DELAYED GREENING 409 encodes a dual-localized pentatricopeptide repeat protein required for chloroplast and mitochondrial development

Abstract In flowering plants, hundreds of RNA editing events occur in the chloroplasts and mitochondria during posttranscriptional processes. Although several pentatricopeptide repeat (PPR) proteins have been shown to form the editosome core, the precise interactions between the different editing factors are still obscure. Here, we isolated an Arabidopsis (Arabidopsis thaliana) PPR protein, designated DELAYED GREENING 409 (DG409), that was dually targeted to chloroplasts and mitochondria. This protein consists of 409 amino acids with 7 PPR motifs but lacks a C-terminal E, E+, or DYW domain. A mild dg409 knockdown mutant displays a sickly phenotype. In this mutant, the young leaves are pale green and turn green at maturity, and the development of chloroplasts and mitochondria is severely disrupted. Complete loss of DG409 function results in defective embryos. Transcriptomic analysis of the dg409 knockdown plants showed some editing defects in genes from both organelles, including CASEINOLYTIC PROTEASE P (clpP)-559, RNA POLYMERASE SUBUNIT ALPHA (rpoA)-200, ACETYL-COA CARBOXYLASE CARBOXYL TRANSFERASE SUBUNIT BETA (accD)-1568, NADH DEHYDROGENASE SUBUNIT 7 (nad7)-1505, and RIBOSOMAL PROTEIN S3 (rps3)-1344. RNA immunoprecipitation showed that DG409 was associated with the targeted transcripts in vivo. Interaction assays revealed that DG409 directly interacted with 2 DYW-type PPR proteins (EARLY CHLOROPLAST BIOGENESIS2 [AtECB2] and DYW DOMAIN PROTEIN2 [DYW2]) and 3 multiple organellar RNA editing factors (MORF2, MORF8, and MORF9). These results indicate that DG409 is involved in RNA editing via protein complexes and is therefore essential for chloroplast and mitochondrial development.