Skip to main content
PMC home page
The EMBO Journal logoLink to The EMBO Journal
. 1990 Nov;9(11):3667–3673. doi: 10.1002/j.1460-2075.1990.tb07578.x

The nematode spliced leader RNA participates in trans-splicing as an Sm snRNP.

P A Maroney 1, G J Hannon 1, J A Denker 1, T W Nilsen 1
PMCID: PMC552121  PMID: 2145151

Abstract

The trans-spliced leader RNA (SL RNA) of nematodes resembles U snRNAs both in cap structure and in the presence of a consensus Sm binding site. We show here that synthetic SL RNA, synthesized by in vitro transcription, is efficiently used as a spliced leader donor in trans-splicing reactions catalyzed by a cell free extract prepared from developing embryos of the parasitic nematode, Ascaris lumbricoides. Efficient utilization of synthetic SL RNA requires a functional Sm binding site. Mutations within the Sm binding sequence that prevent immunoprecipitation by Sm antisera and prevent cap trimethylation abolish trans-splicing. The effect on trans-splicing is not due to undermethylation of the cap structure.

Full text

PDF
3667

Images in this article

3668Fig.1
on p.3668
3669Fig.2
on p.3669
3669Fig.3
on p.3669
3670Fig.4
on p.3670
3671Fig.5
on p.3671
3671Fig.6
on p.3671

Selected References

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

  1. Anderson G. J., Bach M., Lührmann R., Beggs J. D. Conservation between yeast and man of a protein associated with U5 small nuclear ribonucleoprotein. Nature. 1989 Dec 14;342(6251):819–821. doi: 10.1038/342819a0. [DOI] [PubMed] [Google Scholar]
  2. Bindereif A., Wolff T., Green M. R. Discrete domains of human U6 snRNA required for the assembly of U4/U6 snRNP and splicing complexes. EMBO J. 1990 Jan;9(1):251–255. doi: 10.1002/j.1460-2075.1990.tb08102.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Blumenthal T., Thomas J. Cis and trans mRNA splicing in C. elegans. Trends Genet. 1988 Nov;4(11):305–308. doi: 10.1016/0168-9525(88)90107-2. [DOI] [PubMed] [Google Scholar]
  4. Bruzik J. P., Van Doren K., Hirsh D., Steitz J. A. Trans splicing involves a novel form of small nuclear ribonucleoprotein particles. Nature. 1988 Oct 6;335(6190):559–562. doi: 10.1038/335559a0. [DOI] [PubMed] [Google Scholar]
  5. Chabot B., Black D. L., LeMaster D. M., Steitz J. A. The 3' splice site of pre-messenger RNA is recognized by a small nuclear ribonucleoprotein. Science. 1985 Dec 20;230(4732):1344–1349. doi: 10.1126/science.2933810. [DOI] [PubMed] [Google Scholar]
  6. Fabrizio P., McPheeters D. S., Abelson J. In vitro assembly of yeast U6 snRNP: a functional assay. Genes Dev. 1989 Dec;3(12B):2137–2150. doi: 10.1101/gad.3.12b.2137. [DOI] [PubMed] [Google Scholar]
  7. Hamm J., Dathan N. A., Mattaj I. W. Functional analysis of mutant Xenopus U2 snRNAs. Cell. 1989 Oct 6;59(1):159–169. doi: 10.1016/0092-8674(89)90878-7. [DOI] [PubMed] [Google Scholar]
  8. Hamm J., van Santen V. L., Spritz R. A., Mattaj I. W. Loop I of U1 small nuclear RNA is the only essential RNA sequence for binding of specific U1 small nuclear ribonucleoprotein particle proteins. Mol Cell Biol. 1988 Nov;8(11):4787–4791. doi: 10.1128/mcb.8.11.4787. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Hannon G. J., Maroney P. A., Branch A., Benenfield B. J., Robertson H. D., Nilsen T. W. Accurate processing of human pre-rRNA in vitro. Mol Cell Biol. 1989 Oct;9(10):4422–4431. doi: 10.1128/mcb.9.10.4422. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Hannon G. J., Maroney P. A., Denker J. A., Nilsen T. W. Trans splicing of nematode pre-messenger RNA in vitro. Cell. 1990 Jun 29;61(7):1247–1255. doi: 10.1016/0092-8674(90)90689-c. [DOI] [PubMed] [Google Scholar]
  11. Heinrichs V., Bach M., Winkelmann G., Lührmann R. U1-specific protein C needed for efficient complex formation of U1 snRNP with a 5' splice site. Science. 1990 Jan 5;247(4938):69–72. doi: 10.1126/science.2136774. [DOI] [PubMed] [Google Scholar]
  12. Kunkel T. A., Roberts J. D., Zakour R. A. Rapid and efficient site-specific mutagenesis without phenotypic selection. Methods Enzymol. 1987;154:367–382. doi: 10.1016/0076-6879(87)54085-x. [DOI] [PubMed] [Google Scholar]
  13. Maroney P. A., Hannon G. J., Nilsen T. W. Transcription and cap trimethylation of a nematode spliced leader RNA in a cell-free system. Proc Natl Acad Sci U S A. 1990 Jan;87(2):709–713. doi: 10.1073/pnas.87.2.709. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Mattaj I. W. Cap trimethylation of U snRNA is cytoplasmic and dependent on U snRNP protein binding. Cell. 1986 Sep 12;46(6):905–911. doi: 10.1016/0092-8674(86)90072-3. [DOI] [PubMed] [Google Scholar]
  15. Mattaj I. W., De Robertis E. M. Nuclear segregation of U2 snRNA requires binding of specific snRNP proteins. Cell. 1985 Jan;40(1):111–118. doi: 10.1016/0092-8674(85)90314-9. [DOI] [PubMed] [Google Scholar]
  16. Mattaj I. W., Habets W. J., van Venrooij W. J. Monospecific antibodies reveal details of U2 snRNP structure and interaction between U1 and U2 snRNPs. EMBO J. 1986 May;5(5):997–1002. doi: 10.1002/j.1460-2075.1986.tb04314.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. McPheeters D. S., Fabrizio P., Abelson J. In vitro reconstitution of functional yeast U2 snRNPs. Genes Dev. 1989 Dec;3(12B):2124–2136. doi: 10.1101/gad.3.12b.2124. [DOI] [PubMed] [Google Scholar]
  18. Mount S. M., Pettersson I., Hinterberger M., Karmas A., Steitz J. A. The U1 small nuclear RNA-protein complex selectively binds a 5' splice site in vitro. Cell. 1983 Jun;33(2):509–518. doi: 10.1016/0092-8674(83)90432-4. [DOI] [PubMed] [Google Scholar]
  19. Nilsen T. W., Shambaugh J., Denker J., Chubb G., Faser C., Putnam L., Bennett K. Characterization and expression of a spliced leader RNA in the parasitic nematode Ascaris lumbricoides var. suum. Mol Cell Biol. 1989 Aug;9(8):3543–3547. doi: 10.1128/mcb.9.8.3543. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Nilsen T. W. Trans-splicing in nematodes. Exp Parasitol. 1989 Nov;69(4):413–416. doi: 10.1016/0014-4894(89)90191-4. [DOI] [PubMed] [Google Scholar]
  21. Nishimura S. Minor components in transfer RNA: their characterization, location, and function. Prog Nucleic Acid Res Mol Biol. 1972;12:49–85. [PubMed] [Google Scholar]
  23. Pan Z. Q., Prives C. Assembly of functional U1 and U2 human-amphibian hybrid snRNPs in Xenopus laevis oocytes. Science. 1988 Sep 9;241(4871):1328–1331. doi: 10.1126/science.2970672. [DOI] [PubMed] [Google Scholar]
  24. Patton J. R., Pederson T. The Mr 70,000 protein of the U1 small nuclear ribonucleoprotein particle binds to the 5' stem-loop of U1 RNA and interacts with Sm domain proteins. Proc Natl Acad Sci U S A. 1988 Feb;85(3):747–751. doi: 10.1073/pnas.85.3.747. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Query C. C., Bentley R. C., Keene J. D. A specific 31-nucleotide domain of U1 RNA directly interacts with the 70K small nuclear ribonucleoprotein component. Mol Cell Biol. 1989 Nov;9(11):4872–4881. doi: 10.1128/mcb.9.11.4872. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Ruskin B., Green M. R. An RNA processing activity that debranches RNA lariats. Science. 1985 Jul 12;229(4709):135–140. doi: 10.1126/science.2990042. [DOI] [PubMed] [Google Scholar]
  27. Scherly D., Boelens W., van Venrooij W. J., Dathan N. A., Hamm J., Mattaj I. W. Identification of the RNA binding segment of human U1 A protein and definition of its binding site on U1 snRNA. EMBO J. 1989 Dec 20;8(13):4163–4170. doi: 10.1002/j.1460-2075.1989.tb08601.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Sharp P. A. Trans splicing: variation on a familiar theme? Cell. 1987 Jul 17;50(2):147–148. doi: 10.1016/0092-8674(87)90207-8. [DOI] [PubMed] [Google Scholar]
  29. Tazi J., Alibert C., Temsamani J., Reveillaud I., Cathala G., Brunel C., Jeanteur P. A protein that specifically recognizes the 3' splice site of mammalian pre-mRNA introns is associated with a small nuclear ribonucleoprotein. Cell. 1986 Dec 5;47(5):755–766. doi: 10.1016/0092-8674(86)90518-0. [DOI] [PubMed] [Google Scholar]
  30. Thomas J. D., Conrad R. C., Blumenthal T. The C. elegans trans-spliced leader RNA is bound to Sm and has a trimethylguanosine cap. Cell. 1988 Aug 12;54(4):533–539. doi: 10.1016/0092-8674(88)90075-x. [DOI] [PubMed] [Google Scholar]
  31. Van Doren K., Hirsh D. Trans-spliced leader RNA exists as small nuclear ribonucleoprotein particles in Caenorhabditis elegans. Nature. 1988 Oct 6;335(6190):556–559. doi: 10.1038/335556a0. [DOI] [PubMed] [Google Scholar]
  32. Vieira J., Messing J. Production of single-stranded plasmid DNA. Methods Enzymol. 1987;153:3–11. doi: 10.1016/0076-6879(87)53044-0. [DOI] [PubMed] [Google Scholar]
  33. Yuo C. Y., Weiner A. M. Genetic analysis of the role of human U1 snRNA in mRNA splicing: I. Effect of mutations in the highly conserved stem-loop I of U1. Genes Dev. 1989 May;3(5):697–707. doi: 10.1101/gad.3.5.697. [DOI] [PubMed] [Google Scholar]

Articles from The EMBO Journal are provided here courtesy of Nature Publishing Group

RESOURCES