Poster abstracts
Poster number 41 submitted by Chloe Reavill
Exploring the relationship between tRNA quality control and the Trm10-mediated N1-methylation of tRNA in S. cerevisiae
Chloe A. Reavill (The Ohio State University), Isobel E. Bowles (The Ohio State University), Jane E. Jackman (The Ohio State University)
Abstract:
tRNA methyltransferase 10 (Trm10) performs the m1R9 modification on select tRNA substrates throughout Eukarya and in some archaea. In Saccharomyces cerevisiae, deletion of TRM10 does not cause a growth defect under standard conditions, but these mutants exhibit growth sensitivity when exposed to the commonly used chemotherapeutic 5-fluorouracil (5FU). This drug inhibits thymidylate synthase, resulting in apoptosis of cells, but can also be incorporated into RNA in the form of FUMP, which has been shown to negatively affect RNA function. The growth sensitivity of the trm10Δ strain arises due to selective depletion of the Trm10 substrate tRNATrp. Under standard conditions, the trm10Δ strain exhibits a decrease in tRNATrp abundance, which is exacerbated with 5FU treatment. The mechanism responsible for targeting hypomodified tRNATrp for degradation has not yet been fully elucidated. However, 5-fluorouracil sensitivity is rescued in the trm10Δ met22Δ strain. Met22 functions in methionine biosynthesis, and to perform its role, the enzyme utilizes adenosine 3′,5′-bisphosphate (pAp). When Met22 is deleted, pAp accumulates and inhibits nucleases involved in the rapid tRNA decay (RTD) quality control pathway. However, the canonical RTD nucleases that are targets of met22-dependent inhibition, Xrn1 and Rat1, are not acting on tRNATrp, suggesting a novel Met22-dependent pathway is responsible. We will perform a suppressor screen to determine the tRNA quality control mechanism acting in the trm10Δ strain. We will also use tRNA structure-seq to analyze the structure and abundance of tRNAs in the wildtype and trm10Δ S. cerevisiae strains (with and without 5FU) to determine how the m1G9 modification affects tRNA stability. Structural changes observed by comparing the WT and trm10Δ strains may reveal the mechanism in which certain tRNAs exhibit increased susceptibility to nuclease degradation. This work will reveal insight into how a singular tRNA body modification is important in maintaining structure and function, which is especially critical under stress conditions.
References:
1. Bowles IE, Jackman JE. A tRNA-specific function for tRNA methyltransferase Trm10 is associated with a new tRNA quality control mechanism in Saccharomyces cerevisiae. RNA. 2024;30(2):171-187. doi: 10.1261/rna.079861.123.
2. Chernyakov et al. Degradation of several hypomodified mature tRNA species in Saccharomyces cerevisiae is mediated by Met22 and the 5’-3’ exonucleases Rat1 and Xrn1. Genes Dev. 2008;22(10):1369-1380. doi:10.1101/gad.1654308
3. Hoskins J, Butler JS. RNA-based 5-fluorouracil toxicity requires the pseudouridylation activity of Cbf5p. Genetics. 2008;179(1):323-330. doi:10.1534/genetics.107.082727
4. Yamagami et al. Genome-wide analysis of the in vivo tRNA structurome reveals RNA structural and modification dynamics under heat stress. Proc Natl Acad Sci. 2022;119(25):e2201237119. doi:10.1073/pnas.2201237119
Keywords: TRM10, 5-fluorouracil, RTD