Characterization of interactions among the Cef1p-Prp19p-associated splicing complex.
Schizosaccharomyces pombe (Sp) Cdc5p and its Saccharomyces cerevisiae (Sc) ortholog, Cef1p, are essential components of the spliceosome. In S. cerevisiae, a subcomplex of the spliceosome that includes Cef1p can be isolated on its own; this has been termed the nineteen complex (Ntc) because it contains Prp19p. Components of the Ntc include Cef1p, Snt309p, Syf2p/Ntc31p, Ntc30p/lsy1p, Ntc20p and at least six unidentified proteins. We recently identified approximately 30 proteins that copurified with Cdc5p and Cef1p. Previously unidentified S. pombe proteins in this purification were called Cwfs for complexed with five and novel S. cerevisiae proteins were called Cwcs for complexed with Cef1p. Using these proteomics data coupled with available information regarding Ntc composition, we have investigated protein identities and interactions among Ntc components. Our data indicate that Cwc2p, Prp46p, Clf1p, and Syf1p most likely represent Ntc40p, Ntc50p, Ntc77p, and Ntc90p, respectively. We show that Sc Cwc2p interacts with Prp19p and is involved in pre-mRNA splicing. Sp cwf2+, the homolog of Sc CWC2, is allelic with the previously identified Sp prp3+. We present evidence that Sp Cwf7p, an essential protein with obvious homologs in many eukaryotes but not S. cerevisiae, is a functional counterpart of Sc Snt309p and binds Sp Cwf8p (a homolog of Sc Prp19p). Further, our data indicate that a mutation in the U-box of Prp19p disrupts these numerous protein interactions causing Cef1p degradation and Ntc instability.
Ohi
Melanie D
MD
Howard Hughes Medical Institute and Department of Cell Biology, Vanderbilt University School of Medicine, Nashville, Tennessee 37232, USA.
Gould
Kathleen L
KL
D000595:Amino Acid Sequence; D001483:Base Sequence; D018797:Cell Cycle Proteins / Q000235:genetics / Q000378:metabolism*; D017931:DNA Primers; D005656:Fungal Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D012326:RNA Splicing; D000072260:RNA Splicing Factors; D016601:RNA-Binding Proteins; D012441:Saccharomyces cerevisiae / Q000235:genetics*; D029701:Saccharomyces cerevisiae Proteins; D029702:Schizosaccharomyces pombe Proteins; D017386:Sequence Homology, Amino Acid; D017381:Spliceosomes; D020798:Two-Hybrid System Techniques
C513239:CEF1 protein, S cerevisiae; D018797:Cell Cycle Proteins; D017931:DNA Primers; D005656:Fungal Proteins; C079877:PRP19 protein, S cerevisiae; D000072260:RNA Splicing Factors; D016601:RNA-Binding Proteins; D029701:Saccharomyces cerevisiae Proteins; D029702:Schizosaccharomyces pombe Proteins; C085875:cdc5 protein, S pombe