Functional links between the Prp19-associated complex, U4/U6 biogenesis, and spliceosome recycling.
The Prp19-associated complex, consisting of at least eight protein components, is involved in spliceosome activation by specifying the interaction of U5 and U6 with pre-mRNA for their stable association with the spliceosome after U4 dissociation. We show here that yeast cells depleted of one or two of the Prp19-associated components, accumulate the free form of U4. In NTC25-deleted cells, the level of U6 was also reduced. Extracts prepared from NTC25-deleted cells contained neither free U4 nor U6 and were ineffective in spliceosome recycling in the in vitro splicing reaction. Overexpression of U6 partially rescued the temperature-sensitive growth defect and decreased the relative amount of free U4 in NTC25-deleted cells, indicating that the accumulation of free U4 was a consequence of insufficient amounts of U6 snRNA. Extracts prepared from U6-overproducing NTC25-deleted cells containing free-form U6 were capable of spliceosome recycling, suggesting a role of free U6 RNP in spliceosome recycling. Our results demonstrate that in addition to direct participation in spliceosome activation, the Prp19-associated complex has an indirect role in spliceosome recycling through affecting the biogenesis of U4/U6 snRNP in the in vivo splicing reaction.
Institute of Molecular Biology, Academia Sinica, Nankang, Taiwan, Republic of China.
Chun-Hong
CH
Chen
Der-I
DI
Kao
Shih-Peng
SP
Chan
Tsu-Chung
TC
Kao
Jui-Yen
JY
Lin
Soo-Chen
SC
Cheng
D015152:Blotting, Northern; D015153:Blotting, Western; D004591:Electrophoresis, Polyacrylamide Gel; D017353:Gene Deletion; D005800:Genes, Fungal; D008954:Models, Biological; D010957:Plasmids / Q000235:genetics; D012326:RNA Splicing; D000072260:RNA Splicing Factors; D012333:RNA, Messenger / Q000032:analysis; D017414:Ribonucleoprotein, U4-U6 Small Nuclear / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000378:metabolism*; D017381:Spliceosomes / Q000737:chemistry / Q000378:metabolism
C079877:PRP19 protein, S cerevisiae; D000072260:RNA Splicing Factors; D012333:RNA, Messenger; D017414:Ribonucleoprotein, U4-U6 Small Nuclear; D029701:Saccharomyces cerevisiae Proteins