The influence of miRNAs on mRNA degradation

Post-transcriptional processes play a crucial role in the regulation of eukaryotic gene expression. Over the past decade miRNAs (an abundant class of small non-coding RNAs) emerged as key regulators and have entirely revolutionized the way we think about post-transcriptional gene regulation. miRNAs play an essential role in diverse processes such as development, differentiation, cellular growth control, stress response and disease development (e.g. cancer). In the proposed project we plan to investigate the detailed molecular mechanism underlying the diverse functions of miRNAs. It is well established that miRNAs bind their target mRNA and inhibit its translation and/or accelerate mRNA turnover. In the first part of the project we will establish kinetic tools allowing us to dissect the various steps involved in mRNA degradation. With this powerful tools in hand we can address unresolved questions such as which particular steps of mRNA degradation is accelerated by miRNAs and what determines the varying contribution of mRNA decay to the miRNA response. Secondly, we propose to establish a system using tagged miRNA targets. In combination with mass spectrometry and chemical probing approaches we will monitor changes, induced by miRNA binding, in the protein composition and the RNA conformation of the highly dynamic mRNA-protein (mRNP) complex. The knowledge of structural changes of the mRNP complex upon binding of the miRNA is a fundamental part of our efforts to understand the mechanism of miRNA function at a molecular level. Overall, the results of this project will significantly contribute to our understanding of the general mRNA decay pathway and the miRNA-mediated mRNA decay in particular. While the initial studies will be done in Drosophila cells, the results will be important for studying these pathways in higher eukaryotes.

Posttranscriptional processes play an essential role in the regulation of gene expression in eukaryotes. An essential step is the degradation of mRNAs, which determines the lifetime of an mRNA in the cell. This in turn influences the amount of protein, which can be produced from a single mRNA. In particular under changing environmental conditions the change in mRNA lifetime, e.g. through binding of miRNAs, is an important mechanism to adjust to these changes. It is still unknown which features of an mRNA are important for determining the length of its life. In this project we could establish two systems, which allow us to monitor the half-life of an mRNA in Drosophila cells. Since the degradation of mRNAs is a multi-step process, it is important to note that one of the methods can be used to monitor the various different steps of mRNA degradation in Drosophila cells. Very common environmental changes that organisms have to adapt to are temperature changes. In particular the family of heat shock proteins is known to fulfill important functions under these conditions. While it is known that its mRNA half-life is accelerated or decelerated depending on the environmental conditions, it is entirely unclear how this is achieved in cells. In this project we could get evidence that the binding of a factor to the mRNA of the heat shock protein is important for changes in its mRNA half-life.