Transfer of plastid RNA-editing activity to novel sites suggests a critical role for spacing in editing-site recognition.

RNA editing in higher plant plastids alters mRNA sequences by C-to-U conversions at highly specific sites through an unknown mechanism. 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 reduction of this distance by 1 nt entirely abolishes RNA editing. Surprisingly, deletions or combinations of deletional and point mutations that shift a heterologous cytidine residue in the same distance from the upstream cis-element as the editing site in the wild type result in transfer of the RNA-editing activity to the heterologous cytidine whereas the wild-type site remains unedited. Our results suggest that the molecular identity of at least some editing sites in the chloroplast genome is defined by their distance from an essential upstream sequence element.

[1]  P. Nixon,et al.  Identification of a functional respiratory complex in chloroplasts through analysis of tobacco mutants containing disrupted plastid ndh genes , 1998, The EMBO journal.

[2]  M. Hanson,et al.  A guide to RNA editing. , 1997, RNA.

[3]  R. Bock,et al.  Identification of critical nucleotide positions for plastid RNA editing site recognition. , 1997, RNA.

[4]  R. Bock,et al.  Extraplastidic site‐specific factors mediate RNA editing in chloroplasts , 1997, The EMBO journal.

[5]  K. Stuart,et al.  RNA editing: getting U into RNA. , 1997, Trends in biochemical sciences.

[6]  W. Li,et al.  Apobec-1 and apolipoprotein B mRNA editing. , 1997, Biochimica et biophysica acta.

[7]  P. Maliga,et al.  Sequences directing C to U editing of the plastid psbL mRNA are located within a 22 nucleotide segment spanning the editing site. , 1996, The EMBO journal.

[8]  R. Bock,et al.  In vivo dissection of cis‐acting determinants for plastid RNA editing. , 1996, The EMBO journal.

[9]  H. Carrer,et al.  Site‐specific factor involved in the editing of the psbL mRNA in tobacco plastids. , 1995, The EMBO journal.

[10]  R. Bock,et al.  Introduction of a heterologous editing site into the tobacco plastid genome: the lack of RNA editing leads to a mutant phenotype. , 1994, The EMBO journal.

[11]  P. Maliga,et al.  High-frequency plastid transformation in tobacco by selection for a chimeric aadA gene. , 1993, Proceedings of the National Academy of Sciences of the United States of America.

[12]  A. Brennicke,et al.  RNA editing in plant mitochondria. , 1989, Seminars in cell biology.

[13]  H. Kössel,et al.  Identification of editing positions in the ndhB transcript from maize chloroplasts reveals sequence similarities between editing sites of chloroplasts and plant mitochondria. , 1992, Nucleic acids research.

[14]  J. Kudla,et al.  RNA editing in tobacco chloroplasts leads to the formation of a translatable psbL mRNA by a C to U substitution within the initiation codon. , 1992, The EMBO journal.

[15]  S. Litvak,et al.  An in vitro system for the editing of ATP synthase subunit 9 mRNA using wheat mitochondrial extracts. , 1992, Proceedings of the National Academy of Sciences of the United States of America.

[16]  G. Igloi,et al.  Editing of a chloroplast mRNA by creation of an initiation codon , 1991, Nature.

[17]  P. Maliga,et al.  Stable transformation of plastids in higher plants. , 1990, Proceedings of the National Academy of Sciences of the United States of America.

[18]  W. Lüke,et al.  Improvement of PCR amplified DNA sequencing with the aid of detergents. , 1990, Nucleic acids research.

[19]  J. Doyle,et al.  Isolation of plant DNA from fresh tissue , 1990 .

[20]  Michael W. Gray,et al.  RNA editing in plant mitochondria , 1989, Nature.

[21]  José M. Gualberto,et al.  RNA editing in wheat mitochondria results in the conservation of protein sequences , 1989, Nature.

[22]  R. Benne,et al.  Major transcript of the frameshifted coxll gene from trypanosome mitochondria contains four nucleotides that are not encoded in the DNA , 1986, Cell.

[23]  F. Takaiwa,et al.  The complete nucleotide sequence of the tobacco chloroplast genome: its gene organization and expression , 1986, The EMBO journal.

[24]  F. Skoog,et al.  A revised medium for rapid growth and bio assays with tobacco tissue cultures , 1962 .