Phosphorylation of splicing factor SF1 on Ser20 by cGMP-dependent protein kinase regulates spliceosome assembly.
Splicing factor 1 (SF1) functions at early stages of pre-mRNA splicing and contributes to splice site recognition by interacting with the essential splicing factor U2AF65 and binding to the intron branch site. We have identified an 80 kDa substrate of cGMP-dependent protein kinase-I (PKG-I) isolated from rat brain, which is identical to SF1. PKG phosphorylates SF1 at Ser20, which inhibits the SF1-U2AF65 interaction leading to a block of pre-spliceosome assembly. Mutation of Ser20 to Ala or Thr also inhibits the interaction with U2AF65, indicating that Ser20 is essential for binding. SF1 is phosphorylated in vitro by PKG, but not by cAMP-dependent protein kinase A (PKA). Phosphorylation of SF1 also occurs in cultured neuronal cells and is increased on Ser20 in response to a cGMP analogue. These results suggest a new role for PKG in mammalian pre-mRNA splicing by regulating in a phosphorylation-dependent manner the association of SF1 with U2AF65 and spliceosome assembly.
Wang
X
X
Cell Signalling Unit, Children's Medical Research Institute, Locked Bag 23, Wentworthville 2145, NSW.
Bruderer
S
S
Rafi
Z
Z
Xue
J
J
Milburn
P J
PJ
Krämer
A
A
Robinson
P J
PJ
D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D001921:Brain / Q000378:metabolism; D002417:Cattle; D017869:Cyclic GMP-Dependent Protein Kinases / Q000378:metabolism*; D004268:DNA-Binding Proteins; D006801:Humans; D008969:Molecular Sequence Data; D009687:Nuclear Proteins; D010766:Phosphorylation; D011494:Protein Kinases / Q000378:metabolism; D012326:RNA Splicing; D000072260:RNA Splicing Factors; D016601:RNA-Binding Proteins / Q000378:metabolism*; D051381:Rats; D012261:Ribonucleoproteins / Q000378:metabolism*; D012694:Serine / Q000378:metabolism*; D017381:Spliceosomes / Q000378:metabolism*; D000072668:Splicing Factor U2AF; D013379:Substrate Specificity; D014157:Transcription Factors
D004268:DNA-Binding Proteins; D009687:Nuclear Proteins; D000072260:RNA Splicing Factors; D016601:RNA-Binding Proteins; D012261:Ribonucleoproteins; C107467:SF1 protein, human; C491513:Sf1 protein, rat; D000072668:Splicing Factor U2AF; D014157:Transcription Factors; C000607494:U2AF2 protein, human; D012694:Serine; D011494:Protein Kinases; D017869:Cyclic GMP-Dependent Protein Kinases