Investigation of the selective reprogramming of translation during apoptosis

Protein synthesis is the process by which the information in the genetic material in the cell, DNA is converted via an intermediary substrate mRNA, into proteins. For proteins to be synthesised the mRNA must interact with a large complex called the ribosome which consists of RNAs and proteins. Ribosomes are able to decode the genetic information that is held in the mRNA and carry out the synthesis of the proteins. There are two distinct mechanisms by which mRNAs can interact with the ribosomes. The most common mechanism requires the binding of a protein complex to the 5' end of the mRNA and this complex then recruits the ribosome. However, certain mRNAs contain 5' regions that do not code for sections of proteins (termed untranslated regions; UTRs) and these sequences of RNA harbour the information that is required to form a complex RNA structure. These RNA structures allow the ribosome to be recruited to the mRNA generally a considerable distance from the 5' end and so this method of ribosome recruitment has been termed internal ribosome entry. Interestingly, messages that use internal ribosome entry generally encode proteins that are involved in the processes of cell growth and cell death. We have shown that during cell death there is selective use of internal ribosome entry to allow synthesis of the proteins that are required for this process to proceed. The overall aim of this proposal is to understand how this mechanism functions during apoptosis.

Control of protein synthesis is one of the major mechanisms of regulating gene expression and allows a cell to respond rapidly to changing conditions without the requirement for the synthesis of new mRNA. During many types of patho-physiological stress conditions there is a large decrease in the global rate of protein synthesis, and a re-programming of translation, which allows specific subsets of mRNAs to become polysomally associated. This appears to be essential for either recovery from cell stress, or for the cellular process (e.g. apoptosis or mitosis) to proceed. Our recent data show that during apoptosis there is increased polysomal association of mRNAs that encode proteins which have functions in chromatin remodelling processes, cell cycle and certain cell surface receptors. In contrast, following UV exposure (there is increased recruitment into polysomes of mRNAs that encode enzymes that repair UV damage and a subsequent increase in the synthesis of these proteins. In eukaryotic cells there are two mechanisms that are used to initiation translation, which are termed cap-dependent scanning and internal ribosome entry (mediated by a large complex structural element; IRES). We have shown that 70% of the mRNAs that remain polysomal during apoptosis contain IRESs that are active during this process. However, to be able to understand how selective translational re-programming occurs it is important to identify the specific IRES-trans-acting factors, ITAFs that mediate this process.The overall aims of this proposal are to determine the molecular mechanisms used that permit the association of specific mRNAs with actively translating ribosomes (polysomes) during apoptosis. Thus we propose i) to identify proteins that allow IRES-mediated translation during apoptosis, ii) determine how these ITAFs are in turn regulated, iii) to identify in which subcellular compartments the mRNAs are stored under control conditions and how they re-localise during apoptosis.