Characterization of splicing factors Yju2 and Cwc2 from Saccharomyces cerevisiae

• Main Authors: Yen-Chi Liu, 劉晏旗
• Other Authors: Soo-Chen Cheng
• Format: Others
• Language: zh-TW
• Published: 2007
• Online Access: http://ndltd.ncl.edu.tw/handle/50785680107854489584

博士 === 國立陽明大學 === 微生物及免疫學研究所 === 95 === 英文摘要 The Prp19-associated complex (NTC) is essential for pre-mRNA splicing and is associated with the spliceosome during spliceosome activation. NTC is required for specifying interactions of U5 and U6 with pre-mRNA to stabilize their association with the spliceosome after dissociation of U4. In this thesis, I study two splicing factors, Yju2 and Cwc2. Both proteins are associated with components of NTC, and are required for pre-mRNA splicing both in vivo and in vitro. During spliceosome assembly, Yju2 and Cwc2 are associated with the spliceosome at nearly the same time as NTC. However, Yju2 is destabilized after the first catalytic reaction, whereas Cwc2 and other NTC components remain associated until the reaction is complete. Extracts depleted of Yju2 or Cwc2 could be complemented by recombinant Yju2 or recombinant Cwc2, respectively, suggesting that Yju2 or Cwc2 is not entirely associated with NTC, and vice versa. Neither Yju2 nor Cwc2 is required for the binding of NTC to the spliceosome, while NTC is required for the binding of Yju2 or Cwc2 to the spliceosome. Further analysis reveals that Yju2 and NTC are functionally not linked. My study demonstrates that after Prp2-mediated structural rearrangement of the spliceosome, Yju2 acts in concert with unidentified heat-resistant factor(s) in an ATP-independent manner to promote the first catalytic reaction of pre-mRNA splicing. Moreover, Cwc2 is involved in NTC-mediated spliceosome activation through specifying interactions between U6 and the 5’ splice site, and the RNA recognition motif of Cwc2 might be important for its function.