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PMID: 1648201 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't

Antisense probes containing 2-aminoadenosine allow efficient depletion of U5 snRNP from HeLa splicing extracts.

Nucleic acids research ·Vol. 19 ·No. 12 ·1991-06-25 ·Pages 3193-8

Lamm GM, Blencowe BJ, Sproat BS, Iribarren AM, Ryder U, Lamond AI

Abstract

RNA duplexes containing the modified base 2-amino-adenine in place of adenine are stabilized through the formation of three hydrogen bonds in 2-amino A.U base pairs. Antisense 2'-O-alkyloligoribonucleotide probes incorporating 2-aminoadenosine are thus able to efficiently affinity select RNP particles which are otherwise inaccessible. This has allowed the efficient and specific depletion of U5 snRNP from HeLa cell nuclear splicing extracts. U5 snRNP is shown to be essential for spliceosome assembly and for both steps of pre-mRNA splicing. The absence of U5 snRNP prevents the stable association of U4/U6 but not U1 and U2 snRNPs with pre-mRNA.

MeSH Terms
Adenosine/analogs & derivatives,chemistry Base Sequence DNA Electrophoresis, Polyacrylamide Gel Endoribonucleases/metabolism HeLa Cells Humans Hydrogen Bonding Molecular Sequence Data Nucleic Acid Conformation RNA Precursors/metabolism RNA Probes/chemistry,metabolism RNA Splicing/physiology RNA, Antisense/chemistry,metabolism Ribonuclease H Ribonucleoproteins/metabolism,physiology Ribonucleoproteins, Small Nuclear
Chemicals
RNA Precursors RNA Probes RNA, Antisense Ribonucleoproteins Ribonucleoproteins, Small Nuclear 2-aminoadenosine DNA Endoribonucleases Ribonuclease H Adenosine
Authors & Affiliations
6 authors, click to expand affiliations / ORCID
Lamm G M
EMBL, Heidelberg, FRG.
Blencowe B J
Sproat B S
Iribarren A M
Ryder U
Lamond A I
References (41)
41 references, click to expand
  1. The 5' terminus of the RNA moiety of U1 small nuclear ribonucleoprotein particles is required for the splicing of messenger RNA precursors.
    Cell. 1984 Aug;38(1):299-307 PMID: 6235919
  2. The nuclear 5S RNAs from chicken, rat and man. U5 RNAs are encoded by multiple genes.
    Nucleic Acids Res. 1981 Feb 25;9(4):769-87 PMID: 6164980
  3. Mutations in yeast U5 snRNA alter the specificity of 5' splice-site cleavage.
    Cell. 1991 Apr 5;65(1):115-23 PMID: 2013092
  4. Targeted snRNP depletion reveals an additional role for mammalian U1 snRNP in spliceosome assembly.
    Cell. 1990 Oct 19;63(2):293-302 PMID: 2170025
  5. Universally conserved and yeast-specific U1 snRNA sequences are important but not essential for U1 snRNP function.
    Genes Dev. 1990 Oct;4(10):1766-74 PMID: 2147421
  6. Structure of spliceosomal snRNPs and their role in pre-mRNA splicing.
    Biochim Biophys Acta. 1990 Nov 30;1087(3):265-92 PMID: 2147394
  7. Sequence-specific affinity selection of mammalian splicing complexes.
    Nucleic Acids Res. 1990 Dec 25;18(24):7373-9 PMID: 2147994
  8. 2'-O-alkyl oligoribonucleotides as antisense probes.
    Proc Natl Acad Sci U S A. 1990 Oct;87(19):7747-51 PMID: 2145581
  9. Multiple interactions between the splicing substrate and small nuclear ribonucleoproteins in spliceosomes.
    Mol Cell Biol. 1987 Jan;7(1):281-93 PMID: 2951586
  10. Multiple factors including the small nuclear ribonucleoproteins U1 and U2 are necessary for pre-mRNA splicing in vitro.
    Cell. 1985 Oct;42(3):725-36 PMID: 2996774
  11. A mutational analysis of spliceosome assembly: evidence for splice site collaboration during spliceosome formation.
    Genes Dev. 1987 Aug;1(6):532-43 PMID: 2824284
  12. Spliceosome assembly involves the binding and release of U4 small nuclear ribonucleoprotein.
    Proc Natl Acad Sci U S A. 1988 Jan;85(2):411-5 PMID: 2963332
  13. Affinity chromatography of splicing complexes: U2, U5, and U4 + U6 small nuclear ribonucleoprotein particles in the spliceosome.
    Science. 1986 Sep 19;233(4770):1294-9 PMID: 3638792
  14. Cloning of the RNA8 gene of Saccharomyces cerevisiae, detection of the RNA8 protein, and demonstration that it is essential for nuclear pre-mRNA splicing.
    Mol Cell Biol. 1988 Mar;8(3):1067-75 PMID: 2835658
  15. Specific and stable intron-factor interactions are established early during in vitro pre-mRNA splicing.
    Cell. 1985 Nov;43(1):131-42 PMID: 3000599
  16. Spliceosome assembly in yeast.
    Genes Dev. 1987 Nov;1(9):1014-27 PMID: 2962902
  17. Identification of a yeast snRNP protein and detection of snRNP-snRNP interactions.
    Cell. 1987 Dec 24;51(6):1019-26 PMID: 2961458
  18. U2 as well as U1 small nuclear ribonucleoproteins are involved in premessenger RNA splicing.
    Cell. 1985 Oct;42(3):737-50 PMID: 2996775
  19. Interactions between small nuclear ribonucleoprotein particles in formation of spliceosomes.
    Cell. 1987 Jun 19;49(6):763-74 PMID: 2953438
  20. An ordered pathway of snRNP binding during mammalian pre-mRNA splicing complex assembly.
    EMBO J. 1987 Aug;6(8):2415-24 PMID: 2959470
  21. The role of the mammalian branchpoint sequence in pre-mRNA splicing.
    Genes Dev. 1988 Oct;2(10):1268-76 PMID: 3060403
  22. Antisense probing of the human U4/U6 snRNP with biotinylated 2'-OMe RNA oligonucleotides.
    Cell. 1989 Nov 3;59(3):531-9 PMID: 2478298
  23. The mammalian analogue of the yeast PRP8 splicing protein is present in the U4/5/6 small nuclear ribonucleoprotein particle and the spliceosome.
    Proc Natl Acad Sci U S A. 1989 Nov;86(22):8742-6 PMID: 2479028
  24. Pre-mRNA splicing in vitro requires intact U4/U6 small nuclear ribonucleoprotein.
    Cell. 1986 Aug 29;46(5):697-704 PMID: 2427202
  25. U1, U2, and U4/U6 small nuclear ribonucleoproteins are required for in vitro splicing but not polyadenylation.
    Cell. 1986 Aug 29;46(5):691-6 PMID: 2427201
  26. U5 small nuclear ribonucleoprotein: RNA structure analysis and ATP-dependent interaction with U4/U6.
    Mol Cell Biol. 1989 Aug;9(8):3350-9 PMID: 2552294
  27. Evidence from complementation assays in vitro that U5 snRNP is required for both steps of mRNA splicing.
    EMBO J. 1989 Oct;8(10):3105-12 PMID: 2531074
  28. Identification of functional U1 snRNA-pre-mRNA complexes committed to spliceosome assembly and splicing.
    Cell. 1989 Oct 20;59(2):349-58 PMID: 2529976
  29. Conservation between yeast and man of a protein associated with U5 small nuclear ribonucleoprotein.
    Nature. 1989 Dec 14;342(6251):819-21 PMID: 2532307
  30. Mapping U2 snRNP--pre-mRNA interactions using biotinylated oligonucleotides made of 2'-OMe RNA.
    EMBO J. 1989 Dec 20;8(13):4171-8 PMID: 2531659
  31. Probing the structure and function of U2 snRNP with antisense oligonucleotides made of 2'-OMe RNA.
    Cell. 1989 Jul 28;58(2):383-90 PMID: 2526684
  32. Highly efficient chemical synthesis of 2'-O-methyloligoribonucleotides and tetrabiotinylated derivatives; novel probes that are resistant to degradation by RNA or DNA specific nucleases.
    Nucleic Acids Res. 1989 May 11;17(9):3373-86 PMID: 2726482
  33. An essential yeast snRNA with a U5-like domain is required for splicing in vivo.
    Cell. 1987 Jun 5;49(5):613-24 PMID: 3555841
  34. 2NH2A X T helices in the ribo- and deoxypolynucleotide series. Structural and energetic consequences of 2NH2A substitution.
    Biochemistry. 1984 Dec 18;23(26):6723-32 PMID: 6529579
  35. Electrophoresis of ribonucleoproteins reveals an ordered assembly pathway of yeast splicing complexes.
    Nature. 1986 Nov 27-Dec 3;324(6095):341-5 PMID: 3537805
  36. Specific small nuclear RNAs are associated with yeast spliceosomes.
    Cell. 1986 Jun 20;45(6):869-77 PMID: 3518951
  37. A protected biotin containing deoxycytidine building block for solid phase synthesis of biotinylated oligonucleotides.
    Nucleic Acids Res. 1990 Aug 11;18(15):4355-60 PMID: 1697062
  38. Splicing stories and poly(A) tales: an update on RNA processing and transport.
    Curr Opin Cell Biol. 1990 Jun;2(3):528-38 PMID: 2143075
  39. Affinity purification of spliceosomes reveals that the precursor RNA processing protein PRP8, a protein in the U5 small nuclear ribonucleoprotein particle, is a component of yeast spliceosomes.
    Proc Natl Acad Sci U S A. 1990 Mar;87(6):2216-9 PMID: 2138328
  40. Splicing of messenger RNA precursors is inhibited by antisera to small nuclear ribonucleoprotein.
    Cell. 1983 Nov;35(1):101-7 PMID: 6194895
  41. New synthetic routes to synthons suitable for 2'-O-allyloligoribonucleotide assembly.
    Nucleic Acids Res. 1991 Feb 25;19(4):733-8 PMID: 1708121
Article Info
Journal
Nucleic acids research
Abbr.
Nucleic Acids Res
ISSN
0305-1048
DOI
10.1093/nar/19.12.3193
Published
1991-06-25
Pages
3193-8
Language
English
Region
England
NLM ID
0411011
PMCID
PMC328310
Subset
IM
External Links
PubMed source DOI: 10.1093/nar/19.12.3193 PMC Full Text View journal details
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