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Nucleic Acids Research Advance Access originally published online on December 15, 2007
Nucleic Acids Research 2008 36(3):814-825; doi:10.1093/nar/gkm1098
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Nucleic Acids Research, 2008, Vol. 36, No. 3 814-825
© 2007 The Author(s)
This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/2.0/uk/) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.

RNA

The U1, U2 and U5 snRNAs crosslink to the 5' exon during yeast pre-mRNA splicing

Joanne C. McGrail and Raymond T. O’Keefe*

Faculty of Life Sciences, The University of Manchester, Manchester, M13 9PT, UK

*To whom correspondence should be addressed. Tel: +44 0161 275 7393; Fax: +44 0161 275 5082; Email: rokeefe{at}manchester.ac.uk

Received October 3, 2007. Revised November 26, 2007. Accepted November 26, 2007.

Activation of pre-messenger RNA (pre-mRNA) splicing requires 5' splice site recognition by U1 small nuclear RNA (snRNA), which is replaced by U5 and U6 snRNA. Here we use crosslinking to investigate snRNA interactions with the 5' exon adjacent to the 5' splice site, prior to the first step of splicing. U1 snRNA was found to interact with four different 5' exon positions using one specific sequence adjacent to U1 snRNA helix 1. This novel interaction of U1 we propose occurs before U1-5' splice site base pairing. In contrast, U5 snRNA interactions with the 5' exon of the pre-mRNA progressively shift towards the 5' end of U5 loop 1 as the crosslinking group is placed further from the 5' splice site, with only interactions closest to the 5' splice site persisting to the 5' exon intermediate and the second step of splicing. A novel yeast U2 snRNA interaction with the 5' exon was also identified, which is ATP dependent and requires U2-branchpoint interaction. This study provides insight into the nature and timing of snRNA interactions required for 5' splice site recognition prior to the first step of pre-mRNA splicing.


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