Divergence of satellite DNA and interspersion of dispersed repeats in the genome of the wild beet Beta procumbens

Several repetitive sequences of the genome of Beta procumbens Chr. Sm., a wild beet species of the section Procumbentes of the genus Beta have been isolated. According to their genomic organization, the repeats were assigned to satellite DNA and families of dispersed DNA sequences.The tandem repeats are 229–246 bp long and belong to an AluI restriction satellite designated pAp11. Monomers of this satellite DNA form subfamilies which can be distinguished by the divergence or methylation of an internal restriction site. The satellite is amplified in the section Procumbentes, but is also found in species of the section Beta including cultivated beet (Beta vulgaris). The existence of the pAp11 satellite in distantly related species suggests that the AluI sequence family is an ancient component of Beta genomes and the ancestor of the diverged satellite subfamily pEV4 in B. vulgaris. Comparative fluorescent in-situ hybridization revealed remarkable differences in the chromosomal position between B. procumbens and B. vulgaris, indicating that the pAp11 and pEV4 satellites were most likely involved in the expansion or rearrangement of the intercalary B. vulgaris heterochromatin.Furthermore, we describe the molecular structure, and genomic and chromosomal organization of two repetitive DNA families which were designated pAp4 and pAp22 and are 1354 and 582 bp long, respectively. The families consist of sequence elements which are widely dispersed along B. procumbens chromosomes with local clustering and exclusion from distal euchromatic regions. FISH on meiotic chromosomes showed that both dispersed repeats are colocalized in some chromosomal regions. The interspersion of repeats of the pAp4 and pAp22 family was studied by PCR and enabled the determination of repeat flanking sequences. Sequence analysis revealed that pAp22 is either derived from or part of a long terminal repeat (LTR) of an Athila-like retrotransposon. Southern analysis and FISH with pAp4 and pAp22 showed that both dispersed repeats are species-specific and can be used as DNA probes to discriminate parental genomes in interspecific hybrids. This was tested in the sugar beet hybrid PRO1 which contains a small B. procumbens chromosome fragment.     

The mitochondrial genome of fertile maize (Zea mays L.) contains two copies of the gene encoding the α-subunit of the F1-ATPase

Summary In contrast to the situation in animals and fungi the -subunit of the mitochondrial F₁-ATPase is encoded by two identical mitochondrial genes (ATP A) in male fertile maize (Zea mays L.). Cytoplasmic male sterile (T, C and S) maize mitochondrial genomes only contain a single copy of the gene. Sequence analysis reveals that the uninterrupted coding region of both copies of the gene is 1,524 by long and encodes a polypeptide of 508 amino acids with a molecular weight of 55,117. The predicted amino acid sequence shares over 60% homology with the nuclear encoded -subunit from yeast and bovine ATPases and approx. 50% with the corresponding chloroplast and bacterial polypeptides.     

Organization and nucleotide sequence of ribosomal RNA genes on a circular 73 kbp DNA from the colourless flagellate Astasia longa

Summary Three tandemly arranged repeats (A, B, C) of 16S and 23S rDNA, and one supplementary (S) 16S rDNA adjacent to the 16S rDNA of repeat A, are present within an 18 kbp segment of a circular 73 kbp DNA from the colourless flagellate Astasia longa. The repeat units are separated by a short region containing a 5S rRNA gene and a gene for tRNA-Val (UAC). Sequence comparisons reveal 78%, 81%, and 67% identical nucleotides of the 23S rDNA (A), the 16S rDNA (B), and the 5S rDNA (A), respectively, with the corresponding genes of the Euglena gracilis chloroplast genome. As in Euglena chloroplasts, the 3-terminal protion of the 23S rDNA is homologous to the 4.5S rRNA gene of higher plant chloroplast genomes. These results are supportive of a common evolutionary origin for the Astasia 73 kbp DNA and the Euglena 145 kbp chloroplast DNA.     

The chloroplast gene cluster containing psbF,psbL, petG and rps3 is conserved in Chlamydomonas

We have sequenced a 6.8-kb segment of the Chlamydomonas eugametos chloroplast DNA which contains the psbF, psbL, petG and rps3 genes. As in the distantly related green alga Chlamydomonas reinhardtii, these genes reside in this order (53) on the same DNA strand, suggesting that such a chloroplast gene cluster was present in the most recent common ancestor of all Chlamydomonas species. For each of the four genes, with the exception of rps3, the C. eugametos and C. reinhardtii coding regions were found to be identical, or very similar, in length, whereas each of the intergenic spacers is substantially longer in C. eugametos than in C. reinhardtii. The central portion of both Chlamydomonas rps3 genes features a long extra coding region relative to other rps3 sequences. We have shown that the insertion sequence in the C. eugametos rps3 is not excised at the RNA level.     

Nucleotide sequence and determination of the extremities of the 26S ribosomal RNA gene in wheat mitochondria: evidence for sequence rearrangements in the ribosomal genes of higher plants

Summary The nucleotide sequence of the wheat mitochondrial 26S ribosomal RNA gene and flanking regions was determined and compared with mitochondrial 26S rRNA genes from maize and Oenothera. All three genes exhibit a high degree of homology except within two variable regions. When the plant mitochondrial 26S rRNA genes are compared with Escherichia coli 23S rRNA and chloroplast 23S and 4.5S rRNA genes, a third variable region is apparent close to the 3 end of the gene. The 5 and 3 ends of the wheat mitochondrial gene were determined by S1 nuclease mapping. Computer analysis of the wheat mitochondrial gene revealed several small sequences present either in the 5 region of the 26S rRNA gene or in the 18S rRNA gene.     

The two genes for the P700 chlorophyll a apoproteins on the Euglena gracilis chloroplast genome contain multiple introns

Summary We have determined the complete nucleotide sequence of the two genes encoding the P₇₀₀ chlorophyll a apoproteins of the photosystem I reaction center of the Euglena gracilis chloroplast genome. The two genes are separated by 77 bp, are of the same polarity, and span a region which is greater than 9.0 kbp. The psaA gene (751 codons) is interrupted by three introns and the psaB gene (734 codons) by six introns. The introns range in size from 361 to 590 bp, whereas the exons range in size from 42 to 1,194 bp. The introns are extremely AT rich with a pronounced base bias of T > A > G > C in the RNA-like strand. Like other interrupted protein genes in the Euglena chloroplast genome, the psaA and psaB introns are similar to mitochondrial group II introns in having the splice junction consensus sequence, 5 GTGCGNTTCG ..... INTRON ..... TTAATTTTAT 3 and conserved secondary structural features. Except for the placement of the first intron, the intron-exon organization of these two highly homologous genes is not conserved. The other introns fall at or near putative surface domains of the predicted gene products. The psaA and psaB gene products are 74% homologous to one another and 93% and 95% homologous, respectively, to the psaA and psaB gene products of higher plant chloroplasts. The predicted secondary structure derived from the primary amino acid sequence has 11 potential membrane-spanning domains. Abbreviations and notations: Gene names follow the convention of Hallick and Bottomley (1983: psaA, psaB, genes for the P₇₀₀ apoprotein; psbE an psbF, genes foe the subunits of cytochrome b ₅₅₉; orfN, open reading frame of N condons     

Polymorphism of the mitochondrial L-RNA gene of the basidiomycete Coprinus cinereus

Summary Two unexpectedly small mitochondrial (mt) genomes of Coprinus cinereus, P and S, were compared with the H and J genomes we have described previously. H and J are 42 kb in size and differ in having alternative 1.23 kb insertions in or adjacent to the co-1 gene. P and S DNAs lacked both insertions and had an identical 4.4 kb deletion between the co-1 and L-RNA gene. P DNA contained a 700 by insertion and S DNA a 300 by deletion within a sequence coding the L-RNA gene. This was shown by Southern blot analysis using probes containing the 5 or the 3 exon sequences of the L-RNA gene of Neurospora crassa. These hybridisations showed also that the L-RNA gene and co-1 gene in the C. cinereus mt genome are oppositely orientated and must be transcribed from different DNA strands. No DNA homology was detected using probes containing intron sequences from the L-RNA genes of Saccharomyces cerevisiae or N. crassa. There was no evidence of respiratory deficiency in P and S strains and transfer of nuclei by dikaryon formation made it possible to recombine H nuclei with P and S mitochondria, S nuclei with H and P mitochondria and P nuclei with H mitochondria with no apparent detrimental effect on growth. We conclude that P and S mtDNAs represent naturally occurring variants of the C. cinereus mt genome.     

Gene structure and expression of a novel Euglena gracilis chloroplast operon encoding cytochrome b6 and the β and ε subunits of the H+-ATP synthase complex

The genes encoding cytochrome b6 of the chloroplast cytochrome b6/f complex (petB) and the ATP synthase CF₁- subunit (atpB) and -subunit (atpE) were identified on the EcoD fragment of the Euglena gracilis chloroplast genome. The complete nucleotide sequence of these three genes was determined. The petB-atpB-atpE genes are cotranscribed as a tricistronic operon. This gene organization differs from that of land plants in which atpB-atpE form a discistronic operon, and petB is within the psbB-ycf8-psbH-petB-petD operon. Euglena cytochrome b6 and the -subunit of the chloroplast ATP synthase are very similar in derived amino acid sequence to the corresponding gene products from other organisms. The -subunit of the chloroplast ATP synthase complex is more divergent. In Euglena, the petB-atpB-atpE genes contain introns, including two twintrons, at eight different positions. All of the intron positions were confirmed by analysis of cDNAs. Two independent intercistronic RNA processing events and 11 splicing reactions lead to the accumulation of the mature petB, atpB and atpE monocistronic mRNAs.     

The cytoplasmic male-sterility (CMS) determinant of common bean is widespread in Phaseolus coccineus L. and Phaseolus vulgaris L.

To identify regions of the mitochondrial genome that potentially could specify cytoplasmic male sterility (CMS) in Phaseolus coccineus (including P. polyanthus), and to define differences amongst P. coccineus lines, mitochondrial (mt)DNA restriction patterns and Southern blots of total DNA from sterile and fertile lines were analysed. By restriction endonuclease mapping we isolated a region which was specific to CMS lines flanking an F1-ATPase -subunit (atpA) gene. DNA sequence analysis of this region showed 99.9% homology to the region previously isolated from P. vulgaris CMS Sprite. A high frequency of plants carrying the CMS-fragment was observed in a wild Phaseolus population, perhaps explaining the occurrence of inter- and intraspecific gene flow observed in the autogamous species P. vulgaris.     

Biased inheritance of optional insertions following mitochondrial genome recombination in the basidiomycete fungus Coprinus cinereuss

Summary Two mitochondrial genomes of Coprinus cinereus, H and J, were found to have alternative 1.23 kb insertions. Using the Neurospora crassa cytochrome oxidase-1 (co-1) gene as a probe, the J insertion site was shown to be located within the Coprinus co-1 gene, whereas the H insertion was some 2 kb distant. The insertions showed biased inheritance following mitochondrial genome recombination. Recombination between H and J genomes was detected using the mitochondrial gene mutations acu-10, which causes a cytochrome oxidase defect, and cap-1, which confers chloramphenicol resistance. Fourteen of fifteen independently derived recombinants for these two genes were shown to have both DNA insertions. In a second series of H x J crosses, intragenic recombination between different cap-1 alleles was detected. These mutations are assumed to be in the large ribosomal RNA gene some 6 kb distant from the nearest insertion site. Each of eight independently derived cap-1 ⁺ recombinants had both DNA insertions. Despite their similar size and similar behaviour following recombination the insertions do not share extensive sequence homology.     

Organization and structure of plastome psbF,psbL, petG and ORF712 genes in Chlamydomonas reinhardtii

Summary We have determined the nucleotide sequence of a 5159 base-pair (bp) region of the Chlamydomonas reinhardtii plastome containing three photoelectron transport genes, psbF, psbL and petG, and an unusual open reading frame, ORF712. The photosynthetic genes have an unprecedented arrangement. psbF and psbL are located in close proximity to petG, and are not grouped with two other genes of the cytochrome b₅₅₉ locus, psbE and ORF42. ORF712, located adjacent to psbL, has homology at its 5-and 3-ends to the ribosomal protein rps3 gene, but contains, a central 437 residue domain that lacks similarity to any other known sequence. These sequences add to the growing body of evidence that the chloroplast genome of C. reinhardtii has a significantly different gene arrangement to its counterpart in plants. The structure of ORF712 also provides another example of a phenomenon we have discovered with C. reinhardtii RNA polymerase genes (Fong and Surzycki 1992); namely, that the algal plastome contains chimeric genes in which reading frames with homology to known genes are juxtaposed in-frame with long coding regions of unknown identity.     

A truncated recombination repeat in the mitochondrial genome of a Petunia CMS line

Repeated sequences known as recombination repeats are present in the majority of plant mitochondrial genomes. Two recombination repeat sequences from Petunia have been analyzed. The two repeats are virtually identical over 1.42 kb. One of the repeats is truncated and is likely to have arisen from a rare recombination event in the full-length repeat. Two sequence-blocks within the Petunia repeat are highly similar to sequences in the 5 flank of several plant mitochondrial genes. No sequence motifs are shared by the Petunia repeat and other sequenced plant mitochondrial recombination repeats, suggesting that the recombination occurs by an homologous, rather than a site-specific, mechanism.     

High-frequency inter-parental recombination between mitochondrial genomes of rice cybrids

Analyzing more than 100 independent rice cybrids, we found evidence for inter-molecular recombination between parental mitochondrial genomes occurring at high frequency soon after protoplast fusion. The structure of the region around the atp6 gene showed extensive polymorphism among Indica (MTC-5A), Japonica (Nipponbare), and wild abortive (IR58024A) mitochondrial genomes. Recombination between the mitochondrial genomes of IR58024A and MTC-5A around the atp6 gene was detected by Southern-blot analysis of cybrid plants. Such recombinant mitochondrial molecules were also cloned from IR58024A/Nipponbare cybrid callus. PCR analysis around the atp6 gene demonstrated that inter-parental recombination occurs in practically all cybrid calli within 2 weeks after protoplast fusion. At this point, parental and recombinant mitochondrial genomes coexisted within the callus. Over the course of further cultivation, however, mitochondrial genome diversity decreased as parental and/or recombinant genomes segregated out.     

Chloroplast RNA polymerase genes of Chlamydomonas reinhardtii exhibit an unusual structure and arrangement

Summary Nucleotide sequence analysis of a 17043 basepair (bp) region of the Chlamydomonas reinhardtii plastome indicates the presence of three open reading frames (ORFs) similar to RNA polymerase subunit genes. Two, termed rpoB1 and rpoB2, are homologous to the 5-and 3-halves of the Escherichia coli beta subunit gene, respectively. A third, termed rpoC2, is similar to the 3-half of the bacterial beta' subunit gene. These genes exhibit several unusual features: (1) all three represent chimeric structures in which RNA polymerase gene sequences are juxtaposed in-frame with long sequences of unknown identity; (2) unlike their counterparts in plants and eubacteria, rpoB1 and rpoB2 are separated from rpoC2 by a long (7 kilobase-pair, kbp) region containing genes unrelated to RNA polymerase; (3) DNA homologous to the 5 half of rpoC (termed rpoC1 in other species) is not present at the 5 end of rpoC2 and could not be detected in C. reinhardtii chloroplast DNA. RNA expression could not be detected for any of the RNA polymerase genes, suggesting that they are pseudogenes or genes expressed at stages of the C. reinhardtii life-cycle not investigated. The three genes are flanked by GC-rich repeat elements. We suggest that repeat DNA-mediated chloroplast recombination events may have contributed to their unusual arrangement.     

Organization of ribosomal protein genes rp123, rp12, rps19, rp122 and rps3 on the Euglena gracilis chloroplast genome

Summary The nucleotide sequence (4,814 bp) was determined for a cluster of five ribosomal protein genes and their DNA flanking regions from the chloroplast genome of Euglena gracilis. The genes are organized as rp123 — 150 by spacer — rpl2 — 59 by spacer —rps19 — 110 by spacer — rp122 — 630 by spacer — rps3. The genes are all of the same polarity and reside 148 bp downstream from an operon for two genes of photosystem I and four genes of photosystem II. The Euglena ribosomal protein gene cluster resembles the S-10 ribosomal protein operon of Escherichia coli in gene organization and follows the exact linear order of the analogous genes in the tobacco and liverwort chloroplast genomes. The number and positions of introns in the Euglena ribosomal protein loci are different from their higher plant counterparts. The Euglena rp123, rps19 and rps3 loci are unique in that they contain three, two and two introns, respectively, whereas rp12 and rp122 lack introns. The introns found in rpl23 (106, 99 and 103 bp), rps19 (103 and 97 bp) and rps3 intron 2 (102 bp) appear to represent either a new class of chloroplast intron found only in constitutively expressed genes, or possibly a degenerate version of Euglena chloroplast group II introns. They are deficient in bases C and G and extremely rich in base T, with a base composition of 53–76% T, 25–34% A, 3–10% G and 2–7% C in the mRNA-like strand. These six introns show minimal resemblance to group IT chloroplast introns. They have a degenerate version of the group II intron conserved boundary sequences at their 5 and 3 ends. No conserved internal secondary structures are apparent. By contrast, rps3 intron 1 (409 bp) has a potential group II core secondary structure. The five genes, rpl23 (101 codons), rpl2 (278 codons), rpsl9 (95 codons), rpl22 (114 codons) and rps3 (220 codons) encode lysine-rich polypeptides with predicted molecular weights of 12,152, 31,029, 10,880, 12,819, and 25,238, respectively. The Euglena gene products are 18–50%, and 29–58% identical in primary structure to their E. coli and higher plant counterparts, respectively. Oligonucleotide sequences corresponding to Euglena chloroplast ribosome binding sites are not apparent in the intergenic regions. Inverted repeat sequences are found in the upstream flanking region of rp123 and downstream from rps3. Abreviations: Gene names follow the convention of Hallick and Bottomley (1983): rp123, rpl2, rpl22 are, respectively, genes for the L23, L2, L22 polypeptides of the 50S ribosomal subunit; rps19 and rps3 are genes for the S19 and S3 polypeptides of the 30S ribosomal subunit     

The mitochondrial DNA of land plants: peculiarities in phylogenetic perspective

Land plants exhibit a significant evolutionary plasticity in their mitochondrial DNA (mtDNA), which contrasts with the more conservative evolution of their chloroplast genomes. Frequent genomic rearrangements, the incorporation of foreign DNA from the nuclear and chloroplast genomes, an ongoing transfer of genes to the nucleus in recent evolutionary times and the disruption of gene continuity in introns or exons are the hallmarks of plant mtDNA, at least in flowering plants. Peculiarities of gene expression, most notably RNA editing and trans-splicing, are significantly more pronounced in land plant mitochondria than in chloroplasts. At the same time, mtDNA is generally the most slowly evolving of the three plant cell genomes on the sequence level, with unique exceptions in only some plant lineages. The slow sequence evolution and a variable occurrence of introns in plant mtDNA provide an attractive reservoir of phylogenetic information to trace the phylogeny of older land plant clades, which is as yet not fully resolved. This review attempts to summarize the unique aspects of land plant mitochondrial evolution from a phylogenetic perspective.     

Structure and expression of tomato mitochondrial genes coding for tRNACys (GCA), tRNAAsn (GUU) and tRNATyr (GUA): A native tRNACys gene is present in dicot plants but absent in monocot plants

Summary The nucleotide sequences of tRNA^Asn (GUU) and tRNA^Tyr (GUA) genes from tomato mitochondria and their flanking regions have been determined. The tomato mitochondrial tRNA^Asn gene is located 2.1 kb downstream from the tRNA^Cys gene reported previously (Izuchi and Sugita 1989) and shows a nearly complete identity with the corresponding chloroplast gene. The tRNA^Tyr gene, which shows only 73% homology with the corresponding chloroplast gene, has to be considered a native mitochondrial tRNA gene and is 535 bp from the chloroplast-like tRNA^Asn gene on the same strand. Northern hybridization analysis revealed that the three tRNA genes are transcribed in tomato mitochondria. Southern hybridization analysis of tomato, sugar beet, rice and wheat mitochondrial DNAs, with oligonucleotide probes for mitochondrial or chloroplast tRNA genes, demonstrated that the mitochondrial tRNA^Cys gene found in tomato is present in dicot plants but not in monocots. On the other hand, a chloroplast-like tRNA^Cys gene exists in monocot plants.     

Pea chloroplast genes encoding a 4kDa polypeptide of photosystem I and a putative enzyme of C1 metabolism

Summary The nucleotide sequence of 3.2 kbp of pea chloroplast DNA located upstream from the petA gene for cytochrome f, and previously reported to contain the gene for a photosystem I polypeptide, has been determined. Three open reading frames of 587, 40 and 157 codons have been identified. Orf40 encodes a highly conserved, hydrophobic, membrane-spanning polypeptide, and is identified as the gene psaI for the 4 kDa subunit of photosystem I. Orf587 is an extended version of the gene zfpA previously identified as encoding a conserved putative zinc-finger protein. The product of orf587 shows extensive homology to an unidentified open reading frame cotranscribed with a gene for folate metabolism in Escherichia coli and local homology to a region of the subunit of rat mitochondrial propionyl-CoA carboxylase. It is suggested that the product of orf587 is an enzyme of C₁ metabolism and is unlikely to be a regulatory DNA-binding protein. Orf157 potentially encodes an unidentified basic protein, but the protein sequence is not conserved in other plants.