Investigating the role of 4E-T, an eIF4E-binding protein resident in P-bodies, in gene expression control

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DNA is copied into messenger RNA in the nucleus, and following transfer to the cytoplasm, mRNA is translated into protein. mRNAs are translated into protein at different rates, leading to vastly different protein levels, which in part explains why muscle and red blood cells, for example, are so distinct though sharing the same genes. Regulation of mRNA translation is principally controlled by eIF4E, which binds the unusual cap structure at the 5' end of mRNA. eIF4E is important because it interacts with eIF4G which recruits ribosomes, the translational machinery, to mRNAs. Alternative eIF4E-binding proteins exist, such as 4E-BPs and these prevent eIF4G binding to eIF4E and reduce translation. I propose to investigate a more recently discovered eIF4E-binding protein called 4E-T, which also competes with eIF4G for eIF4E-binding. Interestingly, 4E-T and a fraction of eIF4E is concentrated in punctate cytoplasmic "bodies" visible in the microscope, called P-bodies, which lack ribosomes, and are thought to be sites where mRNAs are stored and returned to translation, or where mRNAs are degraded. Despite considerable interest, the role of these P-bodies remains enigmatic, though they influence for example virus replication. Published and unpublished work from us and other investigators show that 4E-T regulates a sub-set of mRNAs, but it is not known what sort of proteins they encode, nor is it known whether 4E-T has to be in P-bodies to exert this control, what proteins it binds there, or what its modification by phosphorylation means for this control. These are the questions we want to answer, using human cells in culture. We believe this work is highly timely since aberrant levels of eIF4E (and its phosphorylation) has been implicated in cell growth, proliferaion and in cancer development, and therefore investigation of a factor that interacts with eIF4E is important. Moreover, the role of P-bodies is still relatively unexplored, and our work will make a significant contribution to our understanding of how mRNAs are regulated in these foci. Last, we're fortunate to collaborate with Professor Anne Willis from the MRC Toxicology Unit, Leicester, to undertake a part of the analysis of 4E-T mRNA targets, and to extend and benefit from a Translational Resource database there, containing data from our and similar experiments from other investigators. The proposed work is novel, focused and timely, and the outcomes will contribute to our understanding and possible exploitation of the regulation of translation initiation by eIF4E and its interacting factors.