Regulation of hydrogen peroxide-signalling by redox-sensitive peroxiredoxin and thioredoxin proteins

This research will examine how a single cell fungus (yeast) uses antioxidants, that breakdown and detoxify hydrogen peroxide, to sense changes in the levels of hydrogen peroxide and regulate the response of cells to these changes. Hydrogen peroxide is produced in air-living cells as an unwanted byproduct of metabolism. Unchecked, hydrogen peroxide damages DNA, protein and lipid components of cells. This type of damage is involved in the initiation and progression of many diseases including heart disease, diabetes and cancer and also in the ageing process. Because it is so toxic hydrogen peroxide is also produced by plants and immune cells to kill invading pathogenic fungi and bacteria. To limit damage by chemicals such as hydrogen peroxide, cells have developed protective antioxidants which breakdown these chemicals. When cells encounter hydrogen peroxide in the environment this is sensed and signals are relayed that lead to the production of more antioxidants. Our previous research made the surprising discovery that, in addition to detoxifying hydrogen peroxide, one family of abundant, conserved antioxidants, the peroxiredoxins, is required by certain yeast to sense and respond to hydrogen peroxide by switching on these antioxidant defences. Subsequently, we have found that another antioxidant, thioredoxin, also plays a vital role in sensing and signalling environmental increases in hydrogen peroxide. Thus, this project will build on this strong platform of published and pilot data to understand how thioredoxin and peroxiredoxin together coordinate the response of the yeast Schizosaccharomyces pombe to hydrogen peroxide. The outcomes of this project will significantly extend our understanding of how fungi detect and respond to hydrogen peroxide. Given the importance of hydrogen peroxide in eliminating invading pathogenic microbes, we predict that an increased understanding of how antioxidants function in cells could uncover new therapeutic strategies for fungal infections. Although hydrogen peroxide is very toxic it has recently been discovered that plants and animals produce hydrogen peroxide as a signal to stimulate cells to divide, to move or to become specialised. Thus, these studies may also provide insight into how antioxidants affect these signalling processes.

Peroxiredoxins (PRX) and thioredoxin utilise thiol-active cysteines in the detoxification of hydrogen peroxide and the reduction of oxidised proteins, respectively. Indeed, the catalytic breakdown of hydrogen peroxide by PRX involves the reduction of oxidised cysteines in PRX by thioredoxin and results in the oxidation of cysteines in thioredoxin. Hence the redox states of thioredoxin and PRX are closely coupled. Work by us and others have established that PRX and thioredoxin have important roles in hydrogen peroxide-sensing and signalling. For example, we and others have found that, counterintuitively based on a role simply as antioxidant enzymes, PRX are required for hydrogen peroxide-induced gene expression in yeast. In the fission yeast Schizosaccharomyces pombe the transcription factors Atf1 and Pap1 are required for hydrogen peroxide-induced changes in gene expression. Atf1 is directly regulated by the p38/JNK-related MAPK Sty1, whereas Pap1 is activated by oxidation of key cysteine residues. Our studies have revealed that the PRX, Tpx1, is required for the activation of both Sty1 and Pap1 by hydrogen peroxide. Intriguingly, only the regulation of Pap1 oxidation requires both of the catalytic cysteines and the thioredoxin peroxidase activity of Tpx1. Thus, Tpx1 regulates Sty1 and Pap1 by distinct mechanisms. We have obtained pilot data indicating that thioredoxin also plays a vital part in the regulation of both Sty1 and Pap1 by hydrogen peroxide. However, this data indicates that thioredoxin plays very different roles to Tpx1 in the regulation of Pap1. A major goal of this study is to investigate the hypothesis that the redox balance between oxidised and reduced thioredoxin and PRX may be at the core of the sensing/signalling mechanisms that allow the fission yeast S. pombe to adapt and grow in the presence of different levels of hydrogen peroxide.