Cwc21p promotes the second step conformation of the spliceosome and modulates 3' splice site selection.
Pre-mRNA splicing involves two transesterification steps catalyzed by the spliceosome. How RNA substrates are positioned in each step and the molecular rearrangements involved, remain obscure. Here, we show that mutations in PRP16, PRP8, SNU114 and the U5 snRNA that affect this process interact genetically with CWC21, that encodes the yeast orthologue of the human SR protein, SRm300/SRRM2. Our microarray analysis shows changes in 3' splice site selection at elevated temperature in a subset of introns in cwc21Δ cells. Considering all the available data, we propose a role for Cwc21p positioning the 3' splice site at the transition to the second step conformation of the spliceosome, mediated through its interactions with the U5 snRNP. This suggests a mechanism whereby SRm300/SRRM2, might influence splice site selection in human cells.
Wellcome Trust Centre for Cell Biology, University of Edinburgh, King's Buildings, Mayfield Road, Edinburgh, EH9 3BF, UK.
Amit
A
Gautam
Wellcome Trust Centre for Cell Biology, University of Edinburgh, King's Buildings, Mayfield Road, Edinburgh, EH9 3BF, UK.
Richard J
RJ
Grainger
Department of Molecular Genomics, Institute of Molecular Biology of Barcelona (IBMB), 08028 Barcelona, Spain Institució Catalana de Recerca i Estudis Avançats (ICREA), 08010 Barcelona, Spain.
J
J
Vilardell
Wellcome Trust Centre for Cell Biology, University of Edinburgh, King's Buildings, Mayfield Road, Edinburgh, EH9 3BF, UK.
J David
JD
Barrass
Wellcome Trust Centre for Cell Biology, University of Edinburgh, King's Buildings, Mayfield Road, Edinburgh, EH9 3BF, UK jbeggs@ed.ac.uk.
Jean D
JD
Beggs
D000251:Adenosine Triphosphatases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D017398:Alternative Splicing; D000595:Amino Acid Sequence; D002352:Carrier Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D017353:Gene Deletion; D005800:Genes, Fungal; D006801:Humans; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D011487:Protein Conformation; D020365:RNA Helicases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012322:RNA Precursors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D022821:RNA Splice Sites; D012326:RNA Splicing; D000072260:RNA Splicing Factors; D012331:RNA, Fungal / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D016601:RNA-Binding Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D017414:Ribonucleoprotein, U4-U6 Small Nuclear / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D017415:Ribonucleoprotein, U5 Small Nuclear / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D017381:Spliceosomes / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
D002352:Carrier Proteins; C545658:Cwc21 protein, S cerevisiae; C118319:ISY1 protein, S cerevisiae; C077018:PRP8 protein, S cerevisiae; D012322:RNA Precursors; D022821:RNA Splice Sites; D000072260:RNA Splicing Factors; D012331:RNA, Fungal; D016601:RNA-Binding Proteins; D017414:Ribonucleoprotein, U4-U6 Small Nuclear; D017415:Ribonucleoprotein, U5 Small Nuclear; C414618:SRRM2 protein, human; D029701:Saccharomyces cerevisiae Proteins; D000251:Adenosine Triphosphatases; C075669:PRP16 protein, S cerevisiae; D020365:RNA Helicases