Rice mitochondrial genome contains a rearranged chloroplast gene cluster

Summary We have previously reported the isolation and partial sequence analysis of a rice mitochondrial DNA fragment (6.9 kb) which contains a transferred copy of a chloroplast gene cluster coding for the large subunit of ribulose-1,5-bisphosphate carboxylase (rbcL), and subunits of ATPase (atpB and atpE), methionine tRNA (trnM) and valine tRNA (trnV). We have now completely sequenced this 6.9 kb fragment and found it to also contain a sequence homologous to the chloroplast gene coding for the ribosomal protein L2 (rpl2), beginning at a site 430 bp downstream from the termination codon of rbcL. In the chloroplast genome, two copies of rpl2 are located at distances of 20 kb and 40 kb, respectively, from rbcL. We have sequenced these two copies of rice chloroplast rpl2 and found their sequences to be identical. In addition, a 151 bp sequence located upstream of the chloroplast rpl2 coding region is also found in the 3 noncoding region of chloroplast rbcL and other as yet undefined locations in the rice chloroplast genome. Hybridization analysis revealed that this 151 bp repeat sequence identified in rice is also present in several copies in 11 other plant species we have examined. Findings from these studies suggest that the translocation of rpl2 to the rbcL gene cluster found in the rice mitochondrial genome might have occurred through homologous recombination between the 151 bp repeat sequence present in both rpl2 and rbcL.     

Euglena gracilis chloroplast transfer RNA transcription units. Nucleotide sequence analysis of a tRNAThr-tRNAGly-tRNAMet-tRNASer-tRNAGln gene cluster

The arrangement and the nucleotide sequence of the tRNA genes in the 2.0-kilobase-pair EcoRI restriction fragment EcoQ of Euglena gracilis Klebs, strain Z Pringsheim chloroplast DNA have been determined. This fragment, cloned in pBR325 to form the plasmid pEZC300, contains five tRNA genes. The DNA insert of this plasmid, a known tRNA gene locus (Orozco, E.M., Jr., and Hallick, R.B. (1982) J. Biol. Chem. 257, 3258-3264) has been mapped by Southern gel analysis using a 32P-labeled oligodeoxynucleotide tRNA gene probe. The DNA sequence of 870 base pairs (bp) from EcoQ containing the entire tRNA gene locus was determined. The organization of this tRNA gene cluster on the E. gracilis chloroplast chromosome is tRNAUUGGln-14-BP spacer-RNAGCUSer-175-bp spacer-tRNACAUMet-12-bp spacer-tRNAGCCGly-5-bp spacer-tRNAUGUThr. The tRNAUUGGln and tRNAGCUSer gene sequences are of the opposite polarity as the other three gene sequences, but of the same polarity as the rRNA genes. The tRNAMet gene is a putative initiator tRNA. The five tRNA genes are separated and flanked by A-T-rich spacer sequences. This gene arrangement is consistent with the model that E. gracilis chloroplast tRNA genes are transcribed into multicistronic tRNA precursors. The DNA sequences have been used to deduce the primary and secondary structures of the tRNAs.     

Euglena gracilis chloroplast transfer RNA transcription units. II. Nucleotide sequence analysis of a tRNAVal-tRNAAsn-tRNAArg-tRNALeu gene cluster

The tRNA-coding locus of the 8.2-kilobase pair (kbp) Eco RI fragments Eco G of Euglena gracilis Klebs, strain Z Pringsheim chloroplast DNA was chosen for detailed analysis. Two recombinant plasmids, pPG14, containing Eco G and the vector pMB9, and pEZC23, containing the chloroplast DNA fragment HindIII B cloned in pBR322 were employed for the study. The tRNA locus was mapped to an 0.8-kbp region of Eco G also present in HindIII B. The DNA sequence of a 1.6 kbp from HindIII B, containing the entire tRNA gene locus was determined. Four tRNA genes were identified from the DNA sequence. The gene organization is tRNAVal-16 bp spacer-tRNAAsn-3 bp spacer-tRNAArg-45 bp spacer-tRNALeu. The tRNALeu gene is of the opposite polarity as the other three genes. This is the first evidence of such a tRNA cluster for a chloroplast genome. Also evident from the DNA sequence, 132 bp from the 5'-end of the tRNALeu gene, is a putative gene or pseudogene for a chloroplast protein.     

Rice chloroplast DNA molecules are heterogeneous as revealed by DNA sequences of a cluster of genes.

We describe the isolation of two rice chloroplast HindIII fragments (9.5 kb and 5.3 kb) each containing a gene cluster coding for the large subunit of ribulose-1,5-bisphosphate carboxylase (rbcL), beta and epsilon subunits of ATPase (atpB and atpE), tRNAmet (trnM) and tRNAval (trnV). All five genes contained in the 9.5 kb fragment are potentially functional, whereas in the 5.3 kb fragment, rbcL is truncated and atpB is frame-shift mutated. The copy number of the 9.5 kb fragment is 10 times that of the 5.3 kb fragment, indicating that the two fragments are probably located on different chloroplast genomes and represent two different (major and minor) genomic populations. Thus, the rice chloroplast genome appears to be heterogeneous, contrary to general belief. We also describe the isolation of a rice mitochondrial HindIII fragment (6.9 kb) which contains an almost complete transferred copy of this chloroplast gene cluster. In this transferred copy, the coding sequences of rbcL, atpE and trnM contain perfectly normal reading frames, whereas atpB has become grossly defective and trnV is truncated.     

The mitochondrial genome of Priapulus caudatus Lamarck (Priapulida: Priapulidae)

We sequenced and annotated the complete mitochondrial (mt) genome of the priapulid Priapulus caudatus in order to provide a source of phylogenetic characters including an assessment of gene order arrangement. The genome was 14,919 bp in its entirety with few, short non-coding regions. A number of protein-coding and tRNA genes overlapped, making the genome relatively compact. The gene order was: cox1, cox2, trnK, trnD, atp8, atp6, cox3, trnG, nad3, trnA, trnR, trnN, rrnS, trnV, rrnL, trnL(yaa), trnL(nag), nad1, -trnS(nga), -cob, -nad6, trnP, -trnT, nad4L, nad4, trnH, nad5, trnF, -trnE, -trnS(nct), trnI, -trnQ, trnM, nad2, trnW, -trnC, -trnY; where '-' indicates genes transcribed on the opposite strand. The gene order, although unique amongst Metazoa, shared the greatest number of gene boundaries and the longest contiguous fragments with the chelicerate Limulus polyphemus. The mt genomes of these taxa differed only by a single inversion of 18 contiguous genes bounded by rrnS and trnS(nct). Other arthropods and nematodes shared fewer gene boundaries but considerably more than the most similar non-ecdysozoan.     

Molecular analysis of a 21.1-kb fragment of wheat chloroplast DNA bearing RNA polymerase subunit (rpo) genes.

The entire nucleotide sequence of a 21.1-kb fragment of wheat chloroplast (ct) DNA was determined. This fragment carries 18 intact genes and parts of two additional genes, including the three RNA polymerase genes rpoB, rpoC1, and rpoC2. The gene arrangement of this region is conserved in wheat, rice, and maize, but not in non-grass species. Comparison of these 20 genes in wheat, rice, and maize showed that tRNA genes evolved more slowly than protein-coding genes in the chloroplast genome. Intergenic regions evolved much faster than both types of genes. Although the 19 genes of wheat, except for orf42, showed high identity to those of other plants, there were three novel structural features in the wheat rpoC2 gene; a deletion of 81 bp in the middle region, a variable insertion (408 bp), and a nonsense mutation in the 3' terminal region, resulting in truncation of a sequence of ca. 10 amino acids. An intermolecular recombination between the stretches of CTTAT and CTTTT was suggested as the mechanism of the 81-bp deletion in the wheat rpoC2 gene. Evolutionary distance between the chloroplast genomes of wheat and maize was larger than those between wheat and rice and between rice and maize.     

The gene for the large subunit of ribulose-1,5-bisphosphate carboxylase in Euglena gracilis chloroplast DNA: location, polarity, cloning, and evidence for an intervening sequence

The gene for the large subunit (LS) of ribulose-1,5,-bisphosphate carboxylase of Euglena gracilis Z chloroplast DNA has been mapped by heterologous hybridization with DNA restriction fragments containing internal sequences from the Zea mays and Chlamydomonas reinhardii LS genes. The Euglena LS gene which has the same polarity as the Euglena rRNA genes has been located with respect to Pst I, Pvu I, and HindIII sites within the Eco RI fragment Eco A. The region of Euglena chloroplast DNA complementary to an 887 bp internal fragment from the Chlamydomonas chloroplast LS gene is interrupted by a 0.5-1.1 kbp non-complementary sequence. This is the first chloroplast protein gene located on the Euglena genome, and the first evidence for an intervening sequence within any chloroplast protein gene.     

Length and Sequence Heterogeneity in the Mitochondrial Internal Transcribed Spacer of Acanthamoeba spp.

ABSTRACT. The internal transcribed spacer (ITS) between the mitochondrial large (23S rRNA; rnl ) and small (16S rRNA; rns ) subunit ribosomal RNA genes of Acanthamoeba castellanii strain Neff was sequenced previously and was uniquely interesting because it contained tRNA genes with acceptor stem mismatches that underwent RNA editing repair. Our interest in this ITS region was to determine its phylogenetic potential in differentiating between closely related isolates. We analyzed the mitochondrial ITS region for 17 Acanthamoeba isolates and observed extensive sequence and length variability, making this region difficult to align. Acanthamoeba griffini strain S-7 had the shortest ITS (i.e. 559 base pairs [bp]) compared with Acanthamoeba palestinensis strain Reich, which had the longest (i.e. 1,360 bp). The length disparity occurred predominantly between the spacer region of the aspartic acid ( trnD ) and methionine ( trnM ) tRNA genes. Unexpectedly, this region in A. palestinensis Reich was found to contain a duplication of the trnM gene. Additionally, like A. castellanii strain Neff, all isolates examined had tRNAs with mismatches in their acceptor stem. Also, the potential for an additional type of editing not described previously for Acanthamoeba , involving purine to pyrimidine transversions was observed.     

Nicotiana chloroplast genes for components of the photosynthetic apparatus

In order to understand more fully chloroplast genetic systems, we have determined the complete nucleotide sequence (155, 844 bp) of tobacco (Nicotiana tabacum var. Bright Yellow 4) chloroplast DNA. It contains two copies of an identical 25,339 bp inverted repeat, which are separated by 86, 684 bp and 18,482 bp single-copy regions. The genes for 4 different rRNAs, 30 different tRNAs, 44 different proteins and 9 other predicted protein-coding genes have been located. Fifteen different genes contain introns.Twenty-two genes for components of the photosynthetic apparatus have so far been identified. Most of the genes (except the gene for the large subunit of ribulose-1,5-bisphosphate carboxylase/oxygenase) code for thylakoid membrane proteins. Twenty of them are located in the large single-copy region and one gene for a 9-kd polypeptide of photosystem I is located in the small single-copy region. The gene for the 32-kd protein of photosystem II as well as the gene for the large subunit of ribulose-1,5-bisphosphate carboxylase/oxygenase have strong promoters and are transcribed monocistronically while the other genes are transcribed polycistronically. We have found that the predicted amino acid sequences of six DNA sequences resemble those of components of the respiratory-chain NADH dehydrogenase from human mitochondria. As these six sequences are highly transcribed in tobacco chloroplasts, they are probably genes for components of a chloroplast NADH dehydrogenase. These observations suggest the existence of a respiratory-chain in the chloroplast of higher plants.     

Nucleotide sequence of the split tRNAUAALeu gene from Vicia faba chloroplasts: evidence for structural homologies of the chloroplast tRNALeu intron with the intron from the autosplicable Tetrahymena ribosomal RNA precursor

Summary The gene encoding the tRNA _UAA ^Leu from broad bean chloroplasts has been located on a 5.1 kbp long BamHI fragment by analysis of the DNA sequence of an XbaI subfragment. This gene is 536 bp long and is split in the anticodon region. The 451 bp long intron shows high sequence homology over about 100 bp from each end with the corresponding regions of the maize chloroplast tRNA _UAA ^Leu intron. These conserved sequences are probably involved in the splicing reaction, for they can be folded into a secondary structure which is very similar to the postulated structure of the intron from the autosplicable ribosomal RNA precursor of Tetrahymena. Very little sequence conservation is found in the 5-and 3-flanking regions of the broad bean and maize chloroplast tRNA _UAA ^Leu genes.