Inhibition of msl-2 splicing by Sex-lethal reveals interaction between U2AF35 and the 3' splice site AG.
The protein Sex-lethal (SXL) controls dosage compensation in Drosophila by inhibiting the splicing and translation of male-specific-lethal-2 (msl-2) transcripts. Here we report that splicing inhibition of msl-2 requires a binding site for SXL at the polypyrimidine (poly(Y)) tract associated with the 3' splice site, and an unusually long distance between the poly(Y) tract and the conserved AG dinucleotide at the 3' end of the intron. Only this combination allows efficient blockage of U2 small nuclear ribonucleoprotein particle binding and displacement of the large subunit of the U2 auxiliary factor (U2AF65) from the poly(Y) tract by SXL. Crosslinking experiments with ultraviolet light indicate that the small subunit of U2AF (U2AF35) contacts the AG dinucleotide only when located in proximity to the poly(Y) tract. This interaction stabilizes U2AF65 binding such that SXL can no longer displace it from the poly(Y) tract. Our results reveal a novel function for U2AF35, a critical role for the 3' splice site AG at the earliest steps of spliceosome assembly and the need for a weakened U2AF35-AG interaction to regulate intron removal.
Merendino
L
L
Gene Expression Programme, European Molecular Biology Laboratory, Heidelberg, Germany.
Guth
S
S
Bilbao
D
D
Martínez
C
C
Valcárcel
J
J
D000227:Adenine Nucleotides / Q000378:metabolism; D001665:Binding Sites; D004268:DNA-Binding Proteins; D029721:Drosophila Proteins; D006150:Guanine Nucleotides / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D007438:Introns; D009687:Nuclear Proteins / Q000235:genetics*; D011485:Protein Binding; D012322:RNA Precursors / Q000378:metabolism; D012326:RNA Splicing; D016601:RNA-Binding Proteins / Q000378:metabolism*; D012261:Ribonucleoproteins / Q000378:metabolism*; D000072668:Splicing Factor U2AF; D014157:Transcription Factors / Q000235:genetics*
D000227:Adenine Nucleotides; D004268:DNA-Binding Proteins; D029721:Drosophila Proteins; D006150:Guanine Nucleotides; D009687:Nuclear Proteins; D012322:RNA Precursors; D016601:RNA-Binding Proteins; D012261:Ribonucleoproteins; D000072668:Splicing Factor U2AF; C067777:Sxl protein, Drosophila; D014157:Transcription Factors; C076753:U2AF1 protein, human; C000607494:U2AF2 protein, human; C094166:msl-2 protein, Drosophila