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. 2000 Dec;6(12):1737–1749. doi: 10.1017/s1355838200001059

Pre-mRNA processing factors are required for nuclear export.

A S Brodsky 1, P A Silver 1
PMCID: PMC1370044  PMID: 11142374

Abstract

RNA export from the nucleus is thought to be linked to proper processing and packaging into ribonucleoprotein protein complexes. A system to observe mRNA nuclear export in living yeast cells was developed by fusing the U1A RNA-binding protein to the green fluorescent protein to follow specific mRNAs with U1A hairpins engineered into them. RNAs encoding Rpl25, Pgk1, and Ssa4 were examined for the effects of 3' UTRs, introns, RNA processing factors, nucleoporins, and transport factors on their export. All accumulated in the nucleus in mutants affecting components of the nuclear export machinery and certain nucleoporins. However, under conditions of stress, PGK1 and RPL25 transcripts accumulate in the nucleus whereas SSA4 RNA is exported. Moreover, when export is blocked, only RNAs containing the ASH1 3' UTR accumulated in the nucleolus. Mutations in the splicing machinery selectively blocked export of only intron-containing RNAs. Mutations in RNA14, RNA15, and PAP1, which encode factors important for 3' processing, also blocked export of all RNAs, including SSA4, thereby linking export to the process of polyadenlyation. Taken together, these data graphically display the connections between mRNA processing and nuclear export.

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Selected References

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  1. Aebi M., Clark M. W., Vijayraghavan U., Abelson J. A yeast mutant, PRP20, altered in mRNA metabolism and maintenance of the nuclear structure, is defective in a gene homologous to the human gene RCC1 which is involved in the control of chromosome condensation. Mol Gen Genet. 1990 Oct;224(1):72–80. doi: 10.1007/BF00259453. [DOI] [PubMed] [Google Scholar]
  2. Amberg D. C., Fleischmann M., Stagljar I., Cole C. N., Aebi M. Nuclear PRP20 protein is required for mRNA export. EMBO J. 1993 Jan;12(1):233–241. doi: 10.1002/j.1460-2075.1993.tb05649.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Amberg D. C., Goldstein A. L., Cole C. N. Isolation and characterization of RAT1: an essential gene of Saccharomyces cerevisiae required for the efficient nucleocytoplasmic trafficking of mRNA. Genes Dev. 1992 Jul;6(7):1173–1189. doi: 10.1101/gad.6.7.1173. [DOI] [PubMed] [Google Scholar]
  4. Beach D. L., Salmon E. D., Bloom K. Localization and anchoring of mRNA in budding yeast. Curr Biol. 1999 Jun 3;9(11):569–578. doi: 10.1016/s0960-9822(99)80260-7. [DOI] [PubMed] [Google Scholar]
  5. Bertrand E., Chartrand P., Schaefer M., Shenoy S. M., Singer R. H., Long R. M. Localization of ASH1 mRNA particles in living yeast. Mol Cell. 1998 Oct;2(4):437–445. doi: 10.1016/s1097-2765(00)80143-4. [DOI] [PubMed] [Google Scholar]
  6. Birse C. E., Minvielle-Sebastia L., Lee B. A., Keller W., Proudfoot N. J. Coupling termination of transcription to messenger RNA maturation in yeast. Science. 1998 Apr 10;280(5361):298–301. doi: 10.1126/science.280.5361.298. [DOI] [PubMed] [Google Scholar]
  7. Chapon C., Cech T. R., Zaug A. J. Polyadenylation of telomerase RNA in budding yeast. RNA. 1997 Nov;3(11):1337–1351. [PMC free article] [PubMed] [Google Scholar]
  8. Dye M. J., Proudfoot N. J. Terminal exon definition occurs cotranscriptionally and promotes termination of RNA polymerase II. Mol Cell. 1999 Mar;3(3):371–378. doi: 10.1016/s1097-2765(00)80464-5. [DOI] [PubMed] [Google Scholar]
  9. Eckner R., Ellmeier W., Birnstiel M. L. Mature mRNA 3' end formation stimulates RNA export from the nucleus. EMBO J. 1991 Nov;10(11):3513–3522. doi: 10.1002/j.1460-2075.1991.tb04915.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Fornerod M., Ohno M., Yoshida M., Mattaj I. W. CRM1 is an export receptor for leucine-rich nuclear export signals. Cell. 1997 Sep 19;90(6):1051–1060. doi: 10.1016/s0092-8674(00)80371-2. [DOI] [PubMed] [Google Scholar]
  11. 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. doi: 10.1101/gad.6.10.1914. [DOI] [PubMed] [Google Scholar]
  12. Gallouzi I. E., Brennan C. M., Stenberg M. G., Swanson M. S., Eversole A., Maizels N., Steitz J. A. HuR binding to cytoplasmic mRNA is perturbed by heat shock. Proc Natl Acad Sci U S A. 2000 Mar 28;97(7):3073–3078. doi: 10.1073/pnas.97.7.3073. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. González C. I., Ruiz-Echevarría M. J., Vasudevan S., Henry M. F., Peltz S. W. The yeast hnRNP-like protein Hrp1/Nab4 marks a transcript for nonsense-mediated mRNA decay. Mol Cell. 2000 Mar;5(3):489–499. doi: 10.1016/s1097-2765(00)80443-8. [DOI] [PubMed] [Google Scholar]
  14. Gorsch L. C., Dockendorff T. C., Cole C. N. A conditional allele of the novel repeat-containing yeast nucleoporin RAT7/NUP159 causes both rapid cessation of mRNA export and reversible clustering of nuclear pore complexes. J Cell Biol. 1995 May;129(4):939–955. doi: 10.1083/jcb.129.4.939. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Grüter P., Tabernero C., von Kobbe C., Schmitt C., Saavedra C., Bachi A., Wilm M., Felber B. K., Izaurralde E. TAP, the human homolog of Mex67p, mediates CTE-dependent RNA export from the nucleus. Mol Cell. 1998 Apr;1(5):649–659. doi: 10.1016/s1097-2765(00)80065-9. [DOI] [PubMed] [Google Scholar]
  16. Görlich D., Kutay U. Transport between the cell nucleus and the cytoplasm. Annu Rev Cell Dev Biol. 1999;15:607–660. doi: 10.1146/annurev.cellbio.15.1.607. [DOI] [PubMed] [Google Scholar]
  17. Heath C. V., Copeland C. S., Amberg D. C., Del Priore V., Snyder M., Cole C. N. Nuclear pore complex clustering and nuclear accumulation of poly(A)+ RNA associated with mutation of the Saccharomyces cerevisiae RAT2/NUP120 gene. J Cell Biol. 1995 Dec;131(6 Pt 2):1677–1697. doi: 10.1083/jcb.131.6.1677. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Hodge C. A., Colot H. V., Stafford P., Cole C. N. Rat8p/Dbp5p is a shuttling transport factor that interacts with Rat7p/Nup159p and Gle1p and suppresses the mRNA export defect of xpo1-1 cells. EMBO J. 1999 Oct 15;18(20):5778–5788. doi: 10.1093/emboj/18.20.5778. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Hopper A. K., Traglia H. M., Dunst R. W. 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. doi: 10.1083/jcb.111.2.309. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Huang Y., Carmichael G. G. Role of polyadenylation in nucleocytoplasmic transport of mRNA. Mol Cell Biol. 1996 Apr;16(4):1534–1542. doi: 10.1128/mcb.16.4.1534. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Hurt E., Strässer K., Segref A., Bailer S., Schlaich N., Presutti C., Tollervey D., Jansen R. Mex67p mediates nuclear export of a variety of RNA polymerase II transcripts. J Biol Chem. 2000 Mar 24;275(12):8361–8368. doi: 10.1074/jbc.275.12.8361. [DOI] [PubMed] [Google Scholar]
  22. Kadowaki T., Chen S., Hitomi M., Jacobs E., Kumagai C., Liang S., Schneiter R., Singleton D., Wisniewska J., Tartakoff A. M. Isolation and characterization of Saccharomyces cerevisiae mRNA transport-defective (mtr) mutants. J Cell Biol. 1994 Aug;126(3):649–659. doi: 10.1083/jcb.126.3.649. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Kadowaki T., Schneiter R., Hitomi M., Tartakoff A. M. Mutations in nucleolar proteins lead to nucleolar accumulation of polyA+ RNA in Saccharomyces cerevisiae. Mol Biol Cell. 1995 Sep;6(9):1103–1110. doi: 10.1091/mbc.6.9.1103. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Kadowaki T., Zhao Y., Tartakoff A. M. A conditional yeast mutant deficient in mRNA transport from nucleus to cytoplasm. Proc Natl Acad Sci U S A. 1992 Mar 15;89(6):2312–2316. doi: 10.1073/pnas.89.6.2312. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Kessler M. M., Henry M. F., Shen E., Zhao J., Gross S., Silver P. A., Moore C. L. Hrp1, a sequence-specific RNA-binding protein that shuttles between the nucleus and the cytoplasm, is required for mRNA 3'-end formation in yeast. Genes Dev. 1997 Oct 1;11(19):2545–2556. doi: 10.1101/gad.11.19.2545. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Krebber H., Taura T., Lee M. S., Silver P. A. Uncoupling of the hnRNP Npl3p from mRNAs during the stress-induced block in mRNA export. Genes Dev. 1999 Aug 1;13(15):1994–2004. doi: 10.1101/gad.13.15.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. LaGrandeur T., Parker R. The cis acting sequences responsible for the differential decay of the unstable MFA2 and stable PGK1 transcripts in yeast include the context of the translational start codon. RNA. 1999 Mar;5(3):420–433. doi: 10.1017/s1355838299981748. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Legrain P., Rosbash M. Some cis- and trans-acting mutants for splicing target pre-mRNA to the cytoplasm. Cell. 1989 May 19;57(4):573–583. doi: 10.1016/0092-8674(89)90127-x. [DOI] [PubMed] [Google Scholar]
  29. Long R. M., Elliott D. J., Stutz F., Rosbash M., Singer R. H. Spatial consequences of defective processing of specific yeast mRNAs revealed by fluorescent in situ hybridization. RNA. 1995 Dec;1(10):1071–1078. [PMC free article] [PubMed] [Google Scholar]
  30. Luo M. J., Reed R. Splicing is required for rapid and efficient mRNA export in metazoans. Proc Natl Acad Sci U S A. 1999 Dec 21;96(26):14937–14942. doi: 10.1073/pnas.96.26.14937. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Mandart E., Parker R. Effects of mutations in the Saccharomyces cerevisiae RNA14, RNA15, and PAP1 genes on polyadenylation in vivo. Mol Cell Biol. 1995 Dec;15(12):6979–6986. doi: 10.1128/mcb.15.12.6979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Minvielle-Sebastia L., Winsor B., Bonneaud N., Lacroute F. Mutations in the yeast RNA14 and RNA15 genes result in an abnormal mRNA decay rate; sequence analysis reveals an RNA-binding domain in the RNA15 protein. Mol Cell Biol. 1991 Jun;11(6):3075–3087. doi: 10.1128/mcb.11.6.3075. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Neville M., Rosbash M. The NES-Crm1p export pathway is not a major mRNA export route in Saccharomyces cerevisiae. EMBO J. 1999 Jul 1;18(13):3746–3756. doi: 10.1093/emboj/18.13.3746. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Pederson T., Politz J. C. The nucleolus and the four ribonucleoproteins of translation. J Cell Biol. 2000 Mar 20;148(6):1091–1095. doi: 10.1083/jcb.148.6.1091. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Piper P. W. The heat shock and ethanol stress responses of yeast exhibit extensive similarity and functional overlap. FEMS Microbiol Lett. 1995 Dec 15;134(2-3):121–127. doi: 10.1111/j.1574-6968.1995.tb07925.x. [DOI] [PubMed] [Google Scholar]
  36. Proweller A., Butler S. Efficient translation of poly(A)-deficient mRNAs in Saccharomyces cerevisiae. Genes Dev. 1994 Nov 1;8(21):2629–2640. doi: 10.1101/gad.8.21.2629. [DOI] [PubMed] [Google Scholar]
  37. Saavedra C. A., Hammell C. M., Heath C. V., Cole C. N. Yeast heat shock mRNAs are exported through a distinct pathway defined by Rip1p. Genes Dev. 1997 Nov 1;11(21):2845–2856. doi: 10.1101/gad.11.21.2845. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Saavedra C., Tung K. S., Amberg D. C., Hopper A. K., Cole C. N. Regulation of mRNA export in response to stress in Saccharomyces cerevisiae. Genes Dev. 1996 Jul 1;10(13):1608–1620. doi: 10.1101/gad.10.13.1608. [DOI] [PubMed] [Google Scholar]
  39. Sarkar S., Hopper A. K. tRNA nuclear export in saccharomyces cerevisiae: in situ hybridization analysis. Mol Biol Cell. 1998 Nov;9(11):3041–3055. doi: 10.1091/mbc.9.11.3041. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Schneiter R., Kadowaki T., Tartakoff A. M. mRNA transport in yeast: time to reinvestigate the functions of the nucleolus. Mol Biol Cell. 1995 Apr;6(4):357–370. doi: 10.1091/mbc.6.4.357. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Schwer B., Guthrie C. PRP16 is an RNA-dependent ATPase that interacts transiently with the spliceosome. Nature. 1991 Feb 7;349(6309):494–499. doi: 10.1038/349494a0. [DOI] [PubMed] [Google Scholar]
  42. Seedorf M., Damelin M., Kahana J., Taura T., Silver P. A. Interactions between a nuclear transporter and a subset of nuclear pore complex proteins depend on Ran GTPase. Mol Cell Biol. 1999 Feb;19(2):1547–1557. doi: 10.1128/mcb.19.2.1547. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Segref A., Sharma K., Doye V., Hellwig A., Huber J., Lührmann R., Hurt E. Mex67p, a novel factor for nuclear mRNA export, binds to both poly(A)+ RNA and nuclear pores. EMBO J. 1997 Jun 2;16(11):3256–3271. doi: 10.1093/emboj/16.11.3256. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Snay-Hodge C. A., Colot H. V., Goldstein A. L., Cole C. N. Dbp5p/Rat8p is a yeast nuclear pore-associated DEAD-box protein essential for RNA export. EMBO J. 1998 May 1;17(9):2663–2676. doi: 10.1093/emboj/17.9.2663. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. Stade K., Ford C. S., Guthrie C., Weis K. Exportin 1 (Crm1p) is an essential nuclear export factor. Cell. 1997 Sep 19;90(6):1041–1050. doi: 10.1016/s0092-8674(00)80370-0. [DOI] [PubMed] [Google Scholar]
  46. Tang J., Rosbash M. Characterization of yeast U1 snRNP A protein: identification of the N-terminal RNA binding domain (RBD) binding site and evidence that the C-terminal RBD functions in splicing. RNA. 1996 Oct;2(10):1058–1070. [PMC free article] [PubMed] [Google Scholar]
  47. Tseng S. S., Weaver P. L., Liu Y., Hitomi M., Tartakoff A. M., Chang T. H. Dbp5p, a cytosolic RNA helicase, is required for poly(A)+ RNA export. EMBO J. 1998 May 1;17(9):2651–2662. doi: 10.1093/emboj/17.9.2651. [DOI] [PMC free article] [PubMed] [Google Scholar]
  48. Vainberg I. E., Dower K., Rosbash M. Nuclear export of heat shock and non-heat-shock mRNA occurs via similar pathways. Mol Cell Biol. 2000 Jun;20(11):3996–4005. doi: 10.1128/mcb.20.11.3996-4005.2000. [DOI] [PMC free article] [PubMed] [Google Scholar]
  49. Vijayraghavan U., Company M., Abelson J. Isolation and characterization of pre-mRNA splicing mutants of Saccharomyces cerevisiae. Genes Dev. 1989 Aug;3(8):1206–1216. doi: 10.1101/gad.3.8.1206. [DOI] [PubMed] [Google Scholar]
  50. Wente S. R., Rout M. P., Blobel G. A new family of yeast nuclear pore complex proteins. J Cell Biol. 1992 Nov;119(4):705–723. doi: 10.1083/jcb.119.4.705. [DOI] [PMC free article] [PubMed] [Google Scholar]
  51. Winston F., Dollard C., Ricupero-Hovasse S. L. Construction of a set of convenient Saccharomyces cerevisiae strains that are isogenic to S288C. Yeast. 1995 Jan;11(1):53–55. doi: 10.1002/yea.320110107. [DOI] [PubMed] [Google Scholar]
  52. Zeng Q., Hall K. B. Contribution of the C-terminal tail of U1A RBD1 to RNA recognition and protein stability. RNA. 1997 Mar;3(3):303–314. [PMC free article] [PubMed] [Google Scholar]
  53. Zhao J., Hyman L., Moore C. Formation of mRNA 3' ends in eukaryotes: mechanism, regulation, and interrelationships with other steps in mRNA synthesis. Microbiol Mol Biol Rev. 1999 Jun;63(2):405–445. doi: 10.1128/mmbr.63.2.405-445.1999. [DOI] [PMC free article] [PubMed] [Google Scholar]
  54. van Hoof A., Lennertz P., Parker R. Yeast exosome mutants accumulate 3'-extended polyadenylated forms of U4 small nuclear RNA and small nucleolar RNAs. Mol Cell Biol. 2000 Jan;20(2):441–452. doi: 10.1128/mcb.20.2.441-452.2000. [DOI] [PMC free article] [PubMed] [Google Scholar]

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