proteins c-fos / Q000096:biosynthesis; D020569:Proto-Oncogene Proteins c-rel / Q000096:biosynthesis; D051996:Transcription Factor RelA; D014157:Transcription Factors / Q000096:biosynthesis; D014162:Transfection; D014466:Ultraviolet Rays
D019380:Anti-HIV Agents; D016328:NF-kappa B; D052002:NF-kappa B p50 Subunit; D016760:Proto-Oncogene Proteins c-fos; D020569:Proto-Oncogene Proteins c-rel; D051996:Transcription Factor RelA; D014157:Transcription Factors; D019695:Glycyrrhizic Acid; D015500:Chloramphenicol O-Acetyltransferase