Poster abstracts
Poster number 26 submitted by Meredith Purchal
Deciphering mRNA Pseudouridylation in Eukaryotes
Meredith Purchal (Program in Chemical Biology, University of Michigan), Ryan McNassor (Department of Chemistry, University of Michigan ), Taslima Khan (Program in Chemical Biology, University of Michigan), Kristin Koutmou (Department of Chemistry, Program in Chemical Biology, University of Michigan), Markos Koutmos (Department of Chemistry, Department of Biophysics, Program in Chemical Biology, University of Michigan )
Abstract:
Pseudouridine (Y) is among the most abundant post-transcriptional RNA modifications in cells and has long been known to make important contributions to tRNA structure and function. While traditionally pseudouridine was only thought to occur in non-coding RNAs, recent efforts in mapping pseudouridylation across the transcriptome have identified a multitude of sites consistently pseudouridylated across mRNA and dynamically modified under stress conditions. As appreciation for the complexity of the role of this modification increases, a deeper understanding of the enzymes that catalyze the incorporation of this modification is necessary. Our goal is to describe the sequence and structural elements that cooperatively determine the specificity of pseudouridine synthase (Pus) enzymes towards their mRNA targets. We solved the first crystal structure of S. cerevisiae Pus7, a pseudouridine synthase that primarily modifies mRNA under cellular stress. Furthermore, we developed binding and kinetic assays to interrogate wild-type and mutant Pus7 pseudouridylation activity. Our biochemical work reveals several residues important to Pus7 function (K61, F307, F67, and D256). Ultimately, the combination of biophysical and biochemical techniques will allow us to dissect the features determining substrate recognition, and help to lay the foundation for a deeper understanding of RNA post-transcriptional regulation.
Keywords: Pseudouridylation, modification