Analysis of the Plant Mitochondrial Transcriptome

International audienc

Analyses des phosphoprotéines et du phosphoprotéome ( application à l étude de la phosphoénolpyruvate carboxylase kinase et à la carence en azote chez Arabidopsis thaliana )

ORSAY-PARIS 11-BU Sciences (914712101) / SudocSudocFranceF

The Pentatricopeptide Repeat Protein MEF100 Is Required for the Editing of Four Mitochondrial Editing Sites in <i>Arabidopsis</i>

In Arabidopsis thaliana there are more than 600 C-to-U RNA editing events in the mitochondria and at least 44 in the chloroplasts. Pentatricopeptide repeat (PPR) proteins provide the specificity for these reactions. They recognize RNA sequences in a partially predictable fashion via key amino acids at the fifth and last position in each PPR motif that bind to individual ribonucleotides. A combined approach of RNA-Seq, mutant complementation, electrophoresis of mitochondrial protein complexes and Western blotting allowed us to show that MEF100, a PPR protein identified in a genetic screen for mutants resistant to an inhibitor of γ -glutamylcysteine synthetase, is required for the editing of nad1-493, nad4-403, nad7-698 and ccmFN2-356 sites in Arabidopsis mitochondria. The absence of editing in mef100 leads to a decrease in mitochondrial Complex I activity, which probably explains the physiological phenotype. Some plants have lost the requirement for MEF100 at one or more of these sites through mutations in the mitochondrial genome. We show that loss of the requirement for MEF100 editing leads to divergence in the MEF100 binding site

Albinism does not correlate with biparental inheritance of plastid DNA in interspecific hybrids in \u3cem\u3eCicer\u3c/em\u3e species

Cultivated chickpea (Cicer arietinum) was crossed with its wild relatives from the genus Cicer to transfer favorable genes from the wider gene pool into the cultivar. Post-hybridization barriers led to yellowing and subsequent senescence from as early as 5 days after fertilization, however, the ovules of hybrid embryos could be rescued in vitro. Hybrids were classified as green, partially green or albino. The hybrid status of regenerated plantlets in vitro was confirmed by amplification of nuclear DNA markers. To check whether chloroplast development correlated with plastid DNA inheritance in these crosses, primers were designed using conserved plastid gene sequences from wild and cultivated species. All three possible plastid inheritance patterns were observed: paternal, maternal and biparental. This is the first report of biparental inheritance of plastid DNA in Cicer. No correlation was observed between parental origin of the plastid genome and degree of albinism, indicating that chloroplast development in hybrid genotypes was mostly influenced by nuclear factors

The MTL1 Pentatricopeptide repeat protein is required for both translation and splicing of the mitochondrial NADH DEHYDROGENASE SUBUNIT7 mRNA in arabidopsis

International audienceMitochondrial translation involves a complex interplay of ancient bacteria-like features and host-derived functionalities. Although the basic components of the mitochondrial translation apparatus have been recognized, very few protein factors aiding in recruiting ribosomes on mitochondria-encoded messenger RNA (mRNAs) have been identified in higher plants. In this study, we describe the identification of the Arabidopsis (Arabidopsis thaliana) MITOCHONDRIAL TRANSLATION FACTOR1 (MTL1) protein, a new member of the Pentatricopeptide Repeat family, and show that it is essential for the translation of the mitochondrial NADH dehydrogenase subunit7 (nad7) mRNA. We demonstrate that mtl1 mutant plants fail to accumulate the Nad7 protein, even though the nad7 mature mRNA is produced and bears the same 5' and 3' extremities as in wild-type plants. We next observed that polysome association of nad7 mature mRNA is specifically disrupted in mtl1 mutants, indicating that the absence of Nad7 results from a lack of translation of nad7 mRNA. These findings illustrate that mitochondrial translation requires the intervention of gene-specific nucleus-encoded PPR trans-factors and that their action does not necessarily involve the 59 processing of their target mRNA, as observed previously. Interestingly, a partial decrease in nad7 intron 2 splicing was also detected in mtl1 mutants, suggesting that MTL1 is also involved in group II intron splicing. However, this second function appears to be less essential for nad7 expression than its role in translation. MTL1 will be instrumental to understand the multifunctionality of PPR proteins and the mechanisms governing mRNA translation and intron splicing in plant mitochondria

Variation of the Polypeptide Composition of Mitochondria Isolated from Different Potato Tissues

The protein contents of mitochondria from different potato (Solanum tuberosum L.) tissues (tubers, dark-grown shoots, and green leaves) grown in a greenhouse or in vitro were compared by two-dimensional polyacrylamide gel electrophoresis. Two different methods were used: using the method that gave the highest resolution, an average number of 360 polypeptides was revealed on the mitochondrial patterns after silver staining. The mitochondrial protein patterns of etiolated tissues (tubers, dark-grown shoots) are roughly similar but distinct from those of green leaves. The four subunits of the glycine decarboxylase complex (involved in photorespiration) and a few other polypeptides are very abundant in green tissues, compared with nonphotosynthetic tissues. Conversely, some other polypeptides that are abundant in tubers and dark-grown shoots are hardly detectable in green leaf mitochondria. A rabbit antiserum was raised against a 40 kilodalton polypeptide that is among the most characteristic of these nonphotosynthetic tissue-specific polypeptides, and the N-terminal sequence of this polypeptide was determined. No effect of in vitro culture was observed on the protein composition of mitochondria isolated from differentiated tissues. However, the protein patterns of callus and cell suspension mitochondria are distinct from those of any differentiated tissues, although their basic pattern is clearly mitochondrial