Transportins 1 and 2 are redundant nuclear import factors for hnRNP A1 and HuR
Transportins 1 and 2 are redundant nuclear import factors for hnRNP A1 and HuR
复制标题
DOI：
10.1261/rna.5224304
复制
发表时间：
2004-04-01
期刊：
影响因子：
4.5
通讯作者：
Steitz, JA
中科院分区：
文献类型：
--
作者：
Rebane, A;Aab, A;Steitz, JA
一些mRNA结合蛋白，包括hnRNPA1和HuR，含有双向转运信号，介导其核输入和输出。以前，转运蛋白1（Trn1）被确定为hnRNPA1输入的介质，而密切相关的蛋白质转运蛋白2（Trn2）被证明与HuR相互作用。在这里，我们研究了转运蛋白的亚家族，包括Trn1（或Kap beta2A）和两种选择性剪接的Trn2亚型（Trn2a和Trn2b），也称为Trn2和Kap beta2B。这些蛋白质之间的序列差异可以改变它们的货物特异性或它们对RanGTP的反应，从而改变它们作为输入或输出受体的功能。使用体外结合试验，我们表明，hnRNP A1优先结合Trn1和Trn2b相对于Trn2a。HuR与所有三种转运蛋白相互作用，以及与Imp β弱相互作用。分别称为M9和HNS的hnRNP A1和HuR穿梭结构域足以进行这些相互作用。尽管HuR和hnRNP A1与三种转运蛋白的结合存在微小差异，但在存在和不存在RanQ69LGTP的情况下进行的体外相互作用研究表明，所有三种转运蛋白最有可能作为HuR和hnRNP A1的输入因子。在毛地黄皂苷透化的HeLa细胞中，M9和HNS肽都竞争重组hnRNP A1和HuR的输入，表明HuR和hnRNP A1输入途径至少部分重叠。hnRNPA1和HuR可能的核质穿梭机制进行了讨论。
Several mRNA-binding proteins, including hnRNP A1 and HuR, contain bidirectional transport signals that mediate both their nuclear import and export. Previously, Transportin 1 (Trn1) was identified as a mediator of hnRNP A1 import, whereas the closely related protein Transportin 2 (Trn2) was shown to interact with HuR. Here we have investigated the subfamily of transportins that consists of Trn1 (or Kap beta2A) and two alternatively spliced Trn2 isoforms (Trn2a and Trn2b), also called Trn2 and Kap beta2B. The sequence differences among these proteins could alter either their cargo specificity or their response to RanGTP and thus their function as import or export receptors. Using in vitro binding assays, we show that hnRNP A1 preferentially binds Trn1 and Trn2b versus Trn2a. HuR interacts with all three transportins, as well as weakly with Imp beta. The hnRNP A1 and HuR shuttling domains, called M9 and HNS, respectively, are sufficient for these interactions. Despite small differences in the binding of HuR and hnRNP A1 to the three transportins, in vitro interaction studies performed in the presence and absence of RanQ69LGTP indicate that all three transportins most likely act as import factors for HuR and hnRNP A1. In digitonin-permeabilized HeLa cells, both M9 and HNS peptides compete for the import of recombinant hnRNP A1 and HuR, indicating that HuR and hnRNP A1 import pathways are at least partially overlapping. Possible nucleocytoplasmic shuttling mechanisms for hnRNP A1 and HuR are discussed.