Nucleocytoplasmic shuttling of polypyrimidine tract-binding protein is uncoupled from RNA export
Nucleocytoplasmic shuttling of polypyrimidine tract-binding protein is uncoupled from RNA export
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DOI：
10.1091/mbc.12.12.3808
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发表时间：
2001-12-01
影响因子：
3.3
通讯作者：
Huang, S
中科院分区：
文献类型：
--
作者：
Kamath, RV;Leary, DJ;Huang, S
多聚嘧啶束结合蛋白PTB/hnRNP I参与细胞核中的前mRNA加工和细胞质中的RNA定位和翻译。在这份报告中，我们证明，PTB穿梭细胞核和细胞质之间的能量依赖性的方式。缺失突变表明，最低限度的N末端和RNA识别基序（RRM）1和2是必要的核质穿梭。RRM 3和RRM 4（主要负责RNA结合的结构域）的缺失加速了PTB的核质穿梭。单独或所有RNA聚合酶的RNA聚合酶II指导的转录抑制产生了类似的结果。相反，RNA聚合酶I的选择性抑制不影响PTB的穿梭动力学。此外，GFP-PTB的核内流动性，如通过光漂白分析后的荧光恢复所测量的，与转录活性细胞相比，在转录不活跃的细胞中显著增加。这些观察表明核RNA转录和输出对于PTB的穿梭不是必需的。此外，与由RNA聚合酶II和/或III转录的新生RNA结合会阻碍PTB的核输出和核质运动。PTB穿梭和RNA输出的解偶联表明PTB的核质穿梭也可能对其在细胞核和细胞质中的功能起调节作用。
Polypyrimidine tract binding protein, PTB/hnRNP I, is involved in pre-mRNA processing in the nucleus and RNA localization and translation in the cytoplasm. In this report, we demonstrate that PTB shuttles between the nucleus and cytoplasm in an energy-dependent manner. Deletion mutagenesis demonstrated that a minimum of the N terminus and RNA recognition motifs (RRMs) 1 and 2 are necessary for nucleocytoplasmic shuttling. Deletion of RRM3 and 4, domains that are primarily responsible for RNA binding, accelerated the nucleocytoplasmic shuttling of PTB. Inhibition of transcription directed by either RNA polymerase II alone or all RNA polymerases yielded similar results. In contrast, selective inhibition of RNA polymerase I did not influence the shuttling kinetics of PTB. Furthermore, the intranuclear mobility of GFP-PTB, as measured by fluorescence recovery after photobleaching analyses, increased significantly in transcriptionally inactive cells compared with transcriptionally active cells. These observations demonstrate that nuclear RNA transcription and export are not necessary for the shuttling of PTB. In addition, binding to nascent RNAs transcribed by RNA polymerase II and/or III retards both the nuclear export and nucleoplasmic movement of PTB. The uncoupling of PTB shuttling and RNA export suggests that the nucleocytoplasmic shuttling of PTB may also play a regulatory role for its functions in the nucleus and cytoplasm.