Grant: Discovery Grants Program - Individual

One of the most fundamental activities in living systems is to express their genetic information (DNA) into proteins, the molecular machines that carry out most of the biological functions that keep an organism alive. The copying of DNA into proteins occurs in two steps: first, DNA is copied into mRNA (messenger ribonucleic acid; chemically similar to DNA); second, mRNA is used as a template to make specific proteins. For a given gene, the amount of protein is therefore dependent on the amount of mRNA. An excess or shortage of many proteins due to changes in the abundance of mRNA can lead to diseases, such as cancer. The abundance of mRNAs is highly dependent on the rate of its degradation. Therefore, an understanding of how the mRNA degradation machinery works is critical for increasing our basic scientific knowledge of molecular biology, as well as for revealing information on the pathogenesis of some human diseases.In 2009 our lab fortuitously discovered that apurinic/apyrimidinic endonuclease 1 (APE1), an enzyme well-known to play a role in DNA repair, has an ability to degrade specific mRNAs in vitro and can control the expression of specific mRNA in cells. Since our pioneering discovery at least three similar enzymes have been discovered to be at the nexus of DNA and RNA metabolism. For many of these enzymes, including APE1, the biological significance of their enzymatic ribonuclease activity (ability to cleave and degrade RNA) is still unknown.The major goal of my proposed research is to understand the ribonuclease function of the endogenous form as well as the recently discovered secreted form of APE1. We will generate two types of cell systems engineered to carry the wild type of APE1 and its variants to help us test our hypotheses. We will determine whether there is correlation between the ribonuclease activity and impact on cell viability by wild-type APE1 and its variants (Aim I). Using a technology known as RNA-seq, we aim to determine the cellular RNA targets of the ribonuclease activity of APE1 (Aim I). We will determine whether the secreted wild-type APE1 and its most common variant D148E have ribonuclease activity and if this has any effect on cell viability and cell death (Aim II). We will investigate the mechanism whereby APE1 controls an important RNA-binding protein involved in tumorigenesis (Aim III).The proposed research is expected to provide insights into the ribonuclease function of APE1 in mammalian cells, which will help broaden our understanding of the regulation of gene expression and how the processes of DNA repair and RNA metabolism may be linked.