We report here that importin α accumulates reversibly in the nucleus in response to cellular stresses including UV irradiation, oxidative stress, and heat shock. The nuclear accumulation of importin α appears to be triggered by a collapse in the Ran gradient, resulting in the suppression of the nuclear export of importin α. In addition, nuclear retention and the importin β/Ran-independent import of importin α also facilitate its rapid nuclear accumulation. The findings herein show that the classical nuclear import pathway is down-regulated via the removal of importin α from the cytoplasm in response to stress. Moreover, whereas the nuclear accumulation of heat shock cognate 70 is more sensitive to heat shock than the other stresses, importin α is able to accumulate in the nucleus at all the stress conditions tested. These findings suggest that the stress-induced nuclear accumulation of importin α can be involved in a common physiological response to various stress conditions.

The targeting of karyophilic proteins to nuclear pores is mediated via the formation of a nuclear pore-targeting complex, through the interaction of nuclear localization signal (NLS) with its NLS receptor. Recently, a novel human protein, Qip1, was identified from a yeast two-hybrid system with DNA helicase Q1. This study demonstrates that Qip1 is a novel third class of NLS receptor that efficiently recognizes the NLS of the helicase Q1. Moreover, the data obtained in this study show that the specific interaction between Qip1 and the NLS of the helicase Q1 requires its upstream sequence of the minimal essential NLS. By using purified recombinant proteins alone in the digitonin-permeabilized cell-free transport system, it was demonstrated that the two known human NLS receptors, Rch1 and NPI-1, are able to transport all the tested NLS substrates into the nucleus, while Qip1 most efficiently transports the helicase Q1-NLS substrates, which contain its upstream sequence in so far as we have examined the system. Furthermore, in HeLa cell crude cytosol, it was found that endogenous Rch1 binds to all the tested NLS substrates, while the binding of endogenous NPI-1 is restricted to only some NLSs, despite the fact that NPI-1 itself shows binding activity to a variety of NLSs. These results indicate that at least three structurally and functionally distinct NLS receptors exist in the human single cell population, and suggest that the nuclear import of karyophilic proteins may be controlled in a complex manner at the NLS recognition step by the existence of a variety of NLS receptors with various specificities to each NLS.In eukaryotic cells, the selective transport of karyophilic proteins to the nuclei is mediated by short amino acid sequences, which are commonly referred to as nuclear localization signals (NLSs) 1 and which are characteristically rich in basic amino acids (1-3). NLSs can be classified into two major groups. The first is a single type containing 3-5 basic amino acids with the weak consensus Lys-Arg/Lys-X-Arg/Lys, which is similar to the simian virus 40 large T antigen (SV40 T) NLS. The other is a bipartite type NLS containing two clusters of basic regions of 3-4 residues, each separated by approximately 10 amino acids, similar to nucleoplasmin NLS. The NLS functions at various positions within the protein and is capable of directing a non-karyophilic protein into nuclei when conjugated genetically or chemically (4).It is generally thought that the NLS-mediated nuclear transport of karyophilic proteins occurs in at least two steps (5-7). The first step is the NLS-dependent, but energy-and temperature-independent, binding to the cytoplasmic face of the nuclearporecomplex.Thesecondstepisanenergyandtemperaturedependent translocation through the nuclear pore complex.In earlier studies, we found that a karyophilic protein forms a stable complex, the nuclear pore-targeting complex (PTAC) in the cytoplasm to target nuclear pores (4, 8). The complex consists of a karyophilic protein and two essential factors, PTA...

Importin α performs the indispensable role of ferrying proteins from the cytoplasm into the nucleus with a transport carrier, importin β1. Mammalian cells from mouse or human contain either six or seven importin α subtypes, respectively, each with a tightly regulated expression. Therefore, the combination of subtype expression in a cell defines distinct signaling pathways to achieve progressive changes in gene expression essential for cellular events, such as differentiation. Recent studies reveal that, in addition to nucleocytoplasmic transport, importin αs also serve non-transport functions. In this review, we first discuss the physiological significance of importin α as a nuclear transport regulator, and then focus on the functional diversities of importin αs based on their specific subcellular and cellular localizations, such as the nucleus and plasma membrane. These findings enrich our knowledge of how importin αs actively contribute to various cellular events.

Monocytes/macrophages are major targets of human immunodeficiency virus type 1 (HIV-1) infection. The viral preintegration complex (PIC) of HIV-1 enters the nuclei of monocyte-derived macrophages, but very little PIC migrates into the nuclei of immature monocytes. Vpr, one of the accessory gene products of HIV-1, is essential for the nuclear import of PIC in these cells, although the role of Vpr in the entry mechanism of PIC remains to be clarified. We have shown previously that Vpr is targeted to the nuclear envelope and then transported into the nucleus by importin ␣ alone, in an importin ␤-independent manner. Here we demonstrate that the nuclear import of Vpr is strongly promoted by the addition of cytoplasmic extract from macrophages but not of that from monocytes and that the nuclear import activity is lost with immunodepletion of importin ␣ from the cytoplasmic extract. Immunoblot analysis and real-time PCR demonstrate that immature monocytes express importin ␣ at low levels, whereas the expression of three major importin ␣ isoforms markedly increases upon their differentiation into macrophages, indicating that the expression of importin ␣ is required for nuclear import of Vpr. Furthermore, interaction between importin ␣ and the N-terminal ␣-helical domain of Vpr is indispensable, not only for the nuclear import of Vpr but also for HIV-1 replication in macrophages. This study suggests the possibility that the binding of Vpr to importin ␣, preceding a novel nuclear import process, is a potential target for therapeutic intervention.