Date of Award
Doctor of Philosophy (PhD)
Ribosome often faces defective adducts that disrupt its movement along the mRNA template. These adducts are primarily caused by chemical damage to mRNA and are highly detrimental to the decoding process on the ribosome. Hence, unless dealt with, chemical damage to RNA has been hypothesized to lead to the production of toxic protein products. Even more detrimental is the ability of damaged mRNA to drastically affect ribosome homeostasis through stalling. This in turn would lead to greatly diminished translation capacity of cells. Therefore, the inability of cells to recognize and resolve translational-stalling events is detrimental to proteostasis and could even lead to disease development. No-go decay (NGD) is a eukaryotic quality control mechanism that evolved to cope with translational arrests. The process is characterized by an endonucleolytic cleavage near the stall sequence, but the mechanistic details are unclear. It was, however, unclear whether organisms evolved NGD to deal with RNA damage. In the work presented in this thesis, I showed that NGD and the associated ribosome-quality control (RQC) are activated in response to nucleobase alkylation and oxidation. Our findings reveal that these processes are important for clearing chemically modified mRNA and the resulting aberrant-protein products and highlighted the burden of chemically damaged mRNA on cellular homeostasis and suggest that organisms evolved mechanisms to counter their accumulation. In addition to activating RQC, stalling has been recently linked to the activation of the integrated-stress response (ISR) by Gcn2, suggesting that the activation of the two processes is likely to be coordinated. To investigate this potential coordination, I performed genetic and biochemical analysis in yeast. My data revealed that activation of RQC by Hel2 suppresses that of Gcn2. I further showed that Hel2 and Gcn2 are activated by a similar set of agents that cause ribosome stalling, with maximal activation of Hel2 observed at a lower frequency of stalling. Interestingly, inactivation of one pathway was found to result in the overactivation of the other, suggesting that both are activated by the same signal of ribosome collisions. Notably, the processes do not appear to be in direct competition with each other; ISR prefers a vacant A site, whereas RQC displays no preference. Collectively, our findings provide important details about how multiple pathways that recognize stalled ribosomes coordinate to mount the appropriate response.
Chair and Committee
Hani H. Zaher
Joe J. Jez, Nima N. Mosammaparast, True T. Heather, Taylor T. John-Stephen,
Yan, Liewei, "Chemical Damage to mRNA and Its Impact on Ribosome Quality-Control and Stress-Response Pathways in Eukaryotic Cells" (2021). Arts & Sciences Electronic Theses and Dissertations. 2546.