Consensus PP1 binding motifs regulate transcriptional corepression and alternative RNA splicing activities of the steroid receptor coregulators, p54nrb and PSF.

共有 PP1 结合基序调节类固醇受体共调节剂的转录抑制和选择性 RNA 剪接活性 ，p54nrb 和 PSF 。

• 发表时间: Jul 2011
• 来源期刊: Mol Endocrinol
• DOI: 10.1210/me.2010-0517
• PMID: 21566083
• 文章类型: Journal Article
• 中科院分区: Q2
• 方向: 医学
• 作者列表: Liu L,Xie N,Rennie P,Challis JR,Gleave M,Lye SJ,Dong X

Mesh: Alternative Splicing; Binding Sites; Cell Line, Tumor; DNA-Binding Proteins; Gene Expression Regulation; HEK293 Cells; Humans; Hyaluronan Receptors / genetics metabolism; Immunoprecipitation; Nuclear Matrix-Associated Proteins / metabolism; Octamer Transcription Factors / metabolism; PTB-Associated Splicing Factor; Phosphorylation; Protein Binding; Protein Phosphatase 1 / chemistry metabolism; Protein Transport; RNA-Binding Proteins / metabolism; Recombinant Fusion Proteins / metabolism; Transcription, Genetic

Abstract

Originally identified as essential pre-mRNA splicing factors, non-POU-domain-containing, octamer binding protein (p54nrb) and PTB-associated RNA splicing factor (PSF) are also steroid receptor corepressors. The mechanisms by which p54nrb and PSF regulate gene transcription remain unclear. Both p54nrb and PSF contain protein phosphatase 1 (PP1) consensus binding RVxF motifs, suggesting that PP1 may regulate phosphorylation status of p54nrb and PSF and thus their function in gene transcription. In this report, we demonstrated that PP1 forms a protein complex with both p54nrb and PSF. PP1 interacts directly with the RVxF motif only in p54nrb, but not in PSF. Association with PP1 results in dephosphorylation of both p54nrb and PSF in vivo and the loss of their transcriptional corepressor activities. Using the CD44 minigene as a reporter, we showed that PP1 regulates p54nrb and PSF alternative splicing activities that determine exon skipping vs. inclusion in the final mature RNA for translation. In addition, changes in transcriptional corepression and RNA splicing activities of p54nrb and PSF are correlated with alterations in protein interactions of p54nrb and PSF with transcriptional corepressors such as Sin3A and histone deacetylase 1, and RNA splicing factors such as U1A and U2AF. Furthermore, we demonstrated a novel function of the RVxF motif within PSF that enhances its corepression and RNA splicing activities independent of PP1. We conclude that the RVxF motifs play an important role in controlling the multifunctional properties of p54nrb and PSF in the regulation of gene transcription.

摘要

最初被确定为必需的前mRNA剪接因子，非POU域，八聚体结合蛋白（p54nrb）和PTB相关RNA剪接因子（PSF）也是类固醇受体的共抑制因子。p54nrb和PSF调控基因转录的机制尚不清楚。p54nrb和PSF都包含蛋白磷酸酶1（PP1）共有结合RVxF基序，表明PP1可能调节p54nrb和PSF的磷酸化状态，从而调节它们在基因转录中的功能。在这份报告中,我们证明PP1与p54nrb和PSF形成蛋白质复合物。PP1仅在p54nrb中与RVxF基序直接交互，但不是在PSF。与PP1的结合导致体内p54nrb和PSF的去磷酸化以及它们的转录抑制物活性的丧失。使用CD44小基因作为报告基因，我们显示PP1调节p54nrb和PSF选择性剪接活性，这些活性决定外显子跳跃与包含在最终的成熟RNA中用于翻译。此外,p54nrb和PSF的转录抑制和RNA剪接活性的变化与p54nrb和PSF与转录抑制因子如Sin3A和组蛋白脱乙酰酶1以及RNA剪接因子如U1A和U2AF的蛋白质相互作用的变化相关。此外，我们证明了RVxF基序在PSF中的一种新功能，该功能增强了其与PP1无关的协同抑制和RNA剪接活性。我们得出的结论是，RVxF基序在控制p54nrb和PSF在基因转录调控中的多功能特性中起着重要作用。