Poster abstracts
Poster number 40 submitted by Ernesto Roldan-Bonet
Understanding Transfer RNA Nuclear/Cytoplasmic Transport and Its Role in Quality Control
Ernesto J. Roldan-Bonet (Department of Molecular Genetics, Center for RNA Biology, OSU), Paolo L. Sinopoli (Department of Molecular Genetics, Center for RNA Biology, OSU), Anita K. Hopper (Department of Molecular Genetics, Center for RNA Biology, OSU)
Abstract:
Transfer RNAs (tRNAs) are essential adapter molecules that function in translation. Long believed to move unidirectionally from the nucleus to the cytoplasm in eukaryotic cells, it is now well established that cytoplasmic tRNAs undergo constitutive retrograde nuclear import and re-export. This widely conserved nuclear retrograde trafficking pathway has been implicated in many important cellular processes including the addition of select post-transcriptional modifications to tRNAs and regulation of translation under stress conditions. Our work demonstrates that disruption of tRNA nuclear import leads to the accumulation of defective cytoplasmic tRNAs, such as those lacking the post-transcriptional modification m22G26 or bearing unprocessed 5′ leader sequences, implicating nuclear import in the quality control of tRNAs. However, the fate of aberrant tRNAs following nuclear import, as well as the molecular machinery mediating import, remain largely undefined. To address these questions, we are employing various genetic approaches. Nuclear turnover will be assessed by measuring hypomodified tRNA levels in strains lacking nuclear turnover components, while nuclear repair will be investigated using yeast heterokaryons. To identify import factors, we developed a genome-wide screen in Saccharomyces cerevisiae, leveraging the nuclear retrograde pathway dependent wybutosine (yW) modification of tRNAPhe. Screening over 15% of the genome, we have identified proteins known to be involved in yW biosynthesis, validating our assay, and identified additional candidates whose roles in import versus yW biosynthesis are under investigation. Together, these studies will advance our understanding of tRNA nuclear trafficking and its involvement tRNA quality control.
References:
1) Kramer, E. B., & Hopper, A. K. (2013). Retrograde transfer RNA nuclear import provides a new level of tRNA quality control in Saccharomyces cerevisiae. Proceedings of the National Academy of Sciences, 110(52), 21042–21047. https://doi.org/10.1073/pnas.1316579110
2) Nostramo, R. T., & Hopper, A. K. (2020). A novel assay provides insight into tRNAPhe retrograde nuclear import and re-export in S. cerevisiae. Nucleic Acids Research, 48(20), 11577–11588. https://doi.org/10.1093/nar/gkaa879
3) Phizicky, E. M., & Hopper, A. K. (2023). The life and times of a tRNA. https://doi.org/10.1261/rna
4) Shaheen, H. H., & Hopper, A. K. (2005). Retrograde movement of tRNAs from the cytoplasm to the nucleus in Saccharomyces cerevisiae. Proceedings of the National Academy of Sciences, 102(32), 11290–11295. https://doi.org/10.1073/pnas.0503836102
Keywords: RNA Quality Control, Retrograde Nuclear Transport, Wybutosine