The snRNP-free U1A (SF-A) complex(es): identification of the largest subunit as PSF, the polypyrimidine-tract binding protein-associated splicing factor.
We have previously shown that a specific monoclonal antibody prepared against the U1A protein, MAb 12E12, is unique in its ability to recognize a form of U1A which is not associated with the U1snRNP. This unique form of U1A, termed snRNP-free U1A or SF-A, was found to be complexed with a novel set of non-snRNP proteins (O'Connor et al., 1997, RNA 3:1444-1455). Here we demonstrate that the largest protein in these SF-A complex(es), p105, is the polypyrimidine-tract binding protein-associated factor (PSF), an auxiliary splicing factor. We show that PSF copurifies and co-immunoprecipitates with SF-A from 293T cell nucleoplasm and that it interacts with SF-A in vitro. In addition, we show that MAb 12E12 inhibits both splicing and polyadenylation in an in vitro coupled splicing and polyadenylation reaction. This suggests that SF-A and/or the SF-A complex(es) perform an important function in both processing reactions and possibly in last exon definition.
Lutz
C S
CS
Department of Microbiology, University of Pennsylvania, School of Medicine, Philadelphia 19104, USA.
Cooke
C
C
O'Connor
J P
JP
Kobayashi
R
R
Alwine
J C
JC
D000595:Amino Acid Sequence; D002499:Centrifugation, Density Gradient; D005982:Glutathione Transferase / Q000378:metabolism; D008969:Molecular Sequence Data; D000072670:PTB-Associated Splicing Factor; D011061:Poly A / Q000037:antagonists & inhibitors; D011233:Precipitin Tests; D011485:Protein Binding; D012326:RNA Splicing; D016601:RNA-Binding Proteins / Q000737:chemistry / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D017412:Ribonucleoprotein, U1 Small Nuclear / Q000378:metabolism*
D000072670:PTB-Associated Splicing Factor; D016601:RNA-Binding Proteins; D011993:Recombinant Fusion Proteins; D017412:Ribonucleoprotein, U1 Small Nuclear; C075339:U1A protein; D011061:Poly A; D005982:Glutathione Transferase