RNA PROCESSING FACTOR2 Is Required for 5′ End Processing of <i>nad9</i> and <i>cox3</i> mRNAs in Mitochondria of <i>Arabidopsis thaliana</i>

Abstract

In mitochondria of higher plants, the majority of 5' termini of mature mRNAs are generated posttranscriptionally. To gain insight into this process, we analyzed a natural 5' end polymorphism in the species Arabidopsis thaliana. This genetic approach identified the nuclear gene At1g62670, encoding a pentatricopeptide repeat protein. The functional importance of this mitochondrial restorer of fertility-like protein, designated RNA PROCESSING FACTOR2 (RPF2), is confirmed by the analysis of a respective T-DNA knockout mutant and its functional restoration by in vivo complementation. RPF2 fulfills two functions: it is required for the generation of a distinct 5' terminus of transcripts of subunit 9 of the NADH DEHYDROGENASE complex (nad9) and it determines the efficiency of 5' end formation of the mRNAs for subunit 3 of the CYTOCHROME C OXIDASE (cox3), the latter also being influenced by mitochondrial DNA sequences. Accordingly, recombinant RPF2 protein directly binds to a nad9 mRNA fragment in vitro. Two-dimensional gel electrophoresis and immunodetection analyses reveal that altered 5' processing does not influence accumulation of the nad9 and cox3 polypeptides. In accessions C24, Oystese-1, and Yosemite-0, different inactive RPF2 alleles exist, demonstrating the variability of this gene in Arabidopsis. The identification of RPF2 is a major step toward the characterization of 5' mRNA processing in mitochondria of higher plants.