The p53-inhibitor Pifithrin-α inhibits Firefly Luciferase activity in vivo and in vitro

Abstract

BackgroundPifithrin-α is a small molecule inhibitor of p53 transcriptional activity. It has been proposed that the use of pifithrin-α in conjunction with chemotherapeutic and radiation therapies for cancer will reduce the side effects of these treatments in normal tissue that still contains wild type p53. In addition, pifithrin-α provides a useful tool in the laboratory to investigate the function of p53 in model systems.ResultsWhile investigating the effects of pifithrin-α on the transcriptional activity of NF-κB, we observed a strong inhibition of reporter plasmids containing the firefly luciferase gene. Firefly luciferase is one of the most commonly used enzymes in reporter gene assays. In contrast, no inhibition of reporter plasmids containing Renilla luciferase or chloramphenicol acetyltransferase was observed. The inhibition of firefly luciferase activity by pifithrin-α was observed both in vivo and in vitro. Pifithrin-α did not inhibit firefly luciferase protein expression, but rather suppressed light production/emission, since addition of exogenous pifithrin-α to active extracts inhibited this activity. Furthermore, pifithrin-α also inhibited recombinant firefly luciferase protein activity.ConclusionsAmong its other biological activities, pifithrin-α is an inhibitor of firefly luciferase activity. Caution must therefore be taken when using this compound, which has been characterised as an inhibitor of p53 transcriptional activity, to investigate effects on gene expression using transiently transfected reporter plasmids. Furthermore, these results demonstrate that when using novel compounds, the choice of vectors used in the experimental procedures might be of great importance for the correct conclusions to be made.