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Originally published as MBC in Press, 10.1091/mbc.E02-06-0372 on February 6, 2003

Vol. 14, Issue 5, 2104-2115, May 2003

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Importin {beta}-depending Nuclear Import Pathways: Role of the Adapter Proteins in the Docking and Releasing Steps

Christiane Rollenhagen * {dagger}, Petra Mühlhäusser {dagger}, Ulrike Kutay {dagger}, and Nelly Panté * {ddagger}

* Department of Zoology, University of British Columbia, Vancouver, British Columbia, V6T 1Z4 Canada; {dagger} Swiss Federal Institute of Technology Zurich, Institute of Biochemistry, Hönggerberg, CH-8093 Zurich, Switzerland

Submitted June 29, 2002; Revised November 22, 2002; Accepted January 16, 2003
Monitoring Editor: Pamela Silver

Nuclear imports of uridine-rich small nuclear ribonucleoprotein (U1 snRNP) and proteins with classical nuclear localization signal (cNLS-protein) are mediated by importin {beta}. However, due to the presence of different import signals, the adapter protein of the imported molecules and importin {beta} is different for each pathway. Although the adapter for cNLS-protein is importin {alpha}, the adapter for U1 snRNP is snurportin1 (SPN1). Herein, we show that the use of distinct adapters by importin {beta} results in differences at the docking and releasing step for these two import pathways. Nuclear pore complex (NPC) docking of U1 snRNP but not of cNLS-protein was inhibited by an anti-CAN/Nup214 antibody. Thus, the initial NPC-binding site is different for each pathway. Pull-down assays between immobilized SPN1 and two truncated forms of importin {beta} documented that SPN1 and importin {alpha} have different binding sites on importin {beta}. Importin {beta} fragment 1–618, which binds to SPN1 but not to importin {alpha}, was able to support the nuclear import of U1 snRNPs. After the translocation through the NPC, both import complexes associated with the nuclear side of the NPC. However, we found that the nature of the importin {beta}-binding domain of the adapters influences the release of the cargo into the nucleoplasm.


Article published online ahead of print. Mol. Biol. Cell 10.1091/mbc.E02-06-0372. Article and publication date are at www.molbiolcell.org/cgi/doi/10.1091/mbc.E02-06-0372.

Abbreviations used: BSA, bovine serum albumin; cNLS, classical nuclear localization sequence; cNLS-protein, cNLS-bearing protein; EM, electron microscopy; IBB, importin {beta}-binding; IBB{alpha}, recombinant {beta}-galactosidase molecules fused to the IBB domain of importin {alpha}; IBBspn1, recombinant {beta}-galactosidase molecules fused to the IBB domain of snurportin1; NPC, nuclear pore complex; SPN1, snurportin1; WGA, wheat germ agglutinin; zz-tag, two immunoglobulin-binding domains of Staphylococcus aureus protein A.

{ddagger} Corresponding author. E-mail address: pante{at}zoology.ubc.ca.




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