Spliceosome disassembly catalyzed by Prp43 and its associated components Ntr1 and Ntr2

• Published in: Genes & development 2005-12, Vol.19 (24), p.2991-3003
• Main Authors: Tsai, Rong-Tzong, Fu, Ru-Huei, Yeh, Fu-Lung, Tseng, Chi-Kang, Lin, Yu-Chieh, Huang, Yu-Hsin, Cheng, Soo-Chen
• Format: Article
• Language: English
• Subjects: Adenosine Triphosphate - chemistry; Adenosine Triphosphate - metabolism; Cell-Free System - chemistry; Cell-Free System - metabolism; DEAD-box RNA Helicases; Research Papers; RNA Helicases - chemistry; RNA Helicases - isolation & purification; RNA Helicases - metabolism; RNA Splicing - physiology; RNA Splicing Factors; Saccharomyces cerevisiae - chemistry; Saccharomyces cerevisiae - metabolism; Saccharomyces cerevisiae Proteins - chemistry; Saccharomyces cerevisiae Proteins - isolation & purification; Saccharomyces cerevisiae Proteins - metabolism; Spliceosomes - chemistry; Spliceosomes - metabolism
• ISSN: 0890-9369; 1549-5477
• DOI: 10.1101/gad.1377405

Two novel yeast splicing factors required for spliceosome disassembly have been identified. Ntr1 and Ntr2 (NineTeen complex-Related proteins) were identified for their weak association with components of the Prp19-associated complex. Unlike other Prp19-associated components, these two proteins were primarily associated with the intron-containing spliceosome during the splicing reaction. Extracts depleted of Ntr1 or Ntr2 exhibited full splicing activity, but accumulated large amounts of lariat-intron in the spliceosome after splicing, indicating that the normal function of the Prp19-associated complex in spliceosome activation was not affected, but spliceosome disassembly was hindered. Immunoprecipitation analysis revealed that Ntr1 and Ntr2 formed a stable complex with DExD/H-box RNA helicase Prp43 in the splicing extract. Ntr1 interacted with Prp43 through the N-terminal G-patch domain, with Ntr2 through a middle region, and with itself through the carboxyl half of the protein. The affinity-purified Ntr1-Ntr2-Prp43 complex could catalyze disassembly of the spliceosome in an ATP-dependent manner, separating U2, U5, U6, NTC (NineTeen Complex), and lariat-intron. This is the first demonstration of physical disassembly of the spliceosome, catalyzed by a complex containing a DExD/H-box RNA helicase and two accessory factors, which might function in targeting the helicase to the correct substrate.