In plants, protein synthesis occurs in the cytosol, mitochondria, and plastids. Each compartment requires a full set of tRNAs and aminoacyl-tRNA synthetases. We have undertaken a systematic analysis of the targeting of organellar aminoacyl-tRNA synthetases in the model plant Arabidopsis thaliana. Dual targeting appeared to be a general rule. Among the 24 identified organellar aminoacyltRNA synthetases (aaRSs), 15 (and probably 17) are shared between mitochondria and plastids, and 5 are shared between cytosol and mitochondria (one of these aaRSs being present also in chloroplasts). Only two were shown to be uniquely chloroplastic and none to be uniquely mitochondrial. Moreover, there are no examples where the three aaRS genes originating from the three ancestral genomes still coexist. These results indicate that extensive exchange of aaRSs has occurred during evolution and that many are now shared between two or even three compartments. The findings have important implications for studies of the translation machinery in plants and on protein targeting and gene transfer in general.GFP ͉ mitochondria ͉ plastids ͉ protein targeting ͉ translation

In plant mitochondria, transcription proceeds well beyond the region that will become mature 3 extremities of mRNAs, and the mechanisms of 3 maturation are largely unknown. Here, we show the involvement of two exoribonucleases, AtmtPNPase and AtmtRNaseII, in the 3 processing of atp9 mRNAs in Arabidopsis thaliana mitochondria. Down-regulation of AtmtPNPase results in the accumulation of pretranscripts of several times the size of mature atp9 mRNAs, indicating that 3 processing of these transcripts is performed mainly exonucleolytically by AtmtPNPase. This enzyme is however not sufficient to completely process atp9 mRNAs, because with down-regulation of another mitochondrial exoribonuclease, AtmtRNaseII, about half of atp9 transcripts exhibit short 3 nucleotide extensions compared with mature mRNAs. These short extensions can be efficiently removed by AtmtRNaseII in vitro. Taken together, these results show that 3 processing of atp9 mRNAs in Arabidopsis mitochondria is, at least, a twostep phenomenon. First, AtmtPNPase is involved in removing 3 extensions that may reach several kilobases. Second, AtmtRNaseII degrades short nucleotidic extensions to generate the mature 3 -ends.Functional RNAs are generated through a series of complex post-transcriptional processes. The nature of these processes can vary depending on the organism or cellular compartment considered and may include maturation of 5Ј and 3Ј termini, splicing of introns, capping, editing, base modifications, or addition of nongenomically-encoded nucleotides such as polyadenylation. Processing of mRNAs is well documented for bacterial and eukaryotic nuclear mRNAs (1, 2). Our knowledge of maturation of plastid transcripts is also extensive for both Chlamydomonas and higher plants (3). However, mRNA maturation processes in plant mitochondria are far less well characterized. Two lines of evidence suggest that processing of 3Ј extremities occurs for any RNA in plant mitochondria. First, run-on experiments have demonstrated that transcription proceeds beyond the region that corresponds to mature 3Ј-ends of mRNAs and rRNAs (4). Second, it has been shown using an in vitro transcription system that inverted repeats that terminate some mitochondrial mRNAs are not recognized as transcription termination signals (5). These inverted repeats seem to be essential as processing signals in vitro (5) and in vivo (6, 7). However, the ribonucleases involved in 3Ј mRNA maturation processes remain to be identified in plant mitochondria.Escherichia coli represents one of the model organisms for which the function, mechanism, and regulation of ribonucleases (RNases) are intensively studied (for example, see Refs. 1, 8, and 9). Among the eight characterized exoribonucleases in E. coli, polynucleotide phosphorylase (PNPase) 1 and RNaseII are the main exoribonucleases involved in degrading mRNAs in a 3Ј to 5Ј direction but participate also in processing events. PNPase-like proteins are also present in human mitochondria (10) and in chloroplasts. In the latter, it is involved both ...