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Systematic two-hybrid and comparative proteomic analyses reveal novel yeast pre-mRNA splicing factors connected to Prp19.

PloS one | Feb 28, 2011

Prp19 is the founding member of the NineTeen Complex, or NTC, which is a spliceosomal subcomplex essential for spliceosome activation. To define Prp19 connectivity and dynamic protein interactions within the spliceosome, we systematically queried the Saccharomyces cerevisiae proteome for Prp19 WD40 domain interaction partners by two-hybrid analysis. We report that in addition to S. cerevisiae Cwc2, the splicing factor Prp17 binds directly to the Prp19 WD40 domain in a 1:1 ratio. Prp17 binds simultaneously with Cwc2 indicating that it is part of the core NTC complex. We also find that the previously uncharacterized protein Urn1 (Dre4 in Schizosaccharomyces pombe) directly interacts with Prp19, and that Dre4 is conditionally required for pre-mRNA splicing in S. pombe. S. pombe Dre4 and S. cerevisiae Urn1 co-purify U2, U5, and U6 snRNAs and multiple splicing factors, and dre4Δ and urn1Δ strains display numerous negative genetic interactions with known splicing mutants. The S. pombe Prp19-containing Dre4 complex co-purifies three previously uncharacterized proteins that participate in pre-mRNA splicing, likely before spliceosome activation. Our multi-faceted approach has revealed new low abundance splicing factors connected to NTC function, provides evidence for distinct Prp19 containing complexes, and underscores the role of the Prp19 WD40 domain as a splicing scaffold.

Pubmed ID: 21386897 RIS Download

Mesh terms: Models, Biological | Organisms, Genetically Modified | Protein Binding | Protein Structure, Tertiary | Proteomics | RNA Precursors | RNA Splicing | RNA Splicing Factors | Saccharomyces cerevisiae | Saccharomyces cerevisiae Proteins | Schizosaccharomyces | Spliceosomes | Transcription Factors | Two-Hybrid System Techniques | Yeasts

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Associated grants

  • Agency: NCRR NIH HHS, Id: P20RR020171
  • Agency: NCRR NIH HHS, Id: P20 RR020171
  • Agency: NCI NIH HHS, Id: T32 CA119925
  • Agency: NCI NIH HHS, Id: T32CA119925
  • Agency: NCRR NIH HHS, Id: P41 RR011823
  • Agency: NCRR NIH HHS, Id: P41 RR11823
  • Agency: NIH HHS, Id: 1DP2OD004483
  • Agency: Howard Hughes Medical Institute, Id: DP2 OD004483
  • Agency: NIH HHS, Id:

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SGD

A curated database that provides comprehensive integrated biological information for Saccharomyces cerevisiae along with search and analysis tools to explore these data. SGD allows researchers to discover functional relationships between sequence and gene products in fungi and higher organisms. The SGD also maintains the S. cerevisiae Gene Name Registry, a complete list of all gene names used in S. cerevisiae which includes a set of general guidelines to gene naming. Protein Page provides basic protein information calculated from the predicted sequence and contains links to a variety of secondary structure and tertiary structure resources. Yeast Biochemical Pathways allows users to view and search for biochemical reactions and pathways that occur in S. cerevisiae as well as map expression data onto the biochemical pathways. Literature citations are provided where available.

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Pompep

FTP site to access Schizosaccharomyces pombe protein data.

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