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Molecular and Cellular Biology, February 1999, p. 1547-1557, Vol. 19, No. 2
0270-7306/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.

Interactions between a Nuclear Transporter and a Subset of Nuclear Pore Complex Proteins Depend on Ran GTPase

Matthias Seedorf,dagger Marc Damelin, Jason Kahana,Dagger Tetsuya Taura, and Pamela A. Silver*

Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, and The Dana-Farber Cancer Institute, Boston, Massachusetts 02115

Received 12 August 1998/Returned for modification 12 October 1998/Accepted 28 October 1998

Proteins to be transported into the nucleus are recognized by members of the importin-karyopherin nuclear transport receptor family. After docking at the nuclear pore complex (NPC), the cargo-receptor complex moves through the aqueous pore channel. Once cargo is released, the importin then moves back through the channel for new rounds of transport. Thus, importin and exportin, another member of this family involved in export, are thought to continuously shuttle between the nuclear interior and the cytoplasm. In order to understand how nuclear transporters traverse the NPC, we constructed functional protein fusions between several members of the yeast importin family, including Pse1p, Sxm1p, Xpo1p, and Kap95p, and the green fluorescent protein (GFP). Complexes containing nuclear transporters were isolated by using highly specific anti-GFP antibodies. Pse1-GFP was studied in the most detail. Pse1-GFP is in a complex with importin-alpha and -beta (Srp1p and Kap95p in yeast cells) that is sensitive to the nucleotide-bound state of the Ran GTPase. In addition, Pse1p associates with the nucleoporins Nsp1p, Nup159p, and Nup116p, while Sxm1p, Xpo1p, and Kap95p show different patterns of interaction with nucleoporins. Association of Pse1p with nucleoporins also depends on the nucleotide-bound state of Ran; when Ran is in the GTP-bound state, the nucleoporin association is lost. A mutant form of Pse1p that does not bind Ran also fails to interact with nucleoporins. These data indicate that transport receptors such as Pse1p interact in a Ran-dependent manner with certain nucleoporins. These nucleoporins may represent major docking sites for Pse1p as it moves in or out of the nucleus via the NPC.


* Corresponding author. Mailing address: The Dana-Farber Cancer Institute, 44 Binney St., Boston, MA 02115. Phone: (617) 632-5102. Fax: (617) 632-5103. E-mail: pfas.harvard.silver{at}edu.

dagger Present address: ZMBH, Im Neuenheimer Feld 282, D-69120 Heidelberg, Germany.

Dagger Present address: Ludwig Institute for Cancer Research, University of California at San Diego, La Jolla, Calif.


Molecular and Cellular Biology, February 1999, p. 1547-1557, Vol. 19, No. 2
0270-7306/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.



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