Prp8 positioning of U5 snRNA is linked to 5′ splice site recognition

  1. Melissa S. Jurica1,2
  1. 1Department of Molecular Cell and Developmental Biology, University of California, Santa Cruz, California 95064, USA
  2. 2Center for Molecular Biology of RNA, University of California, Santa Cruz, California 95064, USA
  3. 3Department of Biochemistry and Biophysics, University of California, San Francisco, California 94143, USA
  4. 4Department of Pharmaceutical Chemistry, University of California, San Francisco, California 94122, USA
  1. Corresponding author: mjurica{at}ucsc.edu

Abstract

Prp8 is an essential protein that regulates spliceosome assembly and conformation during pre-mRNA splicing. Recent cryo-EM structures of the spliceosome model Prp8 as a scaffold for the spliceosome's catalytic U snRNA components. Using a new amino acid probing strategy, we identified a dynamic region in human Prp8 that is positioned to stabilize the pre-mRNA in the spliceosome active site through interactions with U5 snRNA. Mutagenesis of the identified Prp8 residues in yeast indicates a role in 5′ splice site recognition. Genetic interactions with spliceosome proteins Isy1, which buttresses the intron branch point, and Snu114, a regulatory GTPase that directly contacts Prp8, further corroborate a role for the same Prp8 residues in substrate positioning and activation. Together the data suggest that adjustments in interactions between Prp8 and U5 snRNA help establish proper positioning of the pre-mRNA into the active site to enhance 5′ splice site fidelity.

Keywords

  • Received December 24, 2017.
  • Accepted February 26, 2018.

This article is distributed exclusively by the RNA Society for the first 12 months after the full-issue publication date (see http://rnajournal.cshlp.org/site/misc/terms.xhtml). After 12 months, it is available under a Creative Commons License (Attribution-NonCommercial 4.0 International), as described at http://creativecommons.org/licenses/by-nc/4.0/.

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  1. Published in Advance February 27, 2018, doi: 10.1261/rna.065458.117 RNA 24: 769-777 © 2018 MacRae et al.; Published by Cold Spring Harbor Laboratory Press for the RNA Society

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