Poster abstracts
Poster number 63 submitted by Minli Ruan
Determining the Influence of PUS7 localization on RNA Substrate Selection
Minli Ruan (Biological Chemistry, University of Michigan), Daniel E. Eyler (Chemical Biology, University of Michigan), Kristin S. Koutmou (Chemical Biology, University of Michigan)
Abstract:
Pseudouridine (Ψ), one of the most abundant post-transcriptional RNA modifications in cells, broadly affects RNA structure and function. The insertion of Ψ into RNAs (tRNA, snRNA, rRNA, mRNA) is catalyzed by Pseudouridine Synthase (Pus) enzymes, which can be categorized into six families: TruA, TruB, RluA, RsuA, TruD, and Pus10. Pus enzymes are important for gene expression, as the misregulation of Ψ incorporation is associated with a variety of human diseases and cancers.1-3 For example, mutants of the TruD family member, Pus7, are linked with several inherited diseases, including speech delay, intellectual disability, and microcephaly. Recently we have begun to gain insight onto the dynamic regulation of Pus7, but how it selects its specific substrates is still poorly understood. Elucidating the mechanism of Pus7 substrate selection is important for understanding pathologies linked to Pus7 dysfunction. Pus7 recognizes RNA substrates that possess a consensus UGUAR (R = A/G) sequence. However, transcriptome-wide sequencing demonstrates that < 3% of RNAs containing this consensus sequence are pseudouridylated in cells under normal-growth conditions. However, the fraction of UGUAR sequences modified under heat shock conditions, where Pus7 relocaliezes from the nucleus into the cytoplasm, increases to more than 20%. These data indicate that the consensus sequence alone cannot explain Pus7 substrate selection and further mechanistic investigations, such as enzyme relocalization, are warranted. My work investigates the possibility that Pus7 localization impacts RNA substrate selection to influence protein synthesis in response to specific cellular stresses. I am developing Pus7 constructs capable of being localized in either the nucleus or cytoplasm and by mapping pseudouridylation dynamic changes together with translation efficiency changes in yeast to determine this molecular mechanism.
References:
1. Eyler, D. et al. (2019). Proceedings Of The National Academy Of Sciences, 116(46), 23068-23074.
2. Carlile, T. et al.(2014). Nature, 515(7525), 143-146.
3. Garcia, D. et al. (2021). Elife, 10.
4. Purchal, M. et al. (2022). Proceedings Of The National Academy Of Sciences, 119(4).
5. de Brouwer, A. et al. (2018). The American Journal Of Human Genetics, 103(6), 1045-1052.
6. Carlile, T. et al.(2019). Nature Chemical Biology, 15(10), 966-974.
7. Schwartz, S. et al. (2014). Cell, 159(1), 148-162.
Keywords: Pseudouridine Synthase 7 (Pus7), Pseudouridine, Localization