Skip to main content
PMC home page
Biochemical Journal logoLink to Biochemical Journal
. 2000 Feb 15;346(Pt 1):107–115.

Export and transport of tRNA are coupled to a multi-protein complex.

C Kruse 1, D K Willkomm 1, A Grünweller 1, T Vollbrandt 1, S Sommer 1, S Busch 1, T Pfeiffer 1, J Brinkmann 1, R K Hartmann 1, P K Müller 1
PMCID: PMC1220829  PMID: 10657246

Abstract

Vigilin is a ubiquitous multi heterogeneous nuclear ribonucleoprotein (hnRNP) K homologous (KH)-domain protein. Here we demonstrate that purified recombinant human vigilin binds tRNA molecules with high affinity, although with limited specificity. Nuclear microinjection experiments revealed for the first time that the immuno-affinity-purified nuclear vigilin core complex (VCC(N)) as well as recombinant vigilin accelerate tRNA export from the nucleus in human cells. The nuclear tRNA receptor exportin-t is part of the VCC(N). Elongation factor (EF)-1alpha is enriched in VCC(N) and its cytoplasmic counterpart VCC(C), whereas EF-1beta, EF-1gamma and EF-1delta are basically confined to the VCC(C). Our results suggest further that vigilin and exportin-t might interact during tRNA export, provide evidence that the channeled tRNA cycle is already initiated in the nucleus, and illustrate that intracellular tRNA trafficking is associated with discrete changes in the composition of cellular cytoplasmic multi-protein complexes containing tRNA.

Full Text

The Full Text of this article is available as a PDF (283.5 KB).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Arts G. J., Fornerod M., Mattaj I. W. Identification of a nuclear export receptor for tRNA. Curr Biol. 1998 Mar 12;8(6):305–314. doi: 10.1016/s0960-9822(98)70130-7. [DOI] [PubMed] [Google Scholar]
  2. Arts G. J., Kuersten S., Romby P., Ehresmann B., Mattaj I. W. The role of exportin-t in selective nuclear export of mature tRNAs. EMBO J. 1998 Dec 15;17(24):7430–7441. doi: 10.1093/emboj/17.24.7430. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Aumailley M., Krieg T., Razaka G., Müller P. K., Bricaud H. Influence of cell density on collagen biosynthesis in fibroblast cultures. Biochem J. 1982 Sep 15;206(3):505–510. doi: 10.1042/bj2060505. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Boelens W. C., Palacios I., Mattaj I. W. Nuclear retention of RNA as a mechanism for localization. RNA. 1995 May;1(3):273–283. [PMC free article] [PubMed] [Google Scholar]
  5. Butler R. N., August P., Ferdinand K. C., Phillips R. A., Roccella E. J. Hypertension: setting new goals for lower readings. Geriatrics. 1999 Apr;54(4):20-1, 25-6, 29-30 passim. [PubMed] [Google Scholar]
  6. Cameron V., Uhlenbeck O. C. 3'-Phosphatase activity in T4 polynucleotide kinase. Biochemistry. 1977 Nov 15;16(23):5120–5126. doi: 10.1021/bi00642a027. [DOI] [PubMed] [Google Scholar]
  7. Cheng Y., Dahlberg J. E., Lund E. Diverse effects of the guanine nucleotide exchange factor RCC1 on RNA transport. Science. 1995 Mar 24;267(5205):1807–1810. doi: 10.1126/science.7534442. [DOI] [PubMed] [Google Scholar]
  8. Chiu D. S., Oram J. F., LeBoeuf R. C., Alpers C. E., O'Brien K. D. High-density lipoprotein-binding protein (HBP)/vigilin is expressed in human atherosclerotic lesions and colocalizes with apolipoprotein E. Arterioscler Thromb Vasc Biol. 1997 Nov;17(11):2350–2358. doi: 10.1161/01.atv.17.11.2350. [DOI] [PubMed] [Google Scholar]
  9. Dahlberg J. E., Lund E. Functions of the GTPase Ran in RNA export from the nucleus. Curr Opin Cell Biol. 1998 Jun;10(3):400–408. doi: 10.1016/s0955-0674(98)80017-3. [DOI] [PubMed] [Google Scholar]
  10. Dejgaard K., Leffers H. Characterisation of the nucleic-acid-binding activity of KH domains. Different properties of different domains. Eur J Biochem. 1996 Oct 15;241(2):425–431. doi: 10.1111/j.1432-1033.1996.00425.x. [DOI] [PubMed] [Google Scholar]
  11. Dignam J. D., Lebovitz R. M., Roeder R. G. Accurate transcription initiation by RNA polymerase II in a soluble extract from isolated mammalian nuclei. Nucleic Acids Res. 1983 Mar 11;11(5):1475–1489. doi: 10.1093/nar/11.5.1475. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Dodson R. E., Shapiro D. J. An estrogen-inducible protein binds specifically to a sequence in the 3' untranslated region of estrogen-stabilized vitellogenin mRNA. Mol Cell Biol. 1994 May;14(5):3130–3138. doi: 10.1128/mcb.14.5.3130. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Dodson R. E., Shapiro D. J. Vigilin, a ubiquitous protein with 14 K homology domains, is the estrogen-inducible vitellogenin mRNA 3'-untranslated region-binding protein. J Biol Chem. 1997 May 9;272(19):12249–12252. doi: 10.1074/jbc.272.19.12249. [DOI] [PubMed] [Google Scholar]
  14. Görlich D., Kraft R., Kostka S., Vogel F., Hartmann E., Laskey R. A., Mattaj I. W., Izaurralde E. Importin provides a link between nuclear protein import and U snRNA export. Cell. 1996 Oct 4;87(1):21–32. doi: 10.1016/s0092-8674(00)81319-7. [DOI] [PubMed] [Google Scholar]
  15. Hamm J., Kazmaier M., Mattaj I. W. In vitro assembly of U1 snRNPs. EMBO J. 1987 Nov;6(11):3479–3485. doi: 10.1002/j.1460-2075.1987.tb02672.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Hamm J., Mattaj I. W. An abundant U6 snRNP found in germ cells and embryos of Xenopus laevis. EMBO J. 1989 Dec 20;8(13):4179–4187. doi: 10.1002/j.1460-2075.1989.tb08603.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Harmon S. A., Richards O. C., Summers D. F., Ehrenfeld E. The 5'-terminal nucleotides of hepatitis A virus RNA, but not poliovirus RNA, are required for infectivity. J Virol. 1991 May;65(5):2757–2760. doi: 10.1128/jvi.65.5.2757-2760.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Hatzfeld M., Maier G., Franke W. W. Cytokeratin domains involved in heterotypic complex formation determined by in-vitro binding assays. J Mol Biol. 1987 Sep 20;197(2):237–255. doi: 10.1016/0022-2836(87)90122-7. [DOI] [PubMed] [Google Scholar]
  19. Hurt D. J., Wang S. S., Lin Y. H., Hopper A. K. Cloning and characterization of LOS1, a Saccharomyces cerevisiae gene that affects tRNA splicing. Mol Cell Biol. 1987 Mar;7(3):1208–1216. doi: 10.1128/mcb.7.3.1208. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Izaurralde E., Kutay U., von Kobbe C., Mattaj I. W., Görlich D. The asymmetric distribution of the constituents of the Ran system is essential for transport into and out of the nucleus. EMBO J. 1997 Nov 3;16(21):6535–6547. doi: 10.1093/emboj/16.21.6535. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Jarmolowski A., Boelens W. C., Izaurralde E., Mattaj I. W. Nuclear export of different classes of RNA is mediated by specific factors. J Cell Biol. 1994 Mar;124(5):627–635. doi: 10.1083/jcb.124.5.627. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Kanamori H., Dodson R. E., Shapiro D. J. In vitro genetic analysis of the RNA binding site of vigilin, a multi-KH-domain protein. Mol Cell Biol. 1998 Jul;18(7):3991–4003. doi: 10.1128/mcb.18.7.3991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Kruse C., Emmrich J., Rumpel E., Klinger M. H., Grünweller A., Rohwedel J., Krammer H. J., Kühnel W., Müller P. K. Production of trypsin by cells of the exocrine pancreas is paralleled by the expression of the KH protein vigilin. Exp Cell Res. 1998 Feb 25;239(1):111–118. doi: 10.1006/excr.1997.3891. [DOI] [PubMed] [Google Scholar]
  24. Kruse C., Grünweller A., Notbohm H., Kügler S., Purschke W. G., Müller P. K. Evidence for a novel cytoplasmic tRNA-protein complex containing the KH-multidomain protein vigilin. Biochem J. 1996 Nov 15;320(Pt 1):247–252. doi: 10.1042/bj3200247. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Kruse C., Grünweller A., Willkomm D. K., Pfeiffer T., Hartmann R. K., Müller P. K. tRNA is entrapped in similar, but distinct, nuclear and cytoplasmic ribonucleoprotein complexes, both of which contain vigilin and elongation factor 1 alpha. Biochem J. 1998 Feb 1;329(Pt 3):615–621. doi: 10.1042/bj3290615. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Kutay U., Lipowsky G., Izaurralde E., Bischoff F. R., Schwarzmaier P., Hartmann E., Görlich D. Identification of a tRNA-specific nuclear export receptor. Mol Cell. 1998 Feb;1(3):359–369. doi: 10.1016/s1097-2765(00)80036-2. [DOI] [PubMed] [Google Scholar]
  27. Kügler S., Grünweller A., Probst C., Klinger M., Müller P. K., Kruse C. Vigilin contains a functional nuclear localisation sequence and is present in both the cytoplasm and the nucleus. FEBS Lett. 1996 Mar 18;382(3):330–334. doi: 10.1016/0014-5793(96)00204-9. [DOI] [PubMed] [Google Scholar]
  28. Lipowsky G., Bischoff F. R., Izaurralde E., Kutay U., Schäfer S., Gross H. J., Beier H., Görlich D. Coordination of tRNA nuclear export with processing of tRNA. RNA. 1999 Apr;5(4):539–549. doi: 10.1017/s1355838299982134. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Lund E., Dahlberg J. E. Proofreading and aminoacylation of tRNAs before export from the nucleus. Science. 1998 Dec 11;282(5396):2082–2085. doi: 10.1126/science.282.5396.2082. [DOI] [PubMed] [Google Scholar]
  30. Musco G., Stier G., Joseph C., Castiglione Morelli M. A., Nilges M., Gibson T. J., Pastore A. Three-dimensional structure and stability of the KH domain: molecular insights into the fragile X syndrome. Cell. 1996 Apr 19;85(2):237–245. doi: 10.1016/s0092-8674(00)81100-9. [DOI] [PubMed] [Google Scholar]
  31. Nakielny S., Dreyfuss G. Import and export of the nuclear protein import receptor transportin by a mechanism independent of GTP hydrolysis. Curr Biol. 1998 Jan 15;8(2):89–95. doi: 10.1016/s0960-9822(98)70039-9. [DOI] [PubMed] [Google Scholar]
  32. Negrutskii B. S., Deutscher M. P. Channeling of aminoacyl-tRNA for protein synthesis in vivo. Proc Natl Acad Sci U S A. 1991 Jun 1;88(11):4991–4995. doi: 10.1073/pnas.88.11.4991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Ohno M., Fornerod M., Mattaj I. W. Nucleocytoplasmic transport: the last 200 nanometers. Cell. 1998 Feb 6;92(3):327–336. doi: 10.1016/s0092-8674(00)80926-5. [DOI] [PubMed] [Google Scholar]
  34. Pennisi E. The nucleus's revolving door. Science. 1998 Feb 20;279(5354):1129–1131. doi: 10.1126/science.279.5354.1129. [DOI] [PubMed] [Google Scholar]
  35. Perret V., Garcia A., Puglisi J., Grosjean H., Ebel J. P., Florentz C., Giegé R. Conformation in solution of yeast tRNA(Asp) transcripts deprived of modified nucleotides. Biochimie. 1990 Oct;72(10):735–743. doi: 10.1016/0300-9084(90)90158-d. [DOI] [PubMed] [Google Scholar]
  36. Raleigh E. A., Wilson G. Escherichia coli K-12 restricts DNA containing 5-methylcytosine. Proc Natl Acad Sci U S A. 1986 Dec;83(23):9070–9074. doi: 10.1073/pnas.83.23.9070. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Rumpel E., Kruse C., Müller P. K., Kühnel W. Expression of vigilin in the uterus of ovariectomized steroid-treated rats and during the estrous cycle. Ann Anat. 1996 Aug;178(4):337–344. doi: 10.1016/S0940-9602(96)80089-6. [DOI] [PubMed] [Google Scholar]
  38. Schlegl J., Fürste J. P., Bald R., Erdmann V. A., Hartmann R. K. Cleavage efficiencies of model substrates for ribonuclease P from Escherichia coli and Thermus thermophilus. Nucleic Acids Res. 1992 Nov 25;20(22):5963–5970. doi: 10.1093/nar/20.22.5963. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Stapulionis R., Deutscher M. P. A channeled tRNA cycle during mammalian protein synthesis. Proc Natl Acad Sci U S A. 1995 Aug 1;92(16):7158–7161. doi: 10.1073/pnas.92.16.7158. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Weis K. Importins and exportins: how to get in and out of the nucleus. Trends Biochem Sci. 1998 May;23(5):185–189. doi: 10.1016/s0968-0004(98)01204-3. [DOI] [PubMed] [Google Scholar]
  41. Weiss S., König B., Müller H. J., Seidel H., Goody R. S. Synthetic human tRNA(UUULys3) and natural bovine tRNA(UUULys3) interact with HIV-1 reverse transcriptase and serve as specific primers for retroviral cDNA synthesis. Gene. 1992 Feb 15;111(2):183–197. doi: 10.1016/0378-1119(92)90686-j. [DOI] [PubMed] [Google Scholar]
  42. Wintersberger U., Kühne C., Karwan A. Scp160p, a new yeast protein associated with the nuclear membrane and the endoplasmic reticulum, is necessary for maintenance of exact ploidy. Yeast. 1995 Aug;11(10):929–944. doi: 10.1002/yea.320111004. [DOI] [PubMed] [Google Scholar]
  43. 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. doi: 10.1073/pnas.80.21.6436. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Biochemical Journal are provided here courtesy of The Biochemical Society

RESOURCES