Lawe D C, Hahn C, Wong A J
Department of Microbiology and Immunology, The Kimmel Cancer Institute, Philadelphia, Pennsylvania 19107, USA.
Oncogene. 1997 Jan 16;14(2):223-31. doi: 10.1038/sj.onc.1200821.
SH2/SH3 adaptor proteins are essential components of the signal transduction pathways initiated by tyrosine kinases. Nck is a ubiquitously expressed adaptor protein whose function has been enigmatic. We performed confocal microscopy to localize Nck in NIH3T3 and A431 cells. Surprisingly, Nck was identified in the nucleus as well as the cytoplasm with no visible change in localization due to PDGF or EGF stimulation. Western blot analysis of nuclear and cytosolic fractions confirmed that there was no translocation in response to growth factor and that tyrosine phosphorylation was specific to only cytosolic Nck. Far Western blot analysis with either Nck, the SH2 domain, or the SH3 domains revealed differential binding in nuclear and cytosolic lysates, indicating specific binding partners for each subcellular location. The major target of c-Src during mitosis is SAM68, a RNA-binding protein ordinarily localized to the nucleus. SAM68 was identified as a nuclear specific binding partner of Nck in both nonmitotic and mitotic cells. Several tyrosine kinases can be found in the nucleus but their signal transduction remains undefined. The discovery of an adaptor protein in the nucleus suggests there are signal transduction mechanisms within the nucleus that recapitulate those found in the cytoplasm.
SH2/SH3衔接蛋白是酪氨酸激酶启动的信号转导通路的重要组成部分。Nck是一种广泛表达的衔接蛋白，其功能一直成谜。我们进行了共聚焦显微镜检查，以在NIH3T3和A431细胞中定位Nck。令人惊讶的是，在细胞核和细胞质中均发现了Nck，并且由于血小板衍生生长因子（PDGF）或表皮生长因子（EGF）刺激，其定位没有明显变化。对细胞核和细胞质组分的蛋白质免疫印迹分析证实，对生长因子没有发生易位，并且酪氨酸磷酸化仅对细胞质中的Nck具有特异性。用Nck、SH2结构域或SH3结构域进行的Far Western印迹分析显示，在细胞核和细胞质裂解物中有不同的结合，表明每个亚细胞位置都有特定的结合伴侣。有丝分裂期间c-Src的主要靶标是SAM68，一种通常定位于细胞核的RNA结合蛋白。在非有丝分裂和有丝分裂细胞中，SAM68均被鉴定为Nck的核特异性结合伴侣。在细胞核中可以发现几种酪氨酸激酶，但其信号转导仍不明确。在细胞核中发现一种衔接蛋白表明，细胞核内存在一些信号转导机制，这些机制与细胞质中的信号转导机制相似。
