In Arabidopsis thaliana two co‐adapted cyto‐nuclear systems correlate with distinct ccmC transcript sizes

Abstract

SUMMARYPlant mitochondrial transcripts undergo maturation processes at both termini. Although frequently observed, the post-transcriptional formation of mature 5 0 ends is still poorly understood. We now analyzed the processing of transcripts derived from the mitochondrial ccmC gene, coding for a component of the cytochrome c maturation system. In Arabidopsis thaliana (Arabidopsis) there are two mitochondrial ccmC configurations, discriminated by a 66-bp sequence segment located approximately 500 bp upstream of the ccmC gene. In Arabidopsis accessions these divergent mitochondrial genotypes correlate with the generation of two different 5 0 termini that map to positions around À484 in accession Columbia (Col ccmC genotype) or À390 in accession C24 relative to the translation start codon (C24 ccmC genotype). Previously we identified RNA PROCESSING FACTOR 3 (RPF3), a pentatricopeptide repeat (PPR) protein required for the maturation of ccmC mRNAs with À484 5 0 ends transcribed from the Col ccmC genotype. Now we identified several accessions defective in maturation of ccmC transcripts. Taking advantage of this natural genetic variation we identified RNA PROCESSING FACTOR 6 (RPF6), a PPR protein necessary for the generation of ccmC mRNAs with À390 5 0 ends transcribed from the C24 ccmC genotype. Both Col-type and C24-type accessions encode RPF3 and RPF6 so that they can process ccmC transcripts derived from the two different mitochondrial genotypes. These factors and their cognate RNA recognition sites in the different ccmC genotypes are an intriguing example for the evolution of two co-adapted cyto-nuclear systems required for the same process i.e. 5 0 maturation of ccmC transcripts.

The pentatricopeptide repeat protein MTSF2 stabilizes a nad1 precursor transcript and defines the 3΄ end of its 5΄-half intron

“…Further analyses have revealed that most plant mitochondrial mRNA extremities are generated post-transcriptionally. Several P-type PPR proteins—most of them belonging to the restorer of fertility-like subfamily—have been found to be involved in 5΄ end formation of mitochondrial transcripts in Arabidopsis (37–43). However, the functional significance of 5΄ formation for most mitochondrial mRNAs remains unclear as transcripts with incorrect 5΄ ends (accumulating in ppr mutants) remain stable and correctly translated in most cases, and have no impact on plant development.…”

Maturation of 5′ ends of plant mitochondrial RNAs

“…Interestingly, there is natural variation among the abundance of the different nad4 precursor molecules in Arabidopsis accessions, which can be used to identify the factors involved in the stepwise 5′ maturation process. Multiple processing steps can also be assumed to be required for the generation of other mature mitochondrial mRNAs as indicated by the detection of several precursor molecules with different 5′ ends (Jonietz et al , Hauler et al , Stoll et al , ).…”

“…By using this natural genetic variation in forward genetic approaches, several RPFs were identified (Jonietz et al , Hölzle et al , Stoll et al , ). Based on the results of these studies also reverse genetic strategies were successfully applied to identify and characterize further RPFs (Jonietz et al , Hauler et al ).…”

“…Distinct amino acids at position 6 and 1′ (position 1 of the following repeat) specifically interact with certain nucleotides, albeit there is some degeneration of this code. Nevertheless, applying this code, binding sites for most of the RPFs could be predicted suggesting these recognition sites to be localized within the leader sequences upstream of the 5′ ends of the mature RNAs (Binder et al , Stoll et al ). Up to now, there is no direct experimental evidence supporting the interaction of the RPFs and the predicted recognition sites but there are clear hints that support their localization within the 5′ leader.…”

“…RPF5 is the sole RPF for which a true orthologue can be identified in the potato mitochondrial proteome (XP006346263), although conservation between the mitochondrial sequences surrounding the 5′ ends of the 26S rRNA genes in Arabidopsis and potato is restricted to a small segment between positions −17 and +9, excluding the predicted RPF5 binding site located further upstream (Binder et al , Salvato et al ). Striking support for the binding of RPFs upstream of the processing sites is given also by the identification of RPF3 and RPF6 and their cognate target sites (Stoll et al , Jonietz et al 2011). Here the involvement of one of the two factors correlates with the presence of distinct sequences upstream of the cleavage sites demonstrating the importance of the 5′ upstream region for binding and processing (further details see below).…”