An integrated systems approach to posttranscriptional gene expression

As with many of the essential processes in living cells, a great deal of qualitative information about posttranscriptional gene expression has been accumulated, yet our quantitative understanding of the pathways of translation and mRNA turnover as complete processes (systems) is minimal. This means that we are frequently unable to model adequately either the processes themselves or their regulation, thus limiting potential progress in our understanding of the molecular basis of function and control in the translation and mRNA turnover machineries. This is particularly evident in relation to the control of the switching of mRNA molecules from translatable templates to targets of degradation, which remains completely uncharacterised in quantitative terms. I would like to dedicate five years of my research career to the application of a novel combination of advanced techniques drawn from a number of disciplines in order to place our understanding of these steps of posttranscriptional gene expression in a quantitative framework that can be used as the basis for testable and predictive modelling. This will enable a step-change in our fundamental understanding of these key processes in the cell because all structural and functional data will be built into one coherent framework. The techniques to be applied will include process control analysis, fast reaction kinetic analysis, studies of molecular structure and dynamics, and mathematical modelling. The overall strategy was originally embodied in my review of 1998 (McCarthy, J.E.G. Posttranscriptional control of gene expression in yeast. Microbiol. Molec. Biol. Rev. 62, 1492-1553), but only now has the field reached a sufficiently advanced stage of experimental/theoretical progress where an intensive approach of this scale can be truly effective. In the wider context of the BBSRC's commitment to supporting members of the scientific community who can promote the strategic aims of the research council, this ties in with my more general commitment to promoting interdisciplinary research and training in the UK, as evident from my leadership of the MIB project and of the RSC Chemical Biology Interface Forum, as well as from my managerial involvement with the Manchester Centre for Integrated Systems Biology (MCISB) and its associated Doctoral Training Centre.

This will be a coherent research programme combining several complementary threads of investigation focused on eukaryotic posttranscriptional gene expression, using yeast as a model system. The first component will be the development of a comprehensive systems model for rate control in the eukaryotic translation pathway and in the route that leads mRNA molecules into pathways of degradation. This work will be enhanced to incorporate spatial and temporal resolution into the analysis via an approach termed Spatially (and Temporally) Resolved Molecular Systems Analysis [SReMSA (STReMSA)]. This will provide a detailed, quantitative platform for understanding the dynamic control and regulation of eukaryotic posttranscriptional gene expression. The above systems approach will be complemented by a series of structural, allied to functional, studies at the molecular level that will provide the mechanistic detail that is needed for an understanding of component steps in the pathways. X-ray crystallography, NMR and cryo-EM will all be used to enhance our structural understanding of the ribosome and its complexes with translation factors, and of key components of the mRNA decay pathway. Mechanistic studies will involve the use of FRET, EPR, fast reaction spectroscopy and single molecule studies (particularly AFM and TIRFM). Various forms of modelling will be used to analyse the data generated in this research programme, including rate control modelling, local dynamics and kinetic modelling, intermolecular interactions modelling, and structural modelling.