RanBP1 is crucial for the release of RanGTP from importin β‐related nuclear transport factors

Nucleocytoplasmic transport appears mediated by shuttling transport receptors that bind RanGTP as a means to regulate interactions with their cargoes. The receptor·RanGTP complexes are kinetically very stable with nucleotide exchange and GTP hydrolysis being blocked, predicting that a specific disassembly mechanism exists. Here we show in three cases receptor·RanGTP·RanBP1 complexes to be the key disassembly intermediates, where RanBP1 stimulates the off‐rate at the receptor/RanGTP interface by more than two orders of magnitude. The transiently released RanGTP·RanBP1 complex is then induced by RanGAP to hydrolyse GTP, preventing the receptor to rebind RanGTP. The efficient release of importin β from RanGTP requires importin α, in addition to RanBP1.

[1]  P. D’Eustachio,et al.  Characterization of proteins that interact with the cell-cycle regulatory protein Ran/TC4 , 1993, Nature.

[2]  P. Silver,et al.  The GTP-bound form of the yeast Ran/TC4 homologue blocks nuclear protein import and appearance of poly(A)+ RNA in the cytoplasm. , 1995, Proceedings of the National Academy of Sciences of the United States of America.

[3]  M. Fornerod,et al.  The human homologue of yeast CRM1 is in a dynamic subcomplex with CAN/Nup214 and a novel nuclear pore component Nup88 , 1997, The EMBO journal.

[4]  K. Weber,et al.  A functional homologue of the RNA1 gene product in Schizosaccharomyces pombe: purification, biochemical characterization, and identification of a leucine-rich repeat motif. , 1993, Molecular biology of the cell.

[5]  G. Blobel,et al.  A novel ubiquitin-like modification modulates the partitioning of the Ran-GTPase-activating protein RanGAP1 between the cytosol and the nuclear pore complex , 1996, The Journal of cell biology.

[6]  I. Macara,et al.  Requirement of guanosine triphosphate-bound ran for signal-mediated nuclear protein export. , 1997, Science.

[7]  S. Adam,et al.  RanBP1 stabilizes the interaction of Ran with p97 nuclear protein import , 1996, The Journal of cell biology.

[8]  F. Bischoff,et al.  RNA1 Encodes a GTPase-activating Protein Specific for Gsp1p, the Ran/TC4 Homologue of Saccharomyces cerevisiae(*) , 1995, The Journal of Biological Chemistry.

[9]  G. Blobel,et al.  The GTP-binding protein Ran/TC4 is required for protein import into the nucleus , 1993, Nature.

[10]  S. Adam,et al.  Functional domains in nuclear import factor p97 for binding the nuclear localization sequence receptor and the nuclear pore. , 1997, Molecular biology of the cell.

[11]  G. Blobel,et al.  Disassembly of RanGTP-Karyopherin β Complex, an Intermediate in Nuclear Protein Import* , 1997, The Journal of Biological Chemistry.

[12]  G. Dreyfuss,et al.  A Novel Receptor-Mediated Nuclear Protein Import Pathway , 1996, Cell.

[13]  I. Macara,et al.  Ran-binding Protein 1 (RanBP1) Forms a Ternary Complex with Ran and Karyopherin β and Reduces Ran GTPase-activating Protein (RanGAP) Inhibition by Karyopherin β* , 1997, The Journal of Biological Chemistry.

[14]  F. Bischoff Human RanGTPase activating protein RanGAP1 is a homolog of yeast RNA1p involved in messenger RNA processing and transport , 1995 .

[15]  I. Macara,et al.  The C Terminus of the Nuclear RAN/TC4 GTPase Stabilizes the GDP-bound State and Mediates Interactions with RCC1, RAN-GAP, and HTF9A/RANBP1 (*) , 1995, The Journal of Biological Chemistry.

[16]  P. Bork,et al.  A Novel Class of RanGTP Binding Proteins , 1997, The Journal of cell biology.

[17]  Elena Smirnova,et al.  Yrb4p, a yeast Ran–GTP‐binding protein involved in import of ribosomal protein L25 into the nucleus , 1997, The EMBO journal.

[18]  F. Melchior,et al.  Inhibition of nuclear protein import by nonhydrolyzable analogues of GTP and identification of the small GTPase Ran/TC4 as an essential transport factor [published erratum appears in J Cell Biol 1994 Jan;124(1-2):217] , 1993, The Journal of cell biology.

[19]  G. Blobel,et al.  Nup358, a Cytoplasmically Exposed Nucleoporin with Peptide Repeats, Ran-GTP Binding Sites, Zinc Fingers, a Cyclophilin A Homologous Domain, and a Leucine-rich Region (*) , 1995, The Journal of Biological Chemistry.

[20]  F. Bischoff,et al.  Catalysis of guanine nucleotide exchange on Ran by the mitotic regulator RCC1 , 1991, Nature.

[21]  I. Macara,et al.  A nuclear export signal is essential for the cytosolic localization of the Ran binding protein, RanBP1 , 1996, The Journal of cell biology.

[22]  G. Blobel,et al.  Protein import into nuclei: association and dissociation reactions involving transport substrate, transport factors, and nucleoporins , 1995, Cell.

[23]  Karsten Weis,et al.  Exportin 1 (Crm1p) Is an Essential Nuclear Export Factor , 1997, Cell.

[24]  F. Bischoff,et al.  RanGAP1 induces GTPase activity of nuclear Ras-related Ran. , 1994, Proceedings of the National Academy of Sciences of the United States of America.

[25]  F. Bischoff,et al.  Catalysis of guanine nucleotide exchange of Ran by RCC1 and stimulation of hydrolysis of Ran-bound GTP by Ran-GAP1. , 1995, Methods in enzymology.

[26]  B. Cullen,et al.  Nuclear import of hnRNP A1 is mediated by a novel cellular cofactor related to karyopherin-beta. , 1997, Journal of cell science.

[27]  M. Ohtsubo,et al.  The RCC1 protein, a regulator for the onset of chromosome condensation locates in the nucleus and binds to DNA , 1989, The Journal of cell biology.

[28]  F. Bischoff,et al.  Identification of different roles for RanGDP and RanGTP in nuclear protein import. , 1996, The EMBO journal.

[29]  P. Silver,et al.  Rna1p, a Ran/TC4 GTPase activating protein, is required for nuclear import , 1995, The Journal of cell biology.

[30]  Tatsuya Seki,et al.  A giant nucleopore protein that binds Ran/TC4 , 1995, Nature.

[31]  F. Melchior,et al.  A Small Ubiquitin-Related Polypeptide Involved in Targeting RanGAP1 to Nuclear Pore Complex Protein RanBP2 , 1997, Cell.

[32]  I. Macara,et al.  Ran Binding Domains Promote the Interaction of Ran with p97/β-Karyopherin, Linking the Docking and Translocation Steps of Nuclear Import (*) , 1996, The Journal of Biological Chemistry.

[33]  P. Silver,et al.  Mutants in a yeast Ran binding protein are defective in nuclear transport. , 1995, The EMBO journal.

[34]  P. Silver,et al.  Dynamic localization of the nuclear import receptor and its interactions with transport factors , 1996, The Journal of cell biology.

[35]  F. Bischoff,et al.  Export of Importin α from the Nucleus Is Mediated by a Specific Nuclear Transport Factor , 1997, Cell.

[36]  D. Görlich,et al.  Nuclear protein import. , 1997, Current opinion in cell biology.

[37]  Erich A. Nigg,et al.  Nucleocytoplasmic transport: signals, mechanisms and regulation , 1997, Nature.

[38]  F. Bischoff,et al.  Co‐activation of RanGTPase and inhibition of GTP dissociation by Ran‐GTP binding protein RanBP1. , 1995, The EMBO journal.

[39]  A. Hopper,et al.  The yeast RNA1 gene product necessary for RNA processing is located in the cytosol and apparently excluded from the nucleus , 1990, The Journal of cell biology.

[40]  M. Powers,et al.  Nuclear Export Receptors: From Importin to Exportin , 1997, Cell.

[41]  A. Hopper,et al.  A yeast mutant which accumulates precursor tRNAs , 1978, Cell.

[42]  P. Silver,et al.  Nucleocytoplasmic transport of macromolecules , 1997, Microbiology and molecular biology reviews : MMBR.

[43]  C. Dingwall,et al.  RAN/TC4 mutants identify a common requirement for snRNP and protein import into the nucleus , 1996, The Journal of cell biology.

[44]  G. Schlenstedt Protein import into the nucleus , 1996, FEBS letters.

[45]  I. Macara,et al.  A family of proteins that stabilize the Ran/TC4 GTPase in its GTP-bound conformation. , 1994, The Journal of biological chemistry.

[46]  G. Dreyfuss,et al.  Transportin: nuclear transport receptor of a novel nuclear protein import pathway. , 1996, Experimental cell research.

[47]  F. Bischoff,et al.  Ran-binding protein 5 (RanBP5) is related to the nuclear transport factor importin-beta but interacts differently with RanBP1 , 1997, Molecular and cellular biology.

[48]  P. Silver,et al.  A GTPase Controlling Nuclear Trafficking: Running the Right Way or Walking RANdomly? , 1996, Cell.

[49]  G. Blobel,et al.  The Nuclear Transport Factor Karyopherin Binds Stoichiometrically to Ran-GTP and Inhibits the Ran GTPase Activating Protein (*) , 1996, The Journal of Biological Chemistry.

[50]  C. Klebe,et al.  Functional expression in Escherichia coli of the mitotic regulator proteins p24ran and p45rcc1 and fluorescence measurements of their interaction. , 1993, Biochemistry.

[51]  E. Lund,et al.  Diverse effects of the guanine nucleotide exchange factor RCC1 on RNA transport. , 1995, Science.

[52]  A. Wittinghofer,et al.  Dynamic and equilibrium studies on the interaction of Ran with its effector, RanBP1. , 1997, Biochemistry.

[53]  F. Bischoff,et al.  Human RanGTPase-activating protein RanGAP1 is a homologue of yeast Rna1p involved in mRNA processing and transport. , 1995, Proceedings of the National Academy of Sciences of the United States of America.

[54]  F. Bischoff,et al.  Dominant‐negative mutants of importin‐β block multiple pathways of import and export through the nuclear pore complex , 1997, The EMBO journal.

[55]  Minoru Yoshida,et al.  CRM1 Is an Export Receptor for Leucine-Rich Nuclear Export Signals , 1997, Cell.