Flaherty S M, Fortes P, Izaurralde E, Mattaj I W, Gilmartin G M
Department of Microbiology and Molecular Genetics, The Markey Center for Molecular Genetics, Vermont Cancer Center, University of Vermont, Burlington, VT 05405, USA.
Proc Natl Acad Sci U S A. 1997 Oct 28;94(22):11893-8. doi: 10.1073/pnas.94.22.11893.
Communication between the 5' and 3' ends is a common feature of several aspects of eukaryotic mRNA metabolism. In the nucleus, the pre-mRNA 5' end is bound by the nuclear cap binding complex (CBC). This RNA-protein complex plays an active role in both splicing and RNA export. We provide evidence for participation of CBC in the processing of the 3' end of the message. Depletion of CBC from HeLa cell nuclear extract strongly reduced the endonucleolytic cleavage step of the cleavage and polyadenylation process. Cleavage was restored by addition of recombinant CBC. CBC depletion was found to reduce the stability of poly(A) site cleavage complexes formed in nuclear extract. We also provide evidence that the communication between the 5' and 3' ends of the pre-mRNA during processing is mediated by the physical association of the CBC/cap complex with 3' processing factors bound at the poly(A) site. These observations, along with previous data on the function of CBC in splicing, illustrate the key role played by CBC in pre-mRNA recognition and processing. The data provides further support for the hypothesis that pre-mRNAs and mRNAs may exist and be functional in the form of "closed-loops," due to interactions between factors bound at their 5' and 3' ends.
5'端与3'端之间的通讯是真核生物mRNA代谢多个方面的一个共同特征。在细胞核中，前体mRNA的5'端与核帽结合复合物（CBC）结合。这种RNA-蛋白质复合物在剪接和RNA输出过程中均发挥着积极作用。我们提供了CBC参与信使RNA 3'端加工的证据。从HeLa细胞核提取物中去除CBC会强烈降低切割和多聚腺苷酸化过程中的内切核酸酶切割步骤。通过添加重组CBC可恢复切割。研究发现，去除CBC会降低在核提取物中形成的聚腺苷酸化位点切割复合物的稳定性。我们还提供了证据表明，在前体mRNA加工过程中，5'端与3'端之间的通讯是由CBC/帽复合物与结合在聚腺苷酸化位点的3'加工因子的物理结合介导的。这些观察结果，连同先前关于CBC在剪接中功能的数据，说明了CBC在前体mRNA识别和加工中所起的关键作用。这些数据进一步支持了这样一种假说，即由于在前体mRNA和mRNA的5'端与3'端结合的因子之间存在相互作用，它们可能以“闭环”形式存在并发挥功能。
