RanBP1 stabilizes the interaction of Ran with p97 nuclear protein import.
Three factors have been identified that reconstitute nuclear protein import in a permeabilized cell assay: the NLS receptor, p97, and Ran/TC4. Ran/TC4, in turn, interacts with a number of proteins that are involved in the regulation of GTP hydrolysis or are components of the nuclear pore. Two Ran-binding proteins, RanBP1 ... and RanBP2, form discrete complexes with p97 as demonstrated by immunoadsorption from HeLa cell extracts fractionated by gel filtration chromatography. A > 400-kD complex contains p97, Ran, and RanBP2. Another complex of 150-300 kD was comprised of p97, Ran, and RanBP1. This second trimeric complex could be reconstituted from recombinant proteins. In solution binding assays, Ran-GTP bound p97 with high affinity, but the binding of Ran-GDP to p97 was undetectable. The addition of RanBP1 with Ran-GDP or Ran-GTP increased the affinity of both forms of Ran for p97 to the same level. Binding of Ran-GTP to p97 dissociated p97 from immobilized NLS receptor while the Ran-GDP/RanBP1/p97 complex did not dissociate from the receptor. In a digitonin-permeabilized cell docking assay, RanBP1 stabilizes the receptor complex against temperature-dependent release from the pore. When added to an import assay with recombinant NLS receptor, p97 and Ran-GDP, RanBP1 significantly stimulates transport. These results suggest that RanBP1 promotes both the docking and translocation steps in nuclear protein import by stabilizing the interaction of Ran-GDP with p97.
Mesh Terms:
Biological Transport, Cell Membrane Permeability, Cell Nucleus, DNA-Binding Proteins, Digitonin, GTP-Binding Proteins, Guanosine Diphosphate, Guanosine Triphosphate, HeLa Cells, Humans, Macromolecular Substances, Molecular Chaperones, Nuclear Envelope, Nuclear Pore Complex Proteins, Nuclear Proteins, Protein Binding, Recombinant Fusion Proteins, alpha Karyopherins, beta Karyopherins, ran GTP-Binding Protein
Biological Transport, Cell Membrane Permeability, Cell Nucleus, DNA-Binding Proteins, Digitonin, GTP-Binding Proteins, Guanosine Diphosphate, Guanosine Triphosphate, HeLa Cells, Humans, Macromolecular Substances, Molecular Chaperones, Nuclear Envelope, Nuclear Pore Complex Proteins, Nuclear Proteins, Protein Binding, Recombinant Fusion Proteins, alpha Karyopherins, beta Karyopherins, ran GTP-Binding Protein
J. Cell Biol.
Date: Nov. 01, 1996
PubMed ID: 8909533
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