Analysis of Redox Gene Promoters and Structural Mechanisms, Focusing on Thioredoxin and Methionine Sulfoxide Reductase A

Abstract

When cells are exposed to oxidative stress reactive oxygen species (ROS) are formed, causing severe damage in the cell. To maintain the redox homeostasis, antioxidant systems are switched on. Two of the major redox proteins include thioredoxin (Trx) and Methionine Sulfoxide Reductase (MSR). Trx interacts directly and indirectly with several transcription factors to modulate binding to gene promoters. In particular, Nrf2 is known to be an important transcription factor in redox regulation. To further define the mechanisms by which the Trx promoter is regulated by oxidative stress, the HEC1B endometrial cancer cell line and MDA-­‐MB-­‐231 breast cancer cell line were transfected with various Trx gene promoter constructs that drive expression of a luciferase reporter. When cells were transfected with constructs containing the Trx gene promoter region where the Antioxidant Responsive Element (ARE) was deleted, no induction was observed in response to tBHQ, diamide or H2O2 utilising luciferase reporter activity assays. Thus, the results confirmed the published data that the ARE is one of the most important elements in activating the Trx gene promoter during oxidative stress. Furthermore, it was established that luciferase reporter assays using the same cell line grown in media supplemented with different serum yielded significantly different results. Cells grown in medium supplemented with the serum containing the highest selenium level resulted in the lowest fold induction of the Trx gene promoter, while sera with the lowest level resulted in the highest folld inductions.