title: Splicing factors SF1 and U2AF associate in extraspliceosomal complexes.
abstract: Splicing factors SF1 and U2AF associate cooperatively with pre-mRNA and play a crucial role in 3' splice site recognition during early steps of spliceosome assembly. Formation of the active spliceosome subsequently displaces SF1 in a remodeling process that stabilizes the association of U2 snRNP with pre-mRNA. Fluorescence microscopy shows SF1 and U2AF distributed throughout the nucleoplasm, where transcription occurs, with additional concentration in nuclear speckles, where splicing factors accumulate when not engaged in splicing. Fluorescence recovery after photobleaching analysis in live cells shows that the mobilities of SF1 and the two subunits of U2AF (U2AF(65) and U2AF(35)) are correlated with the abilities of these proteins to interact with each other. Direct binding of SF1 to U2AF(65) was demonstrated by fluorescence resonance energy transfer in both the nucleoplasm and nuclear speckles. This interaction persisted after transcription inhibition, suggesting that SF1 associates with U2AF in a splicing-independent manner. We propose that SF1 and U2AF form extraspliceosomal complexes before and after taking part in the assembly of catalytic spliceosomes.
authors.affiliation: Instituto de Medicina Molecular, Faculdade de Medicina, Av. Prof. Egas Moniz, 1649-028 Lisboa, Portugal.
authors.forename: José
authors.initials: J
authors.lastname: Rino
authors.forename: Joana M P
authors.initials: JM
authors.lastname: Desterro
authors.forename: Teresa R
authors.initials: TR
authors.lastname: Pacheco
authors.forename: Theodorus W J
authors.initials: TW
authors.lastname: Gadella
authors.forename: Maria
authors.initials: M
authors.lastname: Carmo-Fonseca
mesh_terms: D004268:DNA-Binding Proteins / Q000502:physiology*; D006367:HeLa Cells; D006801:Humans; D009687:Nuclear Proteins / Q000502:physiology*; D009707:Nucleosomes / Q000378:metabolism; D011485:Protein Binding; D021381:Protein Transport; D012322:RNA Precursors / Q000378:metabolism; D012326:RNA Splicing; D000072260:RNA Splicing Factors; D012261:Ribonucleoproteins / Q000502:physiology*; D017381:Spliceosomes / Q000378:metabolism*; D000072668:Splicing Factor U2AF; D014157:Transcription Factors / Q000502:physiology*
chemical_list: D004268:DNA-Binding Proteins; D009687:Nuclear Proteins; D009707:Nucleosomes; D012322:RNA Precursors; D000072260:RNA Splicing Factors; D012261:Ribonucleoproteins; C107467:SF1 protein, human; D000072668:Splicing Factor U2AF; D014157:Transcription Factors; C076753:U2AF1 protein, human; C000607494:U2AF2 protein, human