Retroviral reporter systems for the assessment of activity of stress-induced signal transduction pathways controlled by p53, HIF-1 and HSF-1 transcription factors

Abstract

The tumor suppressor p53, hypoxia-inducible factor 1 (HIF-1), and heat-shock factor 1 (HSF-1) are the key transcription factors involved in the stress response to damage of the genetic material, hypoxia, or heat shock, respectively. Since these factors play a considerable role in tumor development and progression, modulation of their activities may be used in cancer therapy. To quantitate the transcriptional activities of p53, HIF-1, and HSF-1, reporter constructs were obtained on the basis of retroviral and lentiviral vectors, allowing generation of reporter cultures from almost all cell types. Induction of the reporter β-galactosidase gene, which reflected the activity of p53 or HIF-1, proved to depend on the concentration of an activating agent and to correlate with induction of the endogenous target genes of the transcription factors. Inhibition of p53 or HIF-1α expression with specific small interfering RNAs (siRNAs) completely abolished the activating effect of stress conditions. The reporter constructs were proposed for screening chemical compounds or genetic elements (siRNA or cDNA libraries) that modulate the activity of p53, HIF-1, or HSF-1 and for studying the components of the relevant signaling pathways.