The Prp19-associated Complex Is Required for Specifying Interactions of U5 and U6 with Pre-mRNA during Spliceosome Activation

• Published in: The Journal of biological chemistry 2005-09, Vol.280 (35), p.31190-31199
• Main Authors: Chan, Shih-Peng, Cheng, Soo-Chen
• Format: Article
• Language: English
• Subjects: Adenosine Triphosphate - metabolism; Base Pairing; Base Sequence; Molecular Sequence Data; Multiprotein Complexes; Nucleic Acid Conformation; Protein Binding; RNA Precursors - metabolism; RNA Splicing; RNA Splicing Factors; RNA, Small Nuclear - metabolism; Saccharomyces cerevisiae - genetics; Saccharomyces cerevisiae - metabolism; Saccharomyces cerevisiae Proteins - genetics; Saccharomyces cerevisiae Proteins - metabolism; Spliceosomes - metabolism
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.M505060200

Activation of the spliceosome involves a major structural change in the spliceosome, including release of U1 and U4 small nuclear ribonucleoprotein particles and the addition of a large protein complex, the Prp19-associated complex. We previously showed that the Prp19-associated complex is required for stable association of U5 and U6 with the spliceosome after U4 is released. Changes within the spliceosome upon binding of the Prp19-associated complex include remodeling of the U6/5′ splice site interaction and destabilization of Lsm proteins to allow further interaction of U6 with the intron sequence. Here, we further analyzed interactions of U5 and U6 with pre-mRNA at various stages of spliceosome assembly from initial binding of tri-small nuclear ribonucleoprotein complex to the activated spliceosome to reveal stepwise changes of interactions. We demonstrate that both U5 and U6 interacted with pre-mRNA in dynamic manners spanning over a large region of U6 and the 5′ exon sequences prior to the activation of the spliceosome. During spliceosome activation, interactions were locked down to small regions, and the Prp19-associated complex was required for defining the specificity of interaction of U5 and U6 with the 5′ splice site to stabilize their association with the spliceosome after U4 is dissociated.