Translation regulation elements in both the 5` and 3` untranslated region; how do they coexist?

Recently, a completely new way of controlling gene expression has been identified. This has come to light after the discovery of a whole new class of genes which, unlike most genes, do not produce proteins. Instead they make very small RNA molecules, called microRNAs. There are at least 800 microRNAs within the human genome which have different effects. They work by binding to the messenger RNA of other genes and inhibiting the production of the proteins made from these genes a process known as translation. Each of these 800 small molecules is believed to interact with 100 other genes, thus adding to the complex regulation of the human genome. Already it has become clear that malfunction of miRNA regulation is associated with a growing list of human diseases, including cancer, diabetes, and viral infections. Other methods of regulating expression of genes at the level of translation also exist, and little is known about how these methods interact with miRNA regulation. This study will analyse the interaction between miRNAs and other control elements in regulating specific genes. It will also look for changes in this regulation that occur when cells are stressed, as many changes in translation occur under these conditions. This study will lead to a greater understanding of how gene expression is controlled and the complexity of the human genome.

Control of protein synthesis can be both global and specific to particular mRNAs. In the latter case it is often mediated by defined elements in the 5' and 3' untranslated regions (UTRs) of the mRNAs. For example, an internal ribosome entry site (IRES) in the 5' UTR of an mRNA can be used to allow translation initiation under conditions of cell stress when cap-dependent scanning is compromised (e.g. during apoptosis, heat shock, genotoxic shock and hypoxia). It has also been shown that protein synthesis can be negatively regulated by microRNAs (miRNAs) which bind to target sites in the 3' UTRs of mRNAs and repress their translation. Data from our laboratory and others have shown that modulation of gene expression can occur by both miRNA-mediated regulation and internal ribosome entry on the same mRNA. However, studies into the 5` and 3` control elements have been performed in isolation from one another and it is likely that these regulatory elements act in concert to coordinate gene expression. The aim of this proposal is to assess the relative contributions that IRESs and miRNA regulation make to the expression of these mRNAs. It is proposed to use c-myc and CAT-1 mRNAs in this study as we and others have shown that both of these mRNAs contain IRESs in their 5' UTRs and miRNA target sites in their 3' UTRs. The following questions will be addressed: 1) How does the presence of an IRES in an mRNA influence the ability of miRNAs to repress translation? 2) Does the relationship between IRESs and miRNA translational control elements change under conditions of cell stress? 3) Why have two distinct miRNA repression mechanisms evolved, one acting at the initiation of protein synthesis and the other post-initiation? It is envisaged that information gained from these studies will be applicable to other mRNAs and biological systems.