Identification and Characterization of Components of the Yeast Prp19p-associated Complex, Ntc85p and Ntc20p

• Main Authors: Tsai Wei-yu, 蔡維育
• Other Authors: Cheng Soo-Chen
• Format: Others
• Language: en_US
• Published: 1999
• Online Access: http://ndltd.ncl.edu.tw/handle/13855467277885641899

博士 === 國立陽明大學 === 微生物暨免疫學研究所 === 87 === The Prp19p protein of the budding yeast Saccharomyces cerevisiae is an essential splicing factor and is associated with the spliceosome during the splicing reaction. We have previously shown that Prp19p is not tightly associated with snRNPs, but is associated with a protein complex consisting of at least eight protein components. By sequencing components of the affinity purified complex, I have identified YBR188c and Cef1p as components of the Prp19p-associated complex, Ntc20p and Ntc85p (NTC represents nineteen complex), respectively. Ntc20p is a protein of 140 amino acid residues with no discernible motif. Ntc85p is a protein of 590 amino acid residues containing a c-Myb DNA binding motif in its amino-terminal region. The NTC85 gene is essential for vegetative yeast growth and the encoded protein is required for pre-mRNA splicing both in vivo and in vitro. Ntc85p is highly homologous to the fission yeast S. pombe Cdc5p with 48% identity in the amino-terminus over a region of 267 amino acid residues. Deletion of any sequence in this region resulted in cell lethality. Ntc85p interacts with Prp19p and Ntc20p both in vivo and in vitro. Both Ntc85p and Ntc20p are associated with the spliceosome in the same manner as Prp19p, i.e., concomitantly with or immediately after dissociation of U4 snRNP. The anti-Ntc85p antibody inhibited binding to the spliceosome of Ntc85p, Prp19p and at least three other components of the Prp19p-associated complex, suggesting that Ntc85p may play a role in promoting binding of the Prp19p-associated complex to the spliceosome. Dissociation of U4 snRNP, however, is not affected by the antibody. These results suggest that U4 snRNP dissociation is independent of binding of the Prp19p-associated complex to the spliceosome, and the Prp19p-associated complex may play a role in mediating events after U4 snRNP dissociation to form the active spliceosome.