Tetrapyrrole biosynthetic enzyme protoporphyrinogen IX oxidase 1 is required for plastid RNA editing

Abstract

RNA editing is a posttranscriptional process that covalently alters the sequence of RNA molecules and plays important biological roles in both animals and land plants. In flowering plants, RNA editing converts specific cytidine residues to uridine in both plastid and mitochondrial transcripts. Previous studies identified pentatricopeptide repeat (PPR) motif-containing proteins as site-specific recognition factors for cytidine targets in RNA sequences. However, the regulatory mechanism underlying RNA editing was largely unknown. Here, we report that protoporphyrinogen IX oxidase 1 (PPO1), an enzyme that catalyzes protoporphyrinogen IX into protoporphyrin IX in the tetrapyrrole biosynthetic pathway, plays an unexpected role in editing multiple sites of plastid RNA transcripts, most of which encode subunits of the NADH dehydrogenase-like complex (NDH), in the reference plant Arabidopsis thaliana. We identified multiple organellar RNA editing factors (MORFs), including MORF2, MORF8, and MORF9, that interact with PPO1. We found that two conserved motifs within the 22-aa region at the N terminus of PPO1 are essential for its interaction with MORFs, its RNA editing function, and subsequently, its effect on NDH activity. However, transgenic plants lacking key domains for the tetrapyrrole biosynthetic activity of PPO1 exhibit normal RNA editing. Furthermore, MORF2 and MORF9 interact with three PPRs or related proteins required for editing of ndhB and ndhD sites. These results reveal that the tetrapyrrole biosynthetic enzyme PPO1 is required for plastid RNA editing, acting as a regulator that promotes the stability of MORF proteins through physical interaction. metabolism | organelle | editosome R NA editing, the process of covalently altering the sequence of an RNA molecule, generates protein diversity in eukaryotes (1, 2). Generally, in land plants, RNA editing highly specifically converts cytidine to uridine nucleotides in transcripts of both plastid and mitochondrial genes (3); 34 cytidine residues in plastids and more than 500 residues in mitochondria have been reported to be editing target sites in Arabidopsis thaliana (4, 5). A series of studies identified members of the PLS subfamily of pentatricopeptide repeat (PPR) motif-containing proteins as the site-specific recognition factors for cytidine targets (6). These specific PPR trans-acting proteins recognize cis elements within a region of ∼30 nt within the sites to be edited (1, 6-11). Although the DYW domains of some PPR factors contain several conserved residues with a cytidine deaminase motif (12), the enzyme that executes the editing reaction is elusive. Two recent reports documented that members of multiple organellar RNA editing factors (MORFs)/RNA editing factor interacting proteins (RIPs) widely affect RNA editing sites in both mitochondria and plastids (13,14). Organelle RNA recognition motif protein 1, which contains two truncated RIP domains, is also essential for plastid RNA editing (15). These studies reveal additional components of the plant organel...

Chloroplast SRP43 acts as a chaperone for glutamyl-tRNA reductase, the rate-limiting enzyme in tetrapyrrole biosynthesis

“…Considering the fact that developing green seedlings require ∼100-fold more Chl than heme, it is understandable that heme synthesis would be much less affected than Chl synthesis when ALA synthesis is compromised. Consistently, the knockout mutant of protoporphyrinogen IX oxidase 1 (ppox1), which encodes the last enzyme in the common pathway to Chl and heme biosynthesis, exhibited a severe albino phenotype but had a WT-like heme level (40). Furthermore, like chaos and chaos/gbp (Figs.…”

Quantitative analysis of motifs contributing to the interaction between PLS‐subfamily members and their target RNA sequences in plastid RNA editing

“…This and previous studies indicate that PLS‐class editing factors are unlikely to interact with this region (Barkan et al ., ; Okuda and Shikanai, ; Yagi et al ., ). The editosome in plant organelles consists of multiple components (Bentolila et al ., ; Sun et al ., ; Takenaka et al ., ; Tillich et al ., ; Zhang et al ., ). Some factor(s) other than PPR proteins may be involved in interactions downstream of target C, or the influence of this region may act through RNA secondary structure.…”

Selective Homo- and Heteromer Interactions between the Multiple Organellar RNA Editing Factor (MORF) Proteins in Arabidopsis thaliana

“…in mutants of one MORF protein where another MORF can perform, but cannot substitute as efficiently as the heteromer. It will now be interesting to determine the scale and specificity of the interactions between the various MORF, PPR, and other proteins (40) in plant organellar RNA editing.…”