Prieve M G, Guttridge K L, Munguia J E, Waterman M L
Department of Microbiology and Molecular Genetics, University of California, Irvine, 92717-4025, USA.
J Biol Chem. 1996 Mar 29;271(13):7654-8. doi: 10.1074/jbc.271.13.7654.
Proteins are directed to the nucleus by their nuclear localization sequences (NLSs) in a multistep process. The first step, which is to dock the NLS-containing protein to the nuclear pore, is carried out in part by a recently identified NLS receptor named Srp1/importin-alpha. Using the high mobility group (HMG) DNA binding domain of human lymphoid enhancer factor-1 (hLEF-1) as bait in a yeast two-hybrid screen, we have identified two different mouse Srp1 proteins (pendulin/importin-alpha and mSrp1) that each bind to a 9-amino acid sequence in hLEF-1 called the B box. We show that the B box of hLEF-1, a region essential for high affinity DNA binding, is also necessary and sufficient for nuclear localization, lending support to the model that NLSs can function both in nuclear transport and DNA binding. Pendulin and mSrp1 are the mouse homologues of hRch1/hSrp1alpha/importin-alpha and hSrp1/karyopherin alpha/NPI-1, respectively, and show considerable sequence divergence from each other. We find a surprising and significant difference in the expression pattern of pendulin and mSrp1 mRNA, suggesting that these two Srp1 proteins are distinguishable in function as well as sequence.
蛋白质通过其核定位序列(NLSs)在一个多步骤过程中被导向细胞核。第一步,即将含有NLS的蛋白质停靠到核孔上,部分是由最近鉴定出的一种名为Srp1/输入蛋白α的NLS受体完成的。在酵母双杂交筛选中,我们使用人类淋巴增强因子-1(hLEF-1)的高迁移率族(HMG)DNA结合结构域作为诱饵,鉴定出两种不同的小鼠Srp1蛋白(pendulin/输入蛋白α和mSrp1),它们各自与hLEF-1中一个名为B框的9个氨基酸序列结合。我们表明,hLEF-1的B框是高亲和力DNA结合所必需的区域,对于核定位也是必需且充分的,这支持了NLSs在核运输和DNA结合中都能发挥作用的模型。Pendulin和mSrp1分别是hRch1/hSrp1α/输入蛋白α和hSrp1/核转运蛋白α/NPI-1的小鼠同源物,并且彼此之间显示出相当大的序列差异。我们发现pendulin和mSrp1 mRNA的表达模式存在惊人且显著的差异,这表明这两种Srp1蛋白在功能以及序列上都是可区分的。