Systems biology analysis of global mRNA translational regulation in Saccharomyces cerevisiae

Abstract

mRNA translation is one of the fundamental and well controlled cellular process requiring the combined function of a large number of molecular components. The three main stages of translation, initiation, elongation and termination are facilitated by more than 20 proteins known as translation factors. Translation is the final step in the flow of genetic information,and regulation at this level allows for an immediate and rapid response to changes in physiological conditions. The control exerted at the systems level of translation has not precisely been characterized. Three different techniques have been employed to quantitative the control exerted by the respective translation factors. In the first approach, employing the microscopic techniques, in vivo intra-cellular distribution of translation elongation and release factors were analysed with TCM and GFP tags. The result indicates that the factors are cytoplasmically distributed which cannot influence the overall translational control. In the second approach, the protein expression levels of the elongation and release factors were titrated progressively to explore their control effects on global translation regulation. The endogenous promoter of each translation factor was substituted by the tetO7 synthetic promoter to regulate the expression level in response to varying concentrations of doxycycline. Measurement of protein synthesis rate and the growth rate at different levels of the elongation and release factors provide insight to system-level control. The results indicate that the elongation factors eEF1A and eEF2 and the release factor eRF1 exert an unexpectedly high degree of control over translation rate. Moreover, these factors, along with elongation factor eEF3 were found to be functionally dedicated to translation, in contrast to eEF1B and eRF3, which is evidently multifunctional. In the third approach, a mathematical model has been developed to represent the control landscape of the translational machinery. This translation model is a powerful tool that will be used in the quantitative analysis of translation when two factors are made limiting at a time. The extensive study carried out on the translational regulation of Saccharomyces cerevisiae reveals an interesting observation of the involvement of each translation factors. For the first time, the quantitative measurement of the translational regulation reveals the translational regulation exerted by individual translation factors.EThOS - Electronic Theses Online ServiceGBUnited Kingdo