Trypanosome RNA editing: simple guide RNA features enhance U deletion 100-fold.
Mol Cell Biol
; 21(3): 884-92, 2001 Feb.
Article
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| MEDLINE
| ID: mdl-11154275
ABSTRACT
Trypanosome RNA editing is a massive processing of mRNA by U deletion and U insertion, directed by trans-acting guide RNAs (gRNAs). A U deletion cycle and a U insertion cycle have been reproduced in vitro using synthetic ATPase (A6) pre-mRNA and gRNA. Here we examine which gRNA features are important for this U deletion. We find that, foremost, this editing depends critically on the single-stranded character of a few gRNA and a few mRNA residues abutting the anchor duplex, a feature not previously appreciated. That plus any base-pairing sequence to tether the upstream mRNA are all the gRNA needs to direct unexpectedly efficient in vitro U deletion, using either the purified editing complex or whole extract. In fact, our optimized gRNA constructs support faithful U deletion up to 100 times more efficiently than the natural gRNA, and they can edit the majority of mRNA molecules. This is a marked improvement of in vitro U deletion, in which previous artificial gRNAs were no more active than natural gRNA and the editing efficiencies were at most a few percent. Furthermore, this editing is not stimulated by most other previously noted gRNA features, including its potential ligation bridge, 3' OH moiety, any U residues in the tether, the conserved structure of the central region, or proteins that normally bind these regions. Our data also have implications about evolutionary forces active in RNA editing.
Texto completo:
1
Bases de datos:
MEDLINE
Asunto principal:
Trypanosoma brucei brucei
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ARN Protozoario
/
Edición de ARN
Límite:
Animals
Idioma:
En
Revista:
Mol Cell Biol
Año:
2001
Tipo del documento:
Article
País de afiliación:
Estados Unidos