Lutz C S, Cooke C, O'Connor J P, Kobayashi R, Alwine J C
Department of Microbiology, University of Pennsylvania, School of Medicine, Philadelphia 19104, USA.
RNA. 1998 Dec;4(12):1493-9. doi: 10.1017/s1355838298981183.
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.
我们之前已经表明，针对U1A蛋白制备的一种特异性单克隆抗体，即单克隆抗体12E12，在识别一种不与U1snRNP相关的U1A形式方面具有独特能力。这种独特的U1A形式，称为无snRNP的U1A或SF-A，被发现与一组新的非snRNP蛋白形成复合物（奥康纳等人，1997年，《RNA》3:1444 - 1455）。在这里，我们证明这些SF-A复合物中最大的蛋白p105是多嘧啶序列结合蛋白相关因子（PSF），一种辅助剪接因子。我们表明PSF与293T细胞核质中的SF-A共纯化并共免疫沉淀，并且它在体外与SF-A相互作用。此外，我们表明单克隆抗体12E12在体外偶联剪接和聚腺苷酸化反应中抑制剪接和聚腺苷酸化。这表明SF-A和/或SF-A复合物在这两个加工反应中以及可能在最后外显子定义中发挥重要作用。
