Suppressors of a U4 snRNA mutation define a novel U6 snRNP protein with RNA-binding motifs.
U4 and U6 small nuclear RNAs are associated by an extensive base-pairing interaction that must be disrupted and reformed with each round of splicing. U4 mutations within the U4/U6 interaction domain destabilize the complex in vitro and cause a cold-sensitive phenotype in vivo. Restabilization of the U4/U6 helix by dominant (gain-of-function), compensatory mutations in U6 results in wild-type growth. Cold-insensitive growth can also be restored by two classes of recessive (loss-of-function) suppressors: (1) mutations in PRP24, which we show to be a U6-specific binding protein of the RNP-consensus family; and (2) mutations in U6, which lie outside the interaction domain and identify putative PRP24-binding sites. Destabilization of the U4/U6 helix causes the accumulation of a PRP24/U4/U6 complex, which is undetectable in wild-type cells. The loss-of-function suppressor mutations inhibit the binding of PRP24 to U6, and thus presumably promote the release of PRP24 from the PRP24/U4/U6 complex and the reformation of the base-paired U4/U6 snRNP. We propose that the PRP24/U4/U6 complex is normally a highly transient intermediate in the spliceosome cycle and that PRP24 promotes the reannealing of U6 with U4.
Shannon
K W
KW
Department of Biochemistry and Biophysics, University of California, San Francisco 94143.
Guthrie
C
C
D000595:Amino Acid Sequence; D001483:Base Sequence; D002352:Carrier Proteins / Q000235:genetics* / Q000378:metabolism; D016384:Consensus Sequence; D005656:Fungal Proteins / Q000235:genetics / Q000378:metabolism; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D010641:Phenotype; D011233:Precipitin Tests; D012326:RNA Splicing; D012331:RNA, Fungal / Q000235:genetics* / Q000378:metabolism; D012342:RNA, Small Nuclear / Q000235:genetics*; D016601:RNA-Binding Proteins; D012261:Ribonucleoproteins / Q000235:genetics* / Q000378:metabolism; D017411:Ribonucleoproteins, Small Nuclear; D012441:Saccharomyces cerevisiae / Q000235:genetics* / Q000254:growth & development; D029701:Saccharomyces cerevisiae Proteins; D016415:Sequence Alignment; D013489:Suppression, Genetic
D002352:Carrier Proteins; D005656:Fungal Proteins; C401305:Prp24 protein, S cerevisiae; D012331:RNA, Fungal; D012342:RNA, Small Nuclear; D016601:RNA-Binding Proteins; D012261:Ribonucleoproteins; D017411:Ribonucleoproteins, Small Nuclear; D029701:Saccharomyces cerevisiae Proteins