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J Mol Biol. 2018 Feb 16;430(4):524-536. doi: 10.1016/j.jmb.2017.10.021. Epub 2017 Oct 24.

Stress-induced Pseudouridylation Alters the Structural Equilibrium of Yeast U2 snRNA Stem II.

Author information

1
Department of Biochemistry, University of Wisconsin-Madison, 433 Babcock Dr., Madison, WI 53706, USA.
2
Department of Biochemistry, University of Wisconsin-Madison, 433 Babcock Dr., Madison, WI 53706, USA. Electronic address: ahoskins@wisc.edu.

Abstract

In yeast, the U2 small nuclear ribonucleic acid (snRNA) component of the spliceosome is targeted for additional post-transcriptional modifications in response to cellular stress. Uridines 56 and 93 are both modified to pseudouridines (Ψ) during nutrient deprivation, while U56 is also pseudouridylated during heat shock. Both positions are located within stem II, which must toggle between two mutually exclusive structures during splicing. Stem IIa forms during spliceosome assembly, and stem IIc forms during the catalytic steps. We have studied how uridine 56 and 93 pseudouridylation impacts conformational switching of stem II. Using single-molecule Förster resonance energy transfer, we show that Ψ56 dampens conformational dynamics of stem II and stabilizes stem IIc. In contrast, Ψ93 increases dynamics of non-stem IIc conformations. Pseudouridylation impacts conformational switching of stem II by Mg2+ or the U2 protein Cus2; however, when Mg2+ and Cus2 are used in combination, the impacts of pseudouridylation can be suppressed. These results show that stress-induced post-transcriptional modification of U56 and U93 alters snRNA conformational dynamics by distinct mechanisms and that protein and metal cofactors of the spliceosome alter how snRNAs respond to these modifications.

KEYWORDS:

dynamics; pseudouridine; single-molecule FRET; snRNA; spliceosome

PMID:
29079482
PMCID:
PMC5805610
DOI:
10.1016/j.jmb.2017.10.021
[Indexed for MEDLINE]
Free PMC Article

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