RNA editing competence of trans‐factor MEF1 is modulated by ecotype‐specific differences but requires the DYW domain

Abstract

a b s t r a c t RNA editing in plant mitochondria posttranscriptionally changes multiple cytidines to uridines. The RNA editing trans-factor MEF1 was identified via ecotype-specific editing polymorphisms in Arabidopsis thaliana. Complementation assays reveal that none of the three amino acid changes between Columbia (Col) and C24 individually alters RNA editing. Only one combination of these polymorphisms lowers editing at two of the three target sites, suggesting additive effects of the involved SNPs. Functional importance of the C-terminal DYW domain was analysed with DYW-truncated and extended constructs. These do not recover RNA editing in protoplasts and regain only low levels in stable transformants. In MEF1, the DYW domain is thus required for full competence in RNA editing and its C-terminus has to be accessible.

Cytidine Deaminase Motifs within the DYW Domain of Two Pentatricopeptide Repeat-containing Proteins Are Required for Site-specific Chloroplast RNA Editing

“…7). These results are consistent with reports that several mitochondrial PPR-DYW editing factors cannot tolerate C-terminal fusions to GFP nor to a His tag (31,32). It is likely that C-terminal tags interfere with the function of the C-terminal DYW motif in some PPR-DYW proteins.…”

RNA Recognition Motif-Containing Protein ORRM4 Broadly Affects Mitochondrial RNA Editing and Impacts Plant Development and Flowering

“…9C). To examine the interaction between ORRM4 and PPR proteins, we selected MEF1 and MEF20, since decreased editing at site rps4 C956 was observed by either mef1 or orrm4 mutation while editing at site rps4 C226 is impaired in both mef20 and orrm4 mutants Zehrmann et al, 2010). The results indicate that ORRM4 does not interact with MEF1 or MEF20 in Y2H (Fig.…”

Mitochondrial RNA editing PPR proteins can tolerate protein tags at E as well as at DYW domain termini

“…Conversely, the DYW domain is essential in the MEF1 protein, truncated versions ending with the E domain cannot recover editing in the respective mutant. Concomitantly, the MEF1 protein does not tolerate a C-terminal extension by a His-tag, suggesting that some essential DYW termini have to be free and are not functional when extended (Zehrmann et al, 2010). The here observed partial complementation of the tagged MEF8 protein may also indicate that masking of the C-terminus by the GFP moiety does inhibit the activity and function of this DYW PPR protein.…”

“…This domain terminates in most instances with the name giving amino acid triplet DYW. In several such PPR RNA editing factors in plastids and mitochondria, the DYW domain can be deleted without compromising the function of the protein in editing (Okuda et al, 2009), while in others the DYW domain is required (Zehrmann et al, 2010). In some of the E class PPR RNA editing factors, addition of a DYW domain does not affect their competence in editing (Verbitskiy et al, 2012a).…”