The Prp19-associated complex is required for specifying interactions of U5 and U6 with pre-mRNA during spliceosome activation.
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.
Institute of Molecular Biology, Academia Sinica, Nankang 115, Taiwan, Republic of China.
Shih-Peng
SP
Chan
Soo-Chen
SC
Cheng
D000255:Adenosine Triphosphate / Q000378:metabolism; D020029:Base Pairing; D001483:Base Sequence; D008969:Molecular Sequence Data; D046912:Multiprotein Complexes; D009690:Nucleic Acid Conformation; D011485:Protein Binding; D012322:RNA Precursors / Q000378:metabolism*; D012326:RNA Splicing; D000072260:RNA Splicing Factors; D012342:RNA, Small Nuclear / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000235:genetics* / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism*; D017381:Spliceosomes / Q000378:metabolism*
D046912:Multiprotein Complexes; C079877:PRP19 protein, S cerevisiae; D012322:RNA Precursors; D000072260:RNA Splicing Factors; D012342:RNA, Small Nuclear; D029701:Saccharomyces cerevisiae Proteins; C405485:U5 small nuclear RNA; C405486:U6 small nuclear RNA; D000255:Adenosine Triphosphate