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PMID: 9396820 Published · ppublish English Journal Article Research Support, U.S. Gov't, Non-P.H.S. Research Support, U.S. Gov't, P.H.S.

Use of an engineered ribozyme to produce a circular human exon.

Nucleic acids research ·Vol. 25 ·No. 24 ·1997-12-15 ·Pages 5085-94

Mikheeva S, Hakim-Zargar M, Carlson D, Jarrell K

Abstract

We report the use of an engineered ribozyme to produce a circular human exon in vitro. Specifically, we have designed a derivative of a yeast self-splicing group II intron that is able to catalyze the formation of a circular exon encoding the first kringle domain (K1) of the human tissue plasminogen activator protein. We show that the circular K1 exon is formed with high fidelity in vitro. Furthermore, the system is designed such that the circular exon that is produced consists entirely of human exon sequence. Thus, our results demonstrate that all yeast exon sequences are dispensable for group II intron catalyzed inverse splicing. This is the first demonstration that an engineered ribozyme can be used to create a circular exon containing only human sequences, linked together at a precise desired ligation point. We expect these results to be generalizable, so that similar ribozymes can be designed to precisely create circular derivatives of any nucleotide sequence.

MeSH Terms
Catalysis Cations, Monovalent/pharmacology DNA, Circular/biosynthesis,chemistry DNA, Fungal/genetics Exons Humans Introns/genetics Kringles/genetics Protein Engineering RNA Splicing/drug effects RNA, Catalytic/metabolism Saccharomyces cerevisiae/genetics Substrate Specificity
Chemicals
Cations, Monovalent DNA, Circular DNA, Fungal RNA, Catalytic
Authors & Affiliations
4 authors, click to expand affiliations / ORCID
Mikheeva S
Department of Pharmacology and Experimental Therapeutics, Boston University Medical Center, 80 East Concord Street, Boston, MA 02118, USA.
Hakim-Zargar M
Carlson D
Jarrell K
References (39)
39 references, click to expand
  1. Inverse splicing of a group II intron.
    Proc Natl Acad Sci U S A. 1993 Sep 15;90(18):8624-7 PMID: 8378340
  2. Circular transcripts of the testis-determining gene Sry in adult mouse testis.
    Cell. 1993 Jun 4;73(5):1019-30 PMID: 7684656
  3. Synthesis of circular RNA in bacteria and yeast using RNA cyclase ribozymes derived from a group I intron of phage T4.
    Proc Natl Acad Sci U S A. 1994 Apr 12;91(8):3117-21 PMID: 7512723
  4. Split genes and RNA splicing.
    Cell. 1994 Jun 17;77(6):805-15 PMID: 7516265
  5. Initiation of protein synthesis by the eukaryotic translational apparatus on circular RNAs.
    Science. 1995 Apr 21;268(5209):415-7 PMID: 7536344
  6. Splicing of a circular yeast pre-mRNA in vitro.
    Nucleic Acids Res. 1995 Apr 11;23(7):1133-9 PMID: 7537871
  7. The multiplicity of domains in proteins.
    Annu Rev Biochem. 1995;64:287-314 PMID: 7574483
  8. Two competing pathways for self-splicing by group II introns: a quantitative analysis of in vitro reaction rates and products.
    J Mol Biol. 1996 Feb 16;256(1):31-49 PMID: 8609612
  9. Generation and characterization of circular Bacillus subtilis RNase P RNA; activation by RNase P protein.
    Nucleic Acids Symp Ser. 1995;(33):92-4 PMID: 8643411
  10. Generation of nuclease resistant circular RNA decoys for HIV-Tat and HIV-Rev by autocatalytic splicing.
    Nucleic Acids Symp Ser. 1995;(33):152-5 PMID: 8643355
  11. Reactions catalyzed by group II introns in vitro.
    Methods Enzymol. 1996;264:66-86 PMID: 8965729
  12. Exon circularization in mammalian nuclear extracts.
    RNA. 1996 Jun;2(6):603-10 PMID: 8718689
  13. Use of engineered ribozymes to catalyze chimeric gene assembly.
    Proc Natl Acad Sci U S A. 1996 Jul 23;93(15):7486-90 PMID: 8755500
  14. Length changes in the joining segment between domains 5 and 6 of a group II intron inhibit self-splicing and alter 3' splice site selection.
    Mol Cell Biol. 1996 Oct;16(10):5896-904 PMID: 8816503
  15. A small circular TAR RNA decoy specifically inhibits Tat-activated HIV-1 transcription.
    Nucleic Acids Res. 1996 Oct 1;24(19):3733-8 PMID: 8871552
  16. Why genes in pieces?
    Nature. 1978 Feb 9;271(5645):501 PMID: 622185
  17. Making ends meet: a model for RNA splicing in fungal mitochondria.
    Nature. 1982 Dec 23;300(5894):719-24 PMID: 6757759
  18. Secondary structure of the Tetrahymena ribosomal RNA intervening sequence: structural homology with fungal mitochondrial intervening sequences.
    Proc Natl Acad Sci U S A. 1983 Jul;80(13):3903-7 PMID: 6306649
  19. Close relationship between certain nuclear and mitochondrial introns. Implications for the mechanism of RNA splicing.
    J Mol Biol. 1983 Jul 5;167(3):595-605 PMID: 6876158
  20. The structure of the human tissue-type plasminogen activator gene: correlation of intron and exon structures to functional and structural domains.
    Proc Natl Acad Sci U S A. 1984 Sep;81(17):5355-9 PMID: 6089198
  21. Genes-in-pieces revisited.
    Science. 1985 May 17;228(4701):823-4 PMID: 4001923
  22. An RNA processing activity that debranches RNA lariats.
    Science. 1985 Jul 12;229(4709):135-40 PMID: 2990042
  23. A self-splicing RNA excises an intron lariat.
    Cell. 1986 Jan 31;44(2):213-23 PMID: 3510741
  24. Excised group II introns in yeast mitochondria are lariats and can be formed by self-splicing in vitro.
    Cell. 1986 Jan 31;44(2):225-34 PMID: 2417726
  25. A role for exon sequences and splice-site proximity in splice-site selection.
    Cell. 1986 Aug 29;46(5):681-90 PMID: 2427200
  26. Improved free-energy parameters for predictions of RNA duplex stability.
    Proc Natl Acad Sci U S A. 1986 Dec;83(24):9373-7 PMID: 2432595
  27. Multiple exon-binding sites in class II self-splicing introns.
    Cell. 1987 Jul 3;50(1):17-29 PMID: 3297351
  28. Group II intron self-splicing. Alternative reaction conditions yield novel products.
    J Biol Chem. 1988 Mar 5;263(7):3432-9 PMID: 2830285
  29. Group II intron self-splicing: development of alternative reaction conditions and identification of a predicted intermediate.
    Cold Spring Harb Symp Quant Biol. 1987;52:223-32 PMID: 3331340
  30. Group II intron domain 5 facilitates a trans-splicing reaction.
    Mol Cell Biol. 1988 Jun;8(6):2361-6 PMID: 3405208
  31. Stability of RNA in developing Xenopus embryos and identification of a destabilizing sequence in TFIIIA messenger RNA.
    Development. 1988 Apr;102(4):837-52 PMID: 2458900
  32. RNA structure, not sequence, determines the 5' splice-site specificity of a group I intron.
    Proc Natl Acad Sci U S A. 1989 Oct;86(19):7402-6 PMID: 2678103
  33. Comparative and functional anatomy of group II catalytic introns--a review.
    Gene. 1989 Oct 15;82(1):5-30 PMID: 2684776
  34. Sequences and classification of group I and group II introns.
    Methods Enzymol. 1989;180:533-45 PMID: 2615660
  35. Tissue plasminogen activator: the biochemistry and pharmacology of variants produced by mutagenesis.
    Annu Rev Pharmacol Toxicol. 1990;30:91-121 PMID: 2111656
  36. Scrambled exons.
    Cell. 1991 Feb 8;64(3):607-13 PMID: 1991322
  37. Splice site selection and role of the lariat in a group II intron.
    J Mol Biol. 1991 Jun 5;219(3):415-28 PMID: 1711123
  38. Mis-splicing yields circular RNA molecules.
    FASEB J. 1993 Jan;7(1):155-60 PMID: 7678559
  39. A circular trans-acting hepatitis delta virus ribozyme.
    Nucleic Acids Res. 1993 Sep 11;21(18):4253-8 PMID: 7692400
Article Info
Journal
Nucleic acids research
Abbr.
Nucleic Acids Res
ISSN
0305-1048
DOI
10.1093/nar/25.24.5085
Published
1997-12-15
Pages
5085-94
Language
English
Region
England
NLM ID
0411011
PMCID
PMC147139
Subset
IM
Grants
NIGMS NIH HHS · GM52409-01A1 · United States
External Links
PubMed source DOI: 10.1093/nar/25.24.5085 PMC Full Text View journal details
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