Identification and functional analysis of cytoplasmic factors involved in nuclear protein transport

亲核蛋白靶向核孔需要几种细胞质因子，包括核定位信号(NLS)结合蛋白。利用洋地黄素通透性的无细胞转运实验，我们获得了含有与亲核蛋白特异结合的因子的细胞质组分，并支持转运的核结合步骤。这一组分中的组分通过与NLS的相互作用与亲核分子形成稳定的络合物。由于这种络合物在没有其他胞质因子的情况下在核进入之前就具有核孔结合活性，所以我们称之为核孔靶向络合物。它由亲核蛋白和分别为54、56、66和90 kDa的4种蛋白组成。在我们的重组实验中，一个由54和90 kDa蛋白质组成的复合体能够将亲核细胞靶向核孔。

Targeting of karyophilic proteins to nuclear pores is known to require several cytoplasmic factors including the nuclear location signal (NLS) -binding protein. Using a digitonin-permeabilized cell-free transport assay, we have obtained a cytoplasmic fraction containing factors that specifically bind to karyophilic protein and support the nuclear binding step of the transport. Components in this fraction form a stable complex with the karyophile through interaction with NLS.Since this complex shows nuclear pore binding activity prior to nuclear entry in the absence of other cytosolic factors, we call it nuclear pore-targeting complex. It consists of karyophilic protein and four proteins of 54,56,66, and 90kDa. In our reconstitution experiments, a complex with 54 and 90kDa proteins is capable of targeting karyophiles to the nuclear pores.