ATP-dependent interaction of yeast U5 snRNA loop 1 with the 5′ splice site

Rizwana K. Alvi, Mette Lund, Raymond T. O'Keefe

    Research output: Contribution to journalArticlepeer-review

    Abstract

    Pre-messenger RNA splicing is a two-step process by which introns are removed and exons joined together. In yeast, the U5 snRNA loop 1 interacts with the 5′ exon before the first step of splicing and with the 5′ and 3′ exons before the second step. In vitro studies revealed that yeast U5 loop 1 is not required for the first step of splicing but is essential for holding the 5′ and 3′ exons for ligation during the second step. It is critical, therefore, that loop 1 contacts the 5′ exon before the first step of splicing to hold this exon following cleavage from the pre-mRNA. At present it is not known how U5 loop 1 is positioned on the 5′ exon prior to the first step of splicing. To address this question, we have used site-specific photoactivated crosslinking in yeast spliceosomes to investigate the interaction of U5 loop 1 with the pre-mRNA prior to the first step of splicing. We have found that the highly conserved uridines in loop 1 make ATP-dependent contacts with an approximately 8-nt region at the 5′ splice site that includes the invariant GU. These interactions are dependent on functional U2 and U6 snRNAs. Our results support a model where U5 snRNA loop 1 interacts with the 5′ exon in two steps during its targeting to the 5′ splice site.
    Original languageEnglish
    Pages (from-to)1013-1023
    Number of pages10
    JournalRNA
    Volume7
    Issue number7
    DOIs
    Publication statusPublished - 2001

    Keywords

    • 5′
    • Pre-mRNA splicing
    • splice site
    • Spliceosome
    • U5 snRNA
    • Yeast

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