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tRNA homeostasis and gene regulation

Final Report Summary - TRNA DYNAMICS (tRNA homeostasis and gene regulation)

Translation of mRNA into proteins is a central process in gene expression. The different genes within a genome, depending on their sequence, might have varying efficiency in using the cellular resources devoted for protein synthesis. In particular, codons that are recognized by abundant tRNAs are expected to be translated more efficiently and reduce the energetic cost associated with proofreading and rejection of non-cognate tRNAs.
The simplest models assume that the tRNA pool remains constant throughout the life of a cell. However, recent reports have shown that the abundance of the total tRNA pool can vary during the cell cycle and this observation is evolutionary conserved. Demonstrating that the tRNA pool in the cell is adaptable will add a new regulatory level with important implications in cell biology. Therefore, uncover the molecular features and signaling pathways on tRNA that are responsible for tRNA regulation is of fundamental importance to understand the cell.
Our work aims to investigate whether the processes that regulate the tRNA availability in the cell are specific and how they can affect protein translation and directly impact cell fitness. Specifically, we aimed to answer:
(i) How the tRNA abundance landscape responds to different stimuli?
(ii) Which processes (synthesis, transport and degradation) are actively involved in tRNA dynamics
and are they specific for individual tRNAs?
(iii) Does tRNA dynamics affect translation and influence cell fitness?

This work aimed to uncover new regulatory mechanisms in the cell that may have broad impact in the understanding of how protein synthesis is regulated. Demonstrating that the tRNA pool in the cell is adaptable will add a new regulatory level with important implications in cell biology. Therefore, uncover the molecular features and signaling pathways on tRNA that are responsible for tRNA regulation is of fundamental importance to understand the cell. The results of this project will also provide an extensive knowledge about tRNA regulation during stress conditions. Since no experimental work has been done to systematically characterize the amounts of individual tRNAs at different conditions, the techniques developed during this project will provide other researchers with the tools needed to investigate the influence of tRNA abundance in multiple cellular processes.

Our results show that individual tRNAs change during stress conditions in yeast in a time dependent manner. As tRNAs are used as building blocks to synthesize new proteins, these changes in tRNA abundance have an impact in protein synthesis, both in initiation and elongation. By using GFP reporters, we confirmed that tRNA changes impact to protein synthesis and thereby should be considered as a new regulatory layer in the cell.
We determined that tRNA abundance changes during stress conditions are mainly regulated by tRNA degradation and we identified RNY1, a stress-related ribonuclease, as a key player in the specific regulation of tRNAs during stress conditions.
Finally, we also identified tRNA shuttling between nucleus and cytoplasm as a new player in regulating tRNA abundance. We have determined that active shuttling takes place during stress and is dependent on the stress condition.
All these results support the idea that tRNA abundance is tightly controlled in the cell and this is reflected in a regulation of the protein synthesis in the ribosome. We believe that our results introduce a new layer of regulation operating in the context of the central dogma.