Katz D A, Theodorakis N G, Cleveland D W, Lindsten T, Thompson C B
Department of Molecular Genetics and Cell Biology, University of Chicago, IL 60637.
Nucleic Acids Res. 1994 Jan 25;22(2):238-46. doi: 10.1093/nar/22.2.238.
The 3'-untranslated regions of many labile transcripts contain AU-rich sequences that serve as cis determinants of mRNA stability and translational efficiency. Using a photocrosslinking technique, our laboratory has previously defined three cytoplasmic RNA-binding activities specific for the AUUUA multimers found in the 3'-untranslated regions of lymphokine mRNAs. One of these activities, AU-A, has an apparent molecular mass of 34 kDa, is constitutively expressed in both primary T cells and the Jurkat T cell leukemia line, and binds to a variety of U-rich RNA sequences. Previous studies had shown that AU-A is more prevalent in the nucleus than the cytoplasm, raising the possibility that AU-A is really a nuclear RNA-binding activity that is found in cytoplasmic extracts because of nuclear leakage during cell fractionation. We now show that AU-A shuttles between the cytoplasm and the nucleus. Our results indicate that AU-A is a candidate protein component of ribonucleoprotein complexes that participate in nucleocytoplasmic transport of mRNA and cytoplasmic mRNA metabolism. The properties of AU-A activity are similar to those of heterogeneous nuclear ribonucleoprotein A1 (hnRNP A1). However, using monoclonal antibodies to hnRNP A1 and protease digestion patterns, we show that AU-A activity and hnRNP A1 protein are distinct. These studies have also allowed us to define a fourth RNA-binding activity of apparent molecular mass 41 kDa with specificity for AUUUA multimers. This activity is restricted to the nucleus and contains the hnRNP C protein.
许多不稳定转录本的3'非翻译区含有富含AU的序列，这些序列作为mRNA稳定性和翻译效率的顺式决定因素。利用光交联技术，我们实验室先前已确定了三种细胞质RNA结合活性，它们对在淋巴因子mRNA的3'非翻译区中发现的AUUUA多聚体具有特异性。其中一种活性，即AU-A，其表观分子量为34 kDa，在原代T细胞和Jurkat T细胞白血病系中均组成性表达，并与多种富含U的RNA序列结合。先前的研究表明，AU-A在细胞核中比在细胞质中更普遍，这增加了AU-A实际上是一种核RNA结合活性的可能性，由于细胞分级分离过程中的核泄漏，这种活性在细胞质提取物中被发现。我们现在表明，AU-A在细胞质和细胞核之间穿梭。我们的结果表明，AU-A是核糖核蛋白复合物的候选蛋白质成分，参与mRNA的核质运输和细胞质mRNA代谢。AU-A活性的特性与异质性核核糖核蛋白A1（hnRNP A1）相似。然而，使用针对hnRNP A1的单克隆抗体和蛋白酶消化模式，我们表明AU-A活性和hnRNP A1蛋白是不同的。这些研究还使我们确定了第四种表观分子量为41 kDa的RNA结合活性，它对AUUUA多聚体具有特异性。这种活性仅限于细胞核，并含有hnRNP C蛋白。
