Translational elongation plays crucial role in mediating the expression of proteins and finally affecting various biological reactions and processes in living organisms. Reversely, the efficiency and rate of translational elongation is commonly regulated by amounts of internal elements, such as special codon or sequences, and external conditions, such as different nutrient conditions. The relevant effects have been well investigated a lot based on transcriptome or proteome analysis and ribosome profiling. Specifically, previous researches commonly identified lessening of protein production and ribosomal loading, which suggest the variation of elongation efficiency or ribosome stalling derived from poor genetic elements or worse nutrient conditions. We have found that, the existence of poly-proline sequence in nuclear-encoded mitochondrial proteins, such as ATP3 and Tim50, shows negative effects in the translation efficiency and the final products, while positive effects in mRNA localization on mitochondria. Since it is well-established that consecutive prolines will result in ribosomal stalling on mRNA, we assumed that the “stalling” may decrease the overall elongation rate, which can be measured and quantified in vivo. In addition, we also intend to monitor the translation variation of Yeast after switching from normal glucose condition to an unfamiliar condition, such different carbon source provide or even nutrient depletion. We finally realized that the translation of Yeasts is going to be impeded after alteration of nutrient supply, then great adaptation to the external “stress” is shown by organisms.

While we have learned that mRNA sequence strongly influences translation and co-translational pathways, the exact mechanisms by which this sequence and the RNA structures encoded impact these steps of gene expression are less understood. Therefore, more investigations are needed to reveal how translation elongation and efficiency is influenced by RNA sequence and how the affected translation regulates and determines a variety of co-translational pathways. In this dissertation, I explore the effects of translational kinetics on a series of co-translational pathways using the budding yeast Saccharomyces cerevisiae as a model organism. In Chapter 2, I developed an in-vivo elongation reporter in Saccharomyces cerevisiae to quantitatively monitor translation elongation duration and protein expression. Using this elongation reporter, I investigated the effects of elongation stalls induced by different types of genetic factors on gene expression and demonstrated that distinct ribosomal stalls may trigger distinct co-translational pathways. In Chapter 3, I studied co-translational mRNA localization to mitochondrion, and proposed that translational kinetics, such as ribosomal stall caused by polyprolines, play an important role in mediating co-translational import. In addition, I further studied the effects of elongation stalls on mitochondrial import stress and triggering of relevant quality control pathways. In Chapter 4, I investigated the mechanism of cytosolic mRNP granule formation under glucose deprivation condition. In this study, I use CRISPRi to knockdown expression of RVB2 and confirm its essential role in deciding the fate of mRNA localization and translatability after glucose depletion. Finally in Chapter 5, I address the enhancements made to the developed method, outline potential future directions for the research presented in this dissertation, and conclude with my final remarks.