
%0 Journal Article
%A MIYAMOTO, Tetsuya 
%A OBOKATA, Junichi 
%A WAKASUGI, Tatsuya 
%A SUGIURA, Masahiro 
%T In vitro analysis of cis-elements and trans-factors for the RNA editing of tobacco chloroplast psbE and petB mRNAs
%J Science Access
%D 2001/01/01
%V 3
%N 1
%P -
%K 
%R https://doi.org/10.1071/SA0403721
%U https://www.publish.csiro.au/paper/SA0403721 
%X RNA editing is one of the posttranscriptional processes in higher plant chloroplasts. Editing is found in many cases in internal protein-coding regions, resulting in amino acid substitutions. Therefore, it is important to understand editing events to use the chloroplast as a factory for protein production. The number of editing sites so far observed is 31 in tobacco and these editing events are C- to -U conversions. To analyze the molecular mechanism, we have developed an in vitro RNA editing system from tobacco green chloroplasts (1). Extracts were prepared from intact chloroplasts isolated from 5-10 cm tobacco leaves. Pre-mRNA portions in which the C residue to be edited was specifically labelled with 32P were used as in vitro substrates. Editing activity was assayed by detecting 32P-labelled U mononucleotides on cellulose TLC after digestion of substrates with nuclease P1.  Using the in vitro system, cis-acting elements for editing were defined in tobacco chloroplast mRNAs from psbE and petB. In vitro competition assays indicated that trans-acting factors interact with these cis-elements and that the trans-factor is site-specific. UV cross-linking assays showed the existence of site-specific trans-factors for these mRNA editing.   (1) T. Hirose and M. Sugiura, Involvement of a site-specific trans-acting factor and a common RNA-binding protein in the editing of chloroplast mRNAs: development of a chloroplast in vitro RNA editing system. EMBO J. 20: 1144-1152 (2001) 
