• We are sorry, but NCBI web applications do not support your browser and may not function properly. More information
Logo of embojLink to Publisher's site
EMBO J. Nov 3, 1997; 16(21): 6535–6547.
PMCID: PMC1170258

The asymmetric distribution of the constituents of the Ran system is essential for transport into and out of the nucleus.

Abstract

The GTPase Ran is essential for nuclear import of proteins with a classical nuclear localization signal (NLS). Ran's nucleotide-bound state is determined by the chromatin-bound exchange factor RCC1 generating RanGTP in the nucleus and the cytoplasmic GTPase activating protein RanGAP1 depleting RanGTP from the cytoplasm. This predicts a steep RanGTP concentration gradient across the nuclear envelope. RanGTP binding to importin-beta has previously been shown to release importin-alpha from -beta during NLS import. We show that RanGTP also induces release of the M9 signal from the second identified import receptor, transportin. The role of RanGTP distribution is further studied using three methods to collapse the RanGTP gradient. Nuclear injection of either RanGAP1, the RanGTP binding protein RanBP1 or a Ran mutant that cannot stably bind GTP. These treatments block major export and import pathways across the nuclear envelope. Different export pathways exhibit distinct sensitivities to RanGTP depletion, but all are more readily inhibited than is import of either NLS or M9 proteins, indicating that the block of export is direct rather than a secondary consequence of import inhibition. Surprisingly, nuclear export of several substrates including importin-alpha and -beta, transportin, HIV Rev and tRNA appears to require nuclear RanGTP but may not require GTP hydrolysis by Ran, suggesting that the energy for their nuclear export is supplied by another source.

Full Text

The Full Text of this article is available as a PDF (508K).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.
  • Aitchison JD, Blobel G, Rout MP. Kap104p: a karyopherin involved in the nuclear transport of messenger RNA binding proteins. Science. 1996 Oct 25;274(5287):624–627. [PubMed]
  • Amberg DC, Fleischmann M, Stagljar I, Cole CN, Aebi M. Nuclear PRP20 protein is required for mRNA export. EMBO J. 1993 Jan;12(1):233–241. [PMC free article] [PubMed]
  • Beddow AL, Richards SA, Orem NR, Macara IG. The Ran/TC4 GTPase-binding domain: identification by expression cloning and characterization of a conserved sequence motif. Proc Natl Acad Sci U S A. 1995 Apr 11;92(8):3328–3332. [PMC free article] [PubMed]
  • Bennett M, Piñol-Roma S, Staknis D, Dreyfuss G, Reed R. Differential binding of heterogeneous nuclear ribonucleoproteins to mRNA precursors prior to spliceosome assembly in vitro. Mol Cell Biol. 1992 Jul;12(7):3165–3175. [PMC free article] [PubMed]
  • Bischoff FR, Ponstingl H. Mitotic regulator protein RCC1 is complexed with a nuclear ras-related polypeptide. Proc Natl Acad Sci U S A. 1991 Dec 1;88(23):10830–10834. [PMC free article] [PubMed]
  • Bischoff FR, Ponstingl H. Catalysis of guanine nucleotide exchange on Ran by the mitotic regulator RCC1. Nature. 1991 Nov 7;354(6348):80–82. [PubMed]
  • Bischoff FR, Klebe C, Kretschmer J, Wittinghofer A, Ponstingl H. RanGAP1 induces GTPase activity of nuclear Ras-related Ran. Proc Natl Acad Sci U S A. 1994 Mar 29;91(7):2587–2591. [PMC free article] [PubMed]
  • Bischoff FR, Krebber H, Kempf T, Hermes I, Ponstingl H. Human RanGTPase-activating protein RanGAP1 is a homologue of yeast Rna1p involved in mRNA processing and transport. Proc Natl Acad Sci U S A. 1995 Feb 28;92(5):1749–1753. [PMC free article] [PubMed]
  • Bischoff FR, Krebber H, Smirnova E, Dong W, Ponstingl H. Co-activation of RanGTPase and inhibition of GTP dissociation by Ran-GTP binding protein RanBP1. EMBO J. 1995 Feb 15;14(4):705–715. [PMC free article] [PubMed]
  • Bogerd HP, Fridell RA, Madore S, Cullen BR. Identification of a novel cellular cofactor for the Rev/Rex class of retroviral regulatory proteins. Cell. 1995 Aug 11;82(3):485–494. [PubMed]
  • Cheng Y, Dahlberg JE, Lund E. Diverse effects of the guanine nucleotide exchange factor RCC1 on RNA transport. Science. 1995 Mar 24;267(5205):1807–1810. [PubMed]
  • Chi NC, Adam EJ, Visser GD, Adam SA. RanBP1 stabilizes the interaction of Ran with p97 nuclear protein import. J Cell Biol. 1996 Nov;135(3):559–569. [PMC free article] [PubMed]
  • Coutavas E, Ren M, Oppenheim JD, D'Eustachio P, Rush MG. Characterization of proteins that interact with the cell-cycle regulatory protein Ran/TC4. Nature. 1993 Dec 9;366(6455):585–587. [PubMed]
  • Darzynkiewicz E, Stepinski J, Ekiel I, Jin Y, Haber D, Sijuwade T, Tahara SM. Beta-globin mRNAs capped with m7G, m2.7(2)G or m2.2.7(3)G differ in intrinsic translation efficiency. Nucleic Acids Res. 1988 Sep 26;16(18):8953–8962. [PMC free article] [PubMed]
  • Dreyfuss G, Matunis MJ, Piñol-Roma S, Burd CG. hnRNP proteins and the biogenesis of mRNA. Annu Rev Biochem. 1993;62:289–321. [PubMed]
  • Fischer U, Meyer S, Teufel M, Heckel C, Lührmann R, Rautmann G. Evidence that HIV-1 Rev directly promotes the nuclear export of unspliced RNA. EMBO J. 1994 Sep 1;13(17):4105–4112. [PMC free article] [PubMed]
  • Fischer U, Huber J, Boelens WC, Mattaj IW, Lührmann R. The HIV-1 Rev activation domain is a nuclear export signal that accesses an export pathway used by specific cellular RNAs. Cell. 1995 Aug 11;82(3):475–483. [PubMed]
  • Fornerod M, van Deursen J, van Baal S, Reynolds A, Davis D, Murti KG, Fransen J, Grosveld G. The human homologue of yeast CRM1 is in a dynamic subcomplex with CAN/Nup214 and a novel nuclear pore component Nup88. EMBO J. 1997 Feb 17;16(4):807–816. [PMC free article] [PubMed]
  • Forrester W, Stutz F, Rosbash M, Wickens M. Defects in mRNA 3'-end formation, transcription initiation, and mRNA transport associated with the yeast mutation prp20: possible coupling of mRNA processing and chromatin structure. Genes Dev. 1992 Oct;6(10):1914–1926. [PubMed]
  • Fridell RA, Truant R, Thorne L, Benson RE, Cullen BR. Nuclear import of hnRNP A1 is mediated by a novel cellular cofactor related to karyopherin-beta. J Cell Sci. 1997 Jun;110(Pt 11):1325–1331. [PubMed]
  • Fritz CC, Zapp ML, Green MR. A human nucleoporin-like protein that specifically interacts with HIV Rev. Nature. 1995 Aug 10;376(6540):530–533. [PubMed]
  • Gerace L. Nuclear export signals and the fast track to the cytoplasm. Cell. 1995 Aug 11;82(3):341–344. [PubMed]
  • Görlich D, Mattaj IW. Nucleocytoplasmic transport. Science. 1996 Mar 15;271(5255):1513–1518. [PubMed]
  • Görlich D, Prehn S, Laskey RA, Hartmann E. Isolation of a protein that is essential for the first step of nuclear protein import. Cell. 1994 Dec 2;79(5):767–778. [PubMed]
  • Görlich D, Panté N, Kutay U, Aebi U, Bischoff FR. Identification of different roles for RanGDP and RanGTP in nuclear protein import. EMBO J. 1996 Oct 15;15(20):5584–5594. [PMC free article] [PubMed]
  • Görlich D, Kraft R, Kostka S, Vogel F, Hartmann E, Laskey RA, Mattaj IW, Izaurralde E. Importin provides a link between nuclear protein import and U snRNA export. Cell. 1996 Oct 4;87(1):21–32. [PubMed]
  • Görlich D, Dabrowski M, Bischoff FR, Kutay U, Bork P, Hartmann E, Prehn S, Izaurralde E. A novel class of RanGTP binding proteins. J Cell Biol. 1997 Jul 14;138(1):65–80. [PMC free article] [PubMed]
  • Hamm J, Mattaj IW. Monomethylated cap structures facilitate RNA export from the nucleus. Cell. 1990 Oct 5;63(1):109–118. [PubMed]
  • Hamm J, Dathan NA, Mattaj IW. Functional analysis of mutant Xenopus U2 snRNAs. Cell. 1989 Oct 6;59(1):159–169. [PubMed]
  • Hayashi N, Yokoyama N, Seki T, Azuma Y, Ohba T, Nishimoto T. RanBP1, a Ras-like nuclear G protein binding to Ran/TC4, inhibits RCC1 via Ran/TC4. Mol Gen Genet. 1995 Jun 25;247(6):661–669. [PubMed]
  • Her LS, Lund E, Dahlberg JE. Inhibition of Ran guanosine triphosphatase-dependent nuclear transport by the matrix protein of vesicular stomatitis virus. Science. 1997 Jun 20;276(5320):1845–1848. [PubMed]
  • Hopper AK, Traglia HM, Dunst RW. The yeast RNA1 gene product necessary for RNA processing is located in the cytosol and apparently excluded from the nucleus. J Cell Biol. 1990 Aug;111(2):309–321. [PMC free article] [PubMed]
  • Izaurralde E, Mattaj IW. RNA export. Cell. 1995 Apr 21;81(2):153–159. [PubMed]
  • Izaurralde E, Lewis J, McGuigan C, Jankowska M, Darzynkiewicz E, Mattaj IW. A nuclear cap binding protein complex involved in pre-mRNA splicing. Cell. 1994 Aug 26;78(4):657–668. [PubMed]
  • Izaurralde E, Lewis J, Gamberi C, Jarmolowski A, McGuigan C, Mattaj IW. A cap-binding protein complex mediating U snRNA export. Nature. 1995 Aug 24;376(6542):709–712. [PubMed]
  • Izaurralde E, Jarmolowski A, Beisel C, Mattaj IW, Dreyfuss G, Fischer U. A role for the M9 transport signal of hnRNP A1 in mRNA nuclear export. J Cell Biol. 1997 Apr 7;137(1):27–35. [PMC free article] [PubMed]
  • Jarmolowski A, Boelens WC, Izaurralde E, Mattaj IW. Nuclear export of different classes of RNA is mediated by specific factors. J Cell Biol. 1994 Mar;124(5):627–635. [PMC free article] [PubMed]
  • Kadowaki T, Goldfarb D, Spitz LM, Tartakoff AM, Ohno M. Regulation of RNA processing and transport by a nuclear guanine nucleotide release protein and members of the Ras superfamily. EMBO J. 1993 Jul;12(7):2929–2937. [PMC free article] [PubMed]
  • Kadowaki T, Chen S, Hitomi M, Jacobs E, Kumagai C, Liang S, Schneiter R, Singleton D, Wisniewska J, Tartakoff AM. Isolation and characterization of Saccharomyces cerevisiae mRNA transport-defective (mtr) mutants. J Cell Biol. 1994 Aug;126(3):649–659. [PMC free article] [PubMed]
  • Kambach C, Mattaj IW. Intracellular distribution of the U1A protein depends on active transport and nuclear binding to U1 snRNA. J Cell Biol. 1992 Jul;118(1):11–21. [PMC free article] [PubMed]
  • Klebe C, Bischoff FR, Ponstingl H, Wittinghofer A. Interaction of the nuclear GTP-binding protein Ran with its regulatory proteins RCC1 and RanGAP1. Biochemistry. 1995 Jan 17;34(2):639–647. [PubMed]
  • Koepp DM, Silver PA. A GTPase controlling nuclear trafficking: running the right way or walking RANdomly? Cell. 1996 Oct 4;87(1):1–4. [PubMed]
  • Koepp DM, Wong DH, Corbett AH, Silver PA. Dynamic localization of the nuclear import receptor and its interactions with transport factors. J Cell Biol. 1996 Jun;133(6):1163–1176. [PMC free article] [PubMed]
  • Kutay U, Izaurralde E, Bischoff FR, Mattaj IW, Görlich D. Dominant-negative mutants of importin-beta block multiple pathways of import and export through the nuclear pore complex. EMBO J. 1997 Mar 17;16(6):1153–1163. [PMC free article] [PubMed]
  • Lounsbury KM, Macara IG. Ran-binding protein 1 (RanBP1) forms a ternary complex with Ran and karyopherin beta and reduces Ran GTPase-activating protein (RanGAP) inhibition by karyopherin beta. J Biol Chem. 1997 Jan 3;272(1):551–555. [PubMed]
  • Mahajan R, Delphin C, Guan T, Gerace L, Melchior F. A small ubiquitin-related polypeptide involved in targeting RanGAP1 to nuclear pore complex protein RanBP2. Cell. 1997 Jan 10;88(1):97–107. [PubMed]
  • Matunis EL, Matunis MJ, Dreyfuss G. Association of individual hnRNP proteins and snRNPs with nascent transcripts. J Cell Biol. 1993 Apr;121(2):219–228. [PMC free article] [PubMed]
  • Matunis MJ, Coutavas E, Blobel G. A novel ubiquitin-like modification modulates the partitioning of the Ran-GTPase-activating protein RanGAP1 between the cytosol and the nuclear pore complex. J Cell Biol. 1996 Dec;135(6 Pt 1):1457–1470. [PMC free article] [PubMed]
  • Melchior F, Paschal B, Evans J, Gerace L. Inhibition of nuclear protein import by nonhydrolyzable analogues of GTP and identification of the small GTPase Ran/TC4 as an essential transport factor. J Cell Biol. 1993 Dec;123(6 Pt 2):1649–1659. [PMC free article] [PubMed]
  • Melchior F, Weber K, Gerke V. A functional homologue of the RNA1 gene product in Schizosaccharomyces pombe: purification, biochemical characterization, and identification of a leucine-rich repeat motif. Mol Biol Cell. 1993 Jun;4(6):569–581. [PMC free article] [PubMed]
  • Michael WM, Choi M, Dreyfuss G. A nuclear export signal in hnRNP A1: a signal-mediated, temperature-dependent nuclear protein export pathway. Cell. 1995 Nov 3;83(3):415–422. [PubMed]
  • Michael WM, Eder PS, Dreyfuss G. The K nuclear shuttling domain: a novel signal for nuclear import and nuclear export in the hnRNP K protein. EMBO J. 1997 Jun 16;16(12):3587–3598. [PMC free article] [PubMed]
  • Moore MS, Blobel G. The GTP-binding protein Ran/TC4 is required for protein import into the nucleus. Nature. 1993 Oct 14;365(6447):661–663. [PubMed]
  • Munro S, Pelham HR. Use of peptide tagging to detect proteins expressed from cloned genes: deletion mapping functional domains of Drosophila hsp 70. EMBO J. 1984 Dec 20;3(13):3087–3093. [PMC free article] [PubMed]
  • Nakielny S, Siomi MC, Siomi H, Michael WM, Pollard V, Dreyfuss G. Transportin: nuclear transport receptor of a novel nuclear protein import pathway. Exp Cell Res. 1996 Dec 15;229(2):261–266. [PubMed]
  • Nigg EA. Nucleocytoplasmic transport: signals, mechanisms and regulation. Nature. 1997 Apr 24;386(6627):779–787. [PubMed]
  • Ohtsubo M, Okazaki H, Nishimoto T. The RCC1 protein, a regulator for the onset of chromosome condensation locates in the nucleus and binds to DNA. J Cell Biol. 1989 Oct;109(4 Pt 1):1389–1397. [PMC free article] [PubMed]
  • Palacios I, Weis K, Klebe C, Mattaj IW, Dingwall C. RAN/TC4 mutants identify a common requirement for snRNP and protein import into the nucleus. J Cell Biol. 1996 May;133(3):485–494. [PMC free article] [PubMed]
  • Panté N, Aebi U. Toward the molecular dissection of protein import into nuclei. Curr Opin Cell Biol. 1996 Jun;8(3):397–406. [PubMed]
  • Pollard VW, Michael WM, Nakielny S, Siomi MC, Wang F, Dreyfuss G. A novel receptor-mediated nuclear protein import pathway. Cell. 1996 Sep 20;86(6):985–994. [PubMed]
  • Rexach M, Blobel G. Protein import into nuclei: association and dissociation reactions involving transport substrate, transport factors, and nucleoporins. Cell. 1995 Dec 1;83(5):683–692. [PubMed]
  • Richards SA, Lounsbury KM, Carey KL, Macara IG. A nuclear export signal is essential for the cytosolic localization of the Ran binding protein, RanBP1. J Cell Biol. 1996 Sep;134(5):1157–1168. [PMC free article] [PubMed]
  • Richards SA, Carey KL, Macara IG. Requirement of guanosine triphosphate-bound ran for signal-mediated nuclear protein export. Science. 1997 Jun 20;276(5320):1842–1844. [PubMed]
  • Saavedra C, Tung KS, Amberg DC, Hopper AK, Cole CN. Regulation of mRNA export in response to stress in Saccharomyces cerevisiae. Genes Dev. 1996 Jul 1;10(13):1608–1620. [PubMed]
  • Saavedra C, Felber B, Izaurralde E. The simian retrovirus-1 constitutive transport element, unlike the HIV-1 RRE, uses factors required for cellular mRNA export. Curr Biol. 1997 Sep 1;7(9):619–628. [PubMed]
  • Schlenstedt G. Protein import into the nucleus. FEBS Lett. 1996 Jun 24;389(1):75–79. [PubMed]
  • Schlenstedt G, Saavedra C, Loeb JD, Cole CN, Silver PA. The GTP-bound form of the yeast Ran/TC4 homologue blocks nuclear protein import and appearance of poly(A)+ RNA in the cytoplasm. Proc Natl Acad Sci U S A. 1995 Jan 3;92(1):225–229. [PMC free article] [PubMed]
  • Siomi H, Dreyfuss G. A nuclear localization domain in the hnRNP A1 protein. J Cell Biol. 1995 May;129(3):551–560. [PMC free article] [PubMed]
  • Stutz F, Neville M, Rosbash M. Identification of a novel nuclear pore-associated protein as a functional target of the HIV-1 Rev protein in yeast. Cell. 1995 Aug 11;82(3):495–506. [PubMed]
  • Traglia HM, Atkinson NS, Hopper AK. Structural and functional analyses of Saccharomyces cerevisiae wild-type and mutant RNA1 genes. Mol Cell Biol. 1989 Jul;9(7):2989–2999. [PMC free article] [PubMed]
  • Weighardt F, Biamonti G, Riva S. Nucleo-cytoplasmic distribution of human hnRNP proteins: a search for the targeting domains in hnRNP A1. J Cell Sci. 1995 Feb;108(Pt 2):545–555. [PubMed]
  • Weis K, Dingwall C, Lamond AI. Characterization of the nuclear protein import mechanism using Ran mutants with altered nucleotide binding specificities. EMBO J. 1996 Dec 16;15(24):7120–7128. [PMC free article] [PubMed]
  • Wen W, Meinkoth JL, Tsien RY, Taylor SS. Identification of a signal for rapid export of proteins from the nucleus. Cell. 1995 Aug 11;82(3):463–473. [PubMed]
  • Wu J, Matunis MJ, Kraemer D, Blobel G, Coutavas E. Nup358, a cytoplasmically exposed nucleoporin with peptide repeats, Ran-GTP binding sites, zinc fingers, a cyclophilin A homologous domain, and a leucine-rich region. J Biol Chem. 1995 Jun 9;270(23):14209–14213. [PubMed]
  • Yokoyama N, Hayashi N, Seki T, Panté N, Ohba T, Nishii K, Kuma K, Hayashida T, Miyata T, Aebi U, et al. A giant nucleopore protein that binds Ran/TC4. Nature. 1995 Jul 13;376(6536):184–188. [PubMed]
  • Zasloff M. tRNA transport from the nucleus in a eukaryotic cell: carrier-mediated translocation process. Proc Natl Acad Sci U S A. 1983 Nov;80(21):6436–6440. [PMC free article] [PubMed]
  • Zolotukhin AS, Felber BK. Mutations in the nuclear export signal of human ran-binding protein RanBP1 block the Rev-mediated posttranscriptional regulation of human immunodeficiency virus type 1. J Biol Chem. 1997 Apr 25;272(17):11356–11360. [PubMed]

Articles from The EMBO Journal are provided here courtesy of The European Molecular Biology Organization

Formats:

Related citations in PubMed

See reviews...See all...

Cited by other articles in PMC

See all...

Links

  • Compound
    Compound
    PubChem Compound links
  • PubMed
    PubMed
    PubMed citations for these articles
  • Substance
    Substance
    PubChem Substance links

Recent Activity

Your browsing activity is empty.

Activity recording is turned off.

Turn recording back on

See more...