Recognition of RNA editing sites in plastid transcripts
Bock, R. and Hermann, M.
Institut für Biologie III, Universität Freiburg, Schänzlestraße 1, D-79104 Freiburg, Germany
RNA editing in plant organelles alters mRNA sequences by pyrimidine-to-pyrimidine conversions at highly specific sites through an unknown mechanism. To gain a better understanding of the molecular interactions involved in organellar RNA editing we have begun to define the cis-acting elements involved in the molecular recognition of plastid editing sites. We constructed a series of chloroplast transformation vectors harboring editing sites in a chimeric context. The constructs were inserted into the chloroplast genome by biolistic transformation and editing in the chimeric mRNAs was examined. Successive deletion analyses led to the identification of an upstream sequence element which is absolutely required for editing of the downstream RNA editing site. In addition, we performed a scanning point mutagenesis to determine individual nucleotide residues that are critical for plastid editing site recognition.
In order to elucidate how the cytidine residues to be edited are specifically recognized and distinguished from other cytidines in close proximity, we have changed in vivo the distances of two plastid RNA editing sites from their essential upstream cis-acting sequence element. Analysis of RNA editing in transgenic chloroplasts revealed that already reduction of this distance by one nucleotide entirely abolishes RNA editing. Surprisingly, mutations that shift a heterologous cytidine residue in the same distance from the upstream cis-element as is the editing site in the wild type, result in transfer of the RNA editing activity to the heterologous cytidine while the wild-type site remains unedited. Our results suggest a two-step model for plant organellar RNA editing in which the molecular identity of the editing sites is precisely defined by their distance from an essential upstream 'mooring' sequence.
Bock, R., Hermann, M. and Kössel, H. (1996). In vivo dissection of cis-acting determinants for plastid RNA editing. EMBO J., 15, 5052-5059.
Bock, R., Hermann, M. and Fuchs, M. (1997). Identification of critical nucleotide positions for plastid RNA editing site recognition. RNA, 3, 1194-1200.
| LOCATION |
DATE |
TIME |
| Lecture Hall I |
Thursday, April 9 |
04:30 pm |