title: Efficiency of importin alpha/beta-mediated nuclear localization sequence recognition and nuclear import. Differential role of NTF2.
abstract: Little quantitative, kinetic information is available with respect to the process of nuclear import of conventional nuclear localization sequence (NLS)-containing proteins, which initially involves recognition and docking at the nuclear pore by importin alpha/beta. This study compares the binding and nuclear import properties of mouse (m) and yeast (y) importin (IMP) subunits with respect to the NLSs from the SV40 large tumor antigen (T-ag), and the Xenopus laevis phosphoprotein N1N2. m- and y-IMPalpha recognized both NLSs, with y-IMPalpha exhibiting higher affinity. m-IMPbeta greatly enhanced the binding of m-IMPalpha to the T-ag and N1N2 NLSs, but y-IMPbeta did not significantly affect the affinity of y-IMPalpha for the T-ag NLS. In contrast, y-IMPbeta enhanced y-IMPalpha binding to the NLS of N1N2, but to a lesser extent than the enhancement of m-IMPalpha binding by m-IMPbeta. NLS-dependent nuclear import was reconstituted in vitro using the different importin subunits together with the transport factors Ran and NTF2. Whereas T-ag NLS-mediated nuclear import did not exhibit an absolute requirement for NTF2, N1N2 NLS-mediated transport strictly required NTF2. High levels of NTF2 inhibited nuclear accumulation conferred by both NLSs. We conclude that different NLSs possess distinct nuclear import properties due to differences in recognition by importin and requirements for NTF2.
authors.affiliation: Nuclear Signalling Laboratory, Division for Biochemistry and Molecular Biology, John Curtin School of Medical Research, Canberra, ACT 2601, Australia.
authors.forename: W
authors.initials: W
authors.lastname: Hu
authors.forename: D A
authors.initials: DA
authors.lastname: Jans
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000952:Antigens, Polyomavirus Transforming / Q000378:metabolism; D001692:Biological Transport; D002352:Carrier Proteins / Q000378:metabolism; D002467:Cell Nucleus / Q000378:metabolism*; D028884:Karyopherins; D007700:Kinetics; D051379:Mice; D008969:Molecular Sequence Data; D009685:Nuclear Envelope / Q000378:metabolism; D019913:Nuclear Localization Signals / Q000235:genetics*; D009687:Nuclear Proteins / Q000378:metabolism*; D029543:Nucleocytoplasmic Transport Proteins; D010750:Phosphoproteins / Q000378:metabolism; D011485:Protein Binding; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D014407:Tumor Cells, Cultured; D014982:Xenopus laevis; D015003:Yeasts; D020931:ran GTP-Binding Protein
chemical_list: D000952:Antigens, Polyomavirus Transforming; D002352:Carrier Proteins; D028884:Karyopherins; D019913:Nuclear Localization Signals; D009687:Nuclear Proteins; D029543:Nucleocytoplasmic Transport Proteins; C503613:Nutf2 protein, mouse; D010750:Phosphoproteins; D011993:Recombinant Fusion Proteins; D020931:ran GTP-Binding Protein