In vivo and in vitro characterization of an RNA replication enhancer in a satellite RNA associated with turnip crinkle virus.
Virology
; 288(2): 315-24, 2001 Sep 30.
Article
en En
| MEDLINE
| ID: mdl-11601903
ABSTRACT
RNA replication enhancers are cis-acting elements that can stimulate replication or transcription of RNA viruses. Turnip crinkle virus (TCV) and satC, a parasitic RNA associated with TCV infections, contain stem-loop structures that are RNA replication enhancers (P. Nagy, J. Pogany, and A. E. Simon, EMBO J. 1999, 18, 5653-5665). We have found that replacement of 28 nt of the satC enhancer, termed the motif1-hairpin, with 28 randomized bases reduced satC accumulation 8- to 13-fold in Arabidopsis thaliana protoplasts. Deletion of single-stranded flanking sequences at either side of the hairpin also affected RNA accumulation with combined alterations at both sides of the hairpin showing the most detrimental effect in protoplasts. In vitro analysis with a partially purified TCV RdRp preparation demonstrated that the motif1-hairpin in its minus-sense orientation was able to stimulate RNA synthesis from the satC hairpin promoter (located at the 3' end of plus strands) by almost twofold. This level of RNA synthesis stimulation is approximately fivefold lower than that observed with a linear promoter, suggesting that a highly stable hairpin promoter is less responsive to the presence of the motif1-hairpin enhancer than a linear promoter. The motif1-hairpin in its plus-sense orientation was only 60% as active in enhancing transcription from the hairpin promoter. Since the motif1-hairpin is a hotspot for RNA recombination during plus-strand synthesis and since satC promoters located on the minus-strand are all short linear sequences, these findings support the hypothesis that the motif1-hairpin is primarily involved in enhancing plus-strand synthesis.
Buscar en Google
Colección:
01-internacional
Base de datos:
MEDLINE
Asunto principal:
ARN Viral
/
Elementos de Facilitación Genéticos
/
Carmovirus
/
Satélite de ARN
Tipo de estudio:
Clinical_trials
/
Risk_factors_studies
Idioma:
En
Revista:
Virology
Año:
2001
Tipo del documento:
Article
País de afiliación:
Estados Unidos