RNA editing complex interactions with a site for full-round U deletion in Trypanosoma brucei

Anastasia Sacharidou, Catherine Cifuentes-Rojas, Kari Halbig, Alfredo Hernandez, Lawrence J. Dangott, Monica De Nova-Ocampo, Jorge Cruz-Reyes

Research output: Contribution to journalArticle

11 Citations (Scopus)

Abstract

Trypanosome U insertion and U deletion RNA editing of mitochondrial pre-mRNAs is catalyzed by multisubunit editing complexes as directed by partially complementary guide RNAs. The basic enzymatic activities and protein composition of these high-molecular mass complexes have been under intense study, but their specific protein interactions with functional pre-mRNA/gRNA substrates remains unknown. We show that editing complexes purified through extensive ion-exchange chromatography and immunoprecipitation make specific cross-linking interactions with A6 pre-mRNA containing a single 32P and photoreactive 4-thioU at the scissile bond of a functional site for full-round U deletion. At least four direct protein-RNA contacts are detected at this site by cross-linking. All four interactions are stimulated by unpaired residues just 5′ of the pre-mRNA/ gRNA anchor duplex, but strongly inhibited by pairing of the editing site region. Furthermore, competition analysis with homologous and heterologous transcripts suggests preferential contacts of the editing complex with the mRNA/gRNA duplex substrate. This apparent structural selectivity suggests that the RNA-protein interactions we observe may be involved in recognition of editing sites and/or catalysis in assembled complexes. Copyright © 2006 RNA Society.
Original languageAmerican English
Pages (from-to)1219-1228
Number of pages10
JournalRNA
DOIs
StatePublished - 10 Jul 2006
Externally publishedYes

Fingerprint

Guide RNA
RNA Editing
Trypanosoma brucei brucei
RNA Precursors
Proteins
RNA
Complementary RNA
Trypanosomiasis
Ion Exchange Chromatography
Catalysis
Immunoprecipitation
Messenger RNA

Cite this

Sacharidou, A., Cifuentes-Rojas, C., Halbig, K., Hernandez, A., Dangott, L. J., De Nova-Ocampo, M., & Cruz-Reyes, J. (2006). RNA editing complex interactions with a site for full-round U deletion in Trypanosoma brucei. RNA, 1219-1228. https://doi.org/10.1261/rna.2295706
Sacharidou, Anastasia ; Cifuentes-Rojas, Catherine ; Halbig, Kari ; Hernandez, Alfredo ; Dangott, Lawrence J. ; De Nova-Ocampo, Monica ; Cruz-Reyes, Jorge. / RNA editing complex interactions with a site for full-round U deletion in Trypanosoma brucei. In: RNA. 2006 ; pp. 1219-1228.
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abstract = "Trypanosome U insertion and U deletion RNA editing of mitochondrial pre-mRNAs is catalyzed by multisubunit editing complexes as directed by partially complementary guide RNAs. The basic enzymatic activities and protein composition of these high-molecular mass complexes have been under intense study, but their specific protein interactions with functional pre-mRNA/gRNA substrates remains unknown. We show that editing complexes purified through extensive ion-exchange chromatography and immunoprecipitation make specific cross-linking interactions with A6 pre-mRNA containing a single 32P and photoreactive 4-thioU at the scissile bond of a functional site for full-round U deletion. At least four direct protein-RNA contacts are detected at this site by cross-linking. All four interactions are stimulated by unpaired residues just 5′ of the pre-mRNA/ gRNA anchor duplex, but strongly inhibited by pairing of the editing site region. Furthermore, competition analysis with homologous and heterologous transcripts suggests preferential contacts of the editing complex with the mRNA/gRNA duplex substrate. This apparent structural selectivity suggests that the RNA-protein interactions we observe may be involved in recognition of editing sites and/or catalysis in assembled complexes. Copyright {\circledC} 2006 RNA Society.",
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Sacharidou, A, Cifuentes-Rojas, C, Halbig, K, Hernandez, A, Dangott, LJ, De Nova-Ocampo, M & Cruz-Reyes, J 2006, 'RNA editing complex interactions with a site for full-round U deletion in Trypanosoma brucei', RNA, pp. 1219-1228. https://doi.org/10.1261/rna.2295706

RNA editing complex interactions with a site for full-round U deletion in Trypanosoma brucei. / Sacharidou, Anastasia; Cifuentes-Rojas, Catherine; Halbig, Kari; Hernandez, Alfredo; Dangott, Lawrence J.; De Nova-Ocampo, Monica; Cruz-Reyes, Jorge.

In: RNA, 10.07.2006, p. 1219-1228.

Research output: Contribution to journalArticle

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T1 - RNA editing complex interactions with a site for full-round U deletion in Trypanosoma brucei

AU - Sacharidou, Anastasia

AU - Cifuentes-Rojas, Catherine

AU - Halbig, Kari

AU - Hernandez, Alfredo

AU - Dangott, Lawrence J.

AU - De Nova-Ocampo, Monica

AU - Cruz-Reyes, Jorge

PY - 2006/7/10

Y1 - 2006/7/10

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AB - Trypanosome U insertion and U deletion RNA editing of mitochondrial pre-mRNAs is catalyzed by multisubunit editing complexes as directed by partially complementary guide RNAs. The basic enzymatic activities and protein composition of these high-molecular mass complexes have been under intense study, but their specific protein interactions with functional pre-mRNA/gRNA substrates remains unknown. We show that editing complexes purified through extensive ion-exchange chromatography and immunoprecipitation make specific cross-linking interactions with A6 pre-mRNA containing a single 32P and photoreactive 4-thioU at the scissile bond of a functional site for full-round U deletion. At least four direct protein-RNA contacts are detected at this site by cross-linking. All four interactions are stimulated by unpaired residues just 5′ of the pre-mRNA/ gRNA anchor duplex, but strongly inhibited by pairing of the editing site region. Furthermore, competition analysis with homologous and heterologous transcripts suggests preferential contacts of the editing complex with the mRNA/gRNA duplex substrate. This apparent structural selectivity suggests that the RNA-protein interactions we observe may be involved in recognition of editing sites and/or catalysis in assembled complexes. Copyright © 2006 RNA Society.

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Sacharidou A, Cifuentes-Rojas C, Halbig K, Hernandez A, Dangott LJ, De Nova-Ocampo M et al. RNA editing complex interactions with a site for full-round U deletion in Trypanosoma brucei. RNA. 2006 Jul 10;1219-1228. https://doi.org/10.1261/rna.2295706