Interactions of the yeast SF3b splicing factor.
The U2 snRNP promotes prespliceosome assembly through interactions that minimally involve the branchpoint binding protein, Mud2p, and the pre-mRNA. We previously showed that seven proteins copurify with the yeast (Saccharomyces cerevisiae) SF3b U2 subcomplex that associates with the pre-mRNA branchpoint region: Rse1p, Hsh155p, Hsh49p, Cus1p, and Rds3p and unidentified subunits p10 and p17. Here proteomic and genetic studies identify Rcp10p as p10 and show that it contributes to SF3b stability and is necessary for normal cellular Cus1p accumulation and for U2 snRNP recruitment in splicing. Remarkably, only the final 53 amino acids of Rcp10p are essential. p17 is shown to be composed of two accessory splicing factors, Bud31p and Ist3p, the latter of which independently associates with the RES complex implicated in the nuclear pre-mRNA retention. A directed two-hybrid screen reveals a network of prospective interactions that includes previously unreported intra-SF3b contacts and SF3b interactions with the RES subunit Bud13p, the Prp5p DExD/H-box protein, Mud2p, and the late-acting nineteen complex. These data establish the concordance of yeast and mammalian SF3b complexes, implicate accessory splicing factors in U2 snRNP function, and support SF3b contribution from early pre-mRNP recognition to late steps in splicing.
Wang
Qiang
Q
Department of Biology, University of Kentucky, Lexington, 40506, USA.
He
Jin
J
Lynn
Bert
B
Rymond
Brian C
BC
D000595:Amino Acid Sequence; D008969:Molecular Sequence Data; D040901:Proteomics; D012326:RNA Splicing; D012331:RNA, Fungal / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D016601:RNA-Binding Proteins / Q000378:metabolism; D017413:Ribonucleoprotein, U2 Small Nuclear / Q000235:genetics / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000235:genetics* / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism*; D017381:Spliceosomes / Q000378:metabolism; D020798:Two-Hybrid System Techniques
C489248:BUD31 protein, S cerevisiae; C097976:CUS1 protein, S cerevisiae; C425663:IST3 protein, S cerevisiae; D012331:RNA, Fungal; D012333:RNA, Messenger; D016601:RNA-Binding Proteins; D017413:Ribonucleoprotein, U2 Small Nuclear; D029701:Saccharomyces cerevisiae Proteins; C507323:YSF3 protein, S cerevisiae