Lee W C, Xue Z X, Mélèse T
Department of Biological Sciences, Columbia University, New York 10027.
J Cell Biol. 1991 Apr;113(1):1-12. doi: 10.1083/jcb.113.1.1.
We previously identified a protein (p67) in the yeast, Saccharomyces cerevisiae, that specifically recognizes nuclear localization sequences. We report here the partial purification of p67, and the isolation, sequencing, and disruption of the gene (NSR1) encoding this protein. p67 was purified using an affinity column conjugated with a peptide containing the histone H2B nuclear localization sequence from yeast. Using antibodies against p67 we have cloned the gene for this protein. The protein encoded by the NSR1 gene recognizes the wild-type H2B nuclear localization sequence, but does not recognize a mutant H2B sequence that is incompetent for nuclear localization in vivo. Interestingly, the NSR1 protein has two RNA recognition motifs, as well as an acidic NH2 terminus containing a series of serine clusters, and a basic COOH terminus containing arg-gly repeats. We have confirmed the nuclear localization of p67 by immunofluorescence and found that a restricted portion of the nucleus is highlighted. We have also shown that NSR1 (p67) is required for normal cell growth.
我们之前在酿酒酵母中鉴定出一种蛋白质(p67),它能特异性识别核定位序列。我们在此报告p67的部分纯化,以及编码该蛋白质的基因(NSR1)的分离、测序和破坏。使用与含有来自酵母的组蛋白H2B核定位序列的肽偶联的亲和柱纯化p67。利用抗p67抗体,我们克隆了该蛋白质的基因。NSR1基因编码的蛋白质识别野生型H2B核定位序列,但不识别在体内无核定位能力的突变型H2B序列。有趣的是,NSR1蛋白有两个RNA识别基序,以及一个含有一系列丝氨酸簇的酸性NH2末端和一个含有精氨酸-甘氨酸重复序列的碱性COOH末端。我们通过免疫荧光证实了p67的核定位,并发现细胞核的一个受限部分被突出显示。我们还表明,正常细胞生长需要NSR1(p67)。