[Show abstract][Hide abstract] ABSTRACT: 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 aminoacyl-tRNA 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.
Proceedings of the National Academy of Sciences 12/2005; 102(45):16484-9. DOI:10.1073/pnas.0504682102 · 9.67 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Two cysteinyl-tRNA synthetases (CysRS) and four asparaginyl-tRNA synthetases (AsnRS) from Arabidopsis thaliana were characterized from genome sequence data, EST sequences, and RACE sequences. For one CysRS and one AsnRS, sequence alignments and prediction programs suggested the presence of an N-terminal organellar targeting peptide. Transient expression of these putative targeting sequences joined to jellyfish green fluorescent protein (GFP) demonstrated that both presequences can efficiently dual-target GFP to mitochondria and plastids. The other CysRS and AsnRSs lack targeting sequences and presumably aminoacylate cytosolic tRNAs. Phylogenetic analysis suggests that the four AsnRSs evolved by repeated duplication of a gene transferred from an ancestral plastid and that the CysRSs also arose by duplication of a transferred organelle gene (possibly mitochondrial). These case histories are the best examples to date of capture of organellar aminoacyl-tRNA synthetases by the cytosolic protein synthesis machinery.
[Show abstract][Hide abstract] ABSTRACT: The translation systems of plant mitochondria differ from those of other mitochondria in that they incorporate tRNAs of three different origins: native mitochondrial tRNAs, plastid tRNAs transcribed from plastid DNA insertions in mitochondrial DNA, and nuclearly encoded imported tRNAs, The complicated evolutionary history of the tRNA replacement events leading up to this situation is slowly being unraveled. Recent research on plant aminoacyl-tRNA synthetases is starting to reveal how the mitochondrial compartment can cope with this unusual mix of tRNAs and has uncovered an unprecedented degree of sharing of isoforms between compartments. Many plant aminoacyl-tRNA synthetases are dual targeted to two compartments, either cytosol/mitochondria or plastids/mitochondria. The molecular basis for some of these cases of dual targeting are described.
Journal of Heredity 05/1999; 90(3):333-337. DOI:10.1093/jhered/90.3.333 · 2.09 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: In plants, all aminoacyl-tRNA synthetases are nuclearly encoded, despite the fact that their activities are required in the three protein-synthesizing cell compartments (cytosol, mitochondria, and chloroplasts). To investigate targeting of these enzymes, we cloned cDNAs encoding alanyl-tRNA synthetase (AlaRS) and the corresponding nuclear gene, ALATS, from Arabidopsis by using degenerate polymerase chain reaction primers based on highly conserved regions shared between known AlaRSs from other organisms. Analysis of the transcription of the gene showed the presence of two potential translation initiation codons in some ALATS mRNAs. Translation from the upstream AUG would generate an N-terminal extension with features characteristic of mitochondrial targeting peptides. A polyclonal antibody raised against part of the Arabidopsis AlaRS revealed that the Arabidopsis cytosolic and mitochondrial AlaRSs are immunologically similar, suggesting that both isoforms are encoded by the ALATS gene. In vitro experiments confirmed that two polypeptides can be translated from AlATS transcripts, with most ribosomes initiating on the downstream AUG to give the shorter polypeptide corresponding in size to the cytosolic enzyme. The ability of the presequence encoded between the two initiation codons to direct polypeptides to mitochondria was demonstrated by expression of fusion proteins in tobacco protoplasts and in yeast. We conclude that the ALATS gene encodes both the cytosolic and the mitochondrial forms of AlaRS, depending on which of the two AUG codons is used to initiate translation.
The Plant Cell 07/1996; 8(6):1027-39. DOI:10.1105/tpc.8.6.1027 · 9.34 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Mitochondrial DNA polymorphism was employed to assess cytoplasmic diversity among cytoypes of the genus Cichorium and related genera of the tribe Lactuceae (Asteraceae). Hybridization patterns of total DNA using six restriction enzymes and five heterologous mtDNA probes were examined. From estimates of mtDNA diversity, Cichorium spinosum appeared as an ecotype of C. intybus rather than a separate species. Interspecific mtDNA polymorphism in the genus Cichorium was higher than that observed in Cicerbita Crepis, Lactuca and Tragopogon. Molecular data seemed to indicate that Catananche is very distant from the other genera examined. Intergeneric comparisons allowed the clustering of Cicerbita, Lactuca and Cichorium, genera which belong to different subtribes. However, further molecular investigations on a larger number of genera are needed to clarify the relationships among genera within and between subtribes of the tribe Lactuceae.
[Show abstract][Hide abstract] ABSTRACT: The mt genome of higher plants (size varying from 200 to 560 kb according to the species) consists of a heterogeneous population of molecules resulting from intra- or inter-molecular recombination. Besides the highly conserved coding sequences of mt genes, other sequences, for example, promiscuous chloroplast sequences, may be found. The genes may be interrupted by introns; in some cases, one or more introns have been split, leading to the scattering of the gene segments to remote parts of the genome; this structure requires trans-splicing steps for the mature messenger RNA to be obtained. Over 25 genes have been identified so far that are usable, as universal polymorphism markers in RFLP (restriction fragment length polymorphism) characterization of mt genomes due to sequence conservation. During the search for new genes we identified 2 new coding sequences, nad5 and nad6; nad6 consists of a single reading frame whereas nad5 is a split gene whose 3 independent segments are independently transcribed.
[Show abstract][Hide abstract] ABSTRACT: A region of the mitochondrial (mt) DNA of wheat was studied because of its homology with other plant mtDNAs. Sequence analysis revealed an open reading frame encoding a polypeptide of 247 amino acids. Comparison of the sequence of the putative polypeptide with the protein sequence data of the Swiss-Prot library reveals homology with subunit 6 of the NADH-ubiquinone complex of mitochondria from Marchantia polymorpha, Podospora anserina, Chlamydomonas reinhardtii and of chloroplasts from M. polymorpha and Oryza sativa. No similarity was detected when compared with the subunit 6 of animal mitochondria, probably due to the rapid evolution of the sequence. A single 1.2 kb transcript appears in northern RNA blots. We found 15 edited sites of which only 13 give amino acid changes. This is the first report of a mt nad6 gene in higher plants.
[Show abstract][Hide abstract] ABSTRACT: Sequence analysis of the Ogura-specific mitochondria) DNA (mtDNA) fragment isolated previously from Brassica cybrids carrying Ogura cytoplasmic male sterility (cms) revealed a tRNAfMet sequence, a putative 138 amino acid open reading frame (orf138), and a 158 amino acid ORF (orf158) previously observed in mitochondria) genomes from several other plant species. Transcription mapping showed that both ORFs are present on a 1.4 kb cms-specific transcript. The orf158 sequence is also transcribed in fertile plants on a different mRNA, and thus is unlikely to be related to cms. On the other hand, fertile revertant plants lack transcripts of the orf138 sequence, whose possible role in the mechanism of Ogura cms is discussed.
MGG - Molecular and General Genetics 10/1992; 235(2):340-348. DOI:10.1007/BF00279379
[Show abstract][Hide abstract] ABSTRACT: The mitochondrial single-copy gene nad5 of wheat and maize consists of 5 exons located on three widely separated regions of the genome that are independently transcribed. The first region contains exons I and II separated by an atypical group II intron; in the second region is exon III (only 22 bp long), which is flanked upstream by a maturase-related open reading frame (ORF) and exon e of the nad1 gene, and downstream by a previously unidentified ORF (ORF143); in the third region are exons IV and V separated by a group II intron. In maize, this last domain is flanked upstream by the genes rps12, nad3, and tRNA(Ser) and downstream by a chloroplast tRNA(Cys). RNA editing occurs in wheat exons IV and V as C-to-U changes. A detailed analysis of the transcription of the nad5 gene in wheat and maize reveals that the exons are assembled into a 2.4-kb mRNA after two cis-splicing (between exons I and II and exons IV and V) and two trans-splicing events. The trans-splicing process involves the sequences flanking exons II, III, and IV that feature group II introns. A model is proposed for the assembly and maturation of the nad5 transcripts.
The Plant Cell 01/1992; 3(12):1363-78. · 9.34 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Five diploid potato clones have been transformed by electroporation of protoplasts with different selectable markers. The resulting diploid regenerated plants have been used in somatic hybridization. It has been shown that hybrid cell selection on the basis of antibiotic or herbicide resistances brought by the two parents of fusion is an efficient method for the recovery of tetraploid somatic hybrids.