title: Structural requirement of Ntc77 for spliceosome activation and first catalytic step.
abstract: The Prp19-associated complex is required for spliceosome activation by stabilizing the binding of U5 and U6 on the spliceosome after the release of U4. The complex comprises at least eight proteins, among which Ntc90 and Ntc77 contain multiple tetratricopeptide repeat (TPR) elements. We have previously shown that Ntc90 is not involved in spliceosome activation, but is required for the recruitment of essential first-step factor Yju2 to the spliceosome. We demonstrate here that Ntc77 has dual functions in both spliceosome activation and the first catalytic step in recruiting Yju2. We have identified an amino-terminal region of Ntc77, which encompasses the N-terminal domain and the first three TPR motifs, dispensable for spliceosome activation but required for stable interaction of Yju2 with the spliceosome. Deletion of this region had no severe effect on the integrity of the NTC, binding of NTC to the spliceosome or spliceosome activation, but impaired splicing and exhibited a dominant-negative growth phenotype. Our data reveal functional roles of Ntc77 in both spliceosome activation and the first catalytic step, and distinct structural domains of Ntc77 required for these two steps.
authors.affiliation: Institute of Molecular Biology, Academia Sinica, Nankang, Taipei, Taiwan 115, Republic of China Institute of Microbiology and Immunology, National Yang-Ming University, Shih-Pai, Taipei, Taiwan 112, Republic of China.
authors.forename: Hsin-Chou
authors.initials: HC
authors.lastname: Chen
authors.affiliation: Institute of Molecular Biology, Academia Sinica, Nankang, Taipei, Taiwan 115, Republic of China.
authors.forename: Kae-Jiun
authors.initials: KJ
authors.lastname: Chang
authors.affiliation: Institute of Molecular Biology, Academia Sinica, Nankang, Taipei, Taiwan 115, Republic of China.
authors.forename: Yu-Lun
authors.initials: YL
authors.lastname: Su
authors.affiliation: Institute of Molecular Biology, Academia Sinica, Nankang, Taipei, Taiwan 115, Republic of China.
authors.forename: Yu-Hsin
authors.initials: YH
authors.lastname: Huang
authors.affiliation: Institute of Molecular Biology, Academia Sinica, Nankang, Taipei, Taiwan 115, Republic of China mbscc@ccvax.sinica.edu.tw.
authors.forename: Soo-Chen
authors.initials: SC
authors.lastname: Cheng
mesh_terms: D055162:Biocatalysis; D018797:Cell Cycle Proteins / Q000737:chemistry* / Q000378:metabolism / Q000502:physiology; D053487:DEAD-box RNA Helicases / Q000378:metabolism; D009687:Nuclear Proteins / Q000378:metabolism; D017434:Protein Structure, Tertiary; D000072260:RNA Splicing Factors; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry* / Q000378:metabolism / Q000502:physiology; D017381:Spliceosomes / Q000378:metabolism*
chemical_list: C419763:CLF1 protein, S cerevisiae; D018797:Cell Cycle Proteins; D009687:Nuclear Proteins; C079877:PRP19 protein, S cerevisiae; D000072260:RNA Splicing Factors; D029701:Saccharomyces cerevisiae Proteins; C419764:Syf1 protein, S cerevisiae; C075469:YJU2 protein, S cerevisiae; C066490:PRP2 protein, S cerevisiae; D053487:DEAD-box RNA Helicases