Ran Binding Domains Promote the Interaction of Ran with p97/-Karyopherin, Linking the Docking and Translocation Steps of Nuclear Import

doi:10.1074/jbc.271.5.2357

Ran Binding Domains Promote the Interaction of Ran with p97/-Karyopherin, Linking the Docking and Translocation Steps of Nuclear Import (*)

From the Department of Pathology and Comprehensive Cancer Center, University of Vermont, Burlington, Vermont 05405

^§Supported by National Research Service Award F32 CA63801 from the National Cancer Institute. To whom correspondence should be addressed. Tel.: 802-656-0394; Fax: 802-656-8892; :klounsbu{at}moose.uvm.edu.

Abstract

Nuclear protein import is accomplished by two sequential events: docking at the nuclear pore complex followed by ATP-dependent translocation across the nuclear envelope. Docking of nuclear targeted proteins requires a 56-kDa nuclear localization signal receptor (α-karyopherin, importin-α, SRP1α) and a 97-kDa protein (β-karyopherin, importin-β). Components necessary for translocation include the Ran/TC4 GTPase and NTF2/B-2. The functions of these factors at a molecular level remain unclear. We have now found that a complex of Ran, in the GTP-bound state, with either the Ran binding protein, RanBP1, or an isolated Ran binding domain binds with high affinity and specificity to β-karyopherin to form a ternary complex. We find that a C-terminal truncation mutant of Ran, Δ-DE Ran, also binds to β-karyopherin and that Δ-DE Ran can associate with a cytosolic, multiprotein complex that contains β-karyopherin and another Δ-DE Ran binding protein of 115/120 kDa. These data suggest a physical link between docking and translocation mediated by a Ran GTPase-Ran binding protein complex.

Footnotes

↵^¶ Supported by National Institutes of Health Environmental Pathology Training Grant EST 3207122.
↵* This work was supported by National Institutes of Health Grant GM 50526 from the National Cancer Institute. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore by hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
↵¹ The abbreviations used are:

NLS

nuclear localization signal

RanBP

Ran binding protein

RanBD

Ran binding domain

Δ-DE Ran

C-terminally deleted Ran

GST

glutathione S-transferase

PAGE

polyacrylamide gel electrophoresis

FPLC

fast performance liquid chromatography

MOPS

3-(N-morpholino)propanesulfonic acid.
↵²K. Carey, S. A. Richards, K. L. Lounsbury, and I. G. Macara, submitted for publication.
- Received October 5, 1995.
- Revision received December 4, 1995.