Ling Yan, Smith Abigail J, Morgan Garry T
Institute of Genetics, University of Nottingham, Queens Medical Centre, Nottingham NG7 2UH, UK.
Nucleic Acids Res. 2006 Apr 28;34(8):2219-29. doi: 10.1093/nar/gkl239. Print 2006.
The three structural domains of transcription elongation factor TFIIS are conserved from yeast to human. Although the N-terminal domain is not needed for transcriptional activity, a similar sequence has been identified previously in other transcription factors. We found this conserved sequence, the LW motif, in another three human proteins that are predominantly nuclear localized. We investigated two examples to determine whether the LW motif is actually a dedicated nuclear targeting signal. However, in one of the newly identified proteins, hIWS1 (human Iws1), a region containing classic nuclear localization signals (NLS) rather than the LW motif was necessary and sufficient for nuclear targeting in HeLa cells. In contrast, human TFIIS does not possess an NLS and only constructs containing the LW motif were efficiently targeted to nuclei. Moreover, mutations in the motif could cause cytoplasmic accumulation of TFIIS and enabled a structure/function assay for the domain based on the efficiency of nuclear targeting. Finally, GST pull-down assays showed that the LW motif is part of a protein-binding domain. We suggest that the targeting role the LW motif plays in TFIIS arises from its more general function as a protein interaction domain, enabling TFIIS to bind a carrier protein(s) that accomplishes nuclear import.
转录延伸因子TFIIS的三个结构域在从酵母到人类的进化过程中是保守的。尽管转录活性不需要N端结构域，但之前在其他转录因子中也发现了类似序列。我们在另外三种主要定位于细胞核的人类蛋白质中发现了这个保守序列，即LW基序。我们研究了两个例子，以确定LW基序是否实际上是一个专门的核定位信号。然而，在新鉴定的蛋白质之一hIWS1（人类Iws1）中，在HeLa细胞中，一个包含经典核定位信号（NLS）而非LW基序的区域对于核定位是必要且充分的。相比之下，人类TFIIS不具有NLS，只有包含LW基序的构建体才能有效地靶向细胞核。此外，该基序中的突变可能导致TFIIS在细胞质中积累，并基于核靶向效率对该结构域进行结构/功能分析。最后，GST下拉实验表明LW基序是蛋白质结合结构域的一部分。我们认为，LW基序在TFIIS中发挥的靶向作用源于其作为蛋白质相互作用结构域的更普遍功能，使TFIIS能够结合完成核输入的载体蛋白。
