Indexing <it>TNF-α </it>gene expression using a gene-targeted reporter cell line

• Main Authors: Engelhardt John F, Lei-Butters Diana, Yan Ziying, Leno Gregory H
• Format: Article
• Language: English
• Published: BMC 2009-02-01
• Series: BMC Biology
• Online Access: http://www.biomedcentral.com/1741-7007/7/8

<p>Abstract</p> <p>Background</p> <p>Current cell-based drug screening technologies utilize randomly integrated reporter genes to index transcriptional activity of an endogenous gene of interest. In this context, reporter expression is controlled by known genetic elements that may only partially capture gene regulation and by unknown features of chromatin specific to the integration site. As an alternative technology, we applied highly efficient gene-targeting with recombinant adeno-associated virus to precisely integrate a luciferase reporter gene into exon 1 of the HeLa cell tumor necrosis factor-alpha (<it>TNF-α</it>) gene. Drugs known to induce <it>TNF-α </it>expression were then used to compare the authenticity of gene-targeted and randomly integrated transcriptional reporters.</p> <p>Results</p> <p><it>TNF-α</it>-targeted reporter activity reflected endogenous <it>TNF-α </it>mRNA expression, whereas randomly integrated <it>TNF-α </it>reporter lines gave variable expression in response to transcriptional and epigenetic regulators. 5,6-Dimethylxanthenone-4-acetic acid (DMXAA), currently used in cancer clinical trials to induce <it>TNF-α </it>gene transcription, was only effective at inducing reporter expression from <it>TNF-α </it>gene-targeted cells.</p> <p>Conclusion</p> <p>We conclude that gene-targeted reporter cell lines provide predictive indexing of gene transcription for drug discovery.</p>