Nucleocytoplasmic shuttling of heterodimeric splicing factor U2AF.
The U2 small nuclear ribonucleoprotein auxiliary factor (U2AF) is a heterodimeric splicing factor composed of 65-kDa (U2AF(65)) and 35-kDa (U2AF(35)) subunits. The large subunit of U2AF recognizes the intronic polypyrimidine tract, a sequence located adjacent to the 3' splice site that serves as an important signal for both constitutive and regulated pre-mRNA splicing. The small subunit U2AF(35) interacts with the 3' splice site dinucleotide AG and is essential for regulated splicing. Like several other proteins involved in constitutive and regulated splicing, both U2AF(65) and U2AF(35) contain an arginine/serine-rich (RS) domain. In the present study we determined the role of RS domains in the subcellular localization of U2AF. Both U2AF(65) and U2AF(35) are shown to shuttle continuously between the nucleus and the cytoplasm by a mechanism that involves carrier receptors and is independent from binding to mRNA. The RS domain on either U2AF(65) or U2AF(35) acts as a nuclear localization signal and is sufficient to target a heterologous protein to the nuclear speckles. Furthermore, the results suggest that the presence of an RS domain in either U2AF subunit is sufficient to trigger the nucleocytoplasmic import of the heterodimeric complex. Shuttling of U2AF between nucleus and cytoplasm possibly represents a means to control the availability of this factor to initiate spliceosome assembly and therefore contribute to regulate splicing.
Gama-Carvalho
M
M
Institute of Histology and Embryology, Faculty of Medicine, University of Lisbon, 1649-028 Lisbon, Portugal. m.gamacarvalho@fm.ul.pt
Carvalho
M P
MP
Kehlenbach
A
A
Valcarcel
J
J
Carmo-Fonseca
M
M
D016475:3T3 Cells; D000595:Amino Acid Sequence; D000818:Animals; D001120:Arginine; D002467:Cell Nucleus / Q000502:physiology*; D003593:Cytoplasm / Q000502:physiology; D017931:DNA Primers; D019281:Dimerization; D049452:Green Fluorescent Proteins; D006367:HeLa Cells; D006801:Humans; D008164:Luminescent Proteins / Q000032:analysis / Q000235:genetics; D051379:Mice; D009687:Nuclear Proteins; D016133:Polymerase Chain Reaction; D021381:Protein Transport; D012326:RNA Splicing; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000378:metabolism; D012261:Ribonucleoproteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D012694:Serine; D000072668:Splicing Factor U2AF; D014158:Transcription, Genetic; D014162:Transfection
D017931:DNA Primers; D008164:Luminescent Proteins; D009687:Nuclear Proteins; D011993:Recombinant Fusion Proteins; D012261:Ribonucleoproteins; D000072668:Splicing Factor U2AF; C000607494:U2AF2 protein, human; C000607495:Zrsr1 protein, mouse; D049452:Green Fluorescent Proteins; D012694:Serine; D001120:Arginine