Small interspersed sequences that serve as recombination sites at the cox2 and atp6 loci in the mitochondrial genome of soybean are widely distributed in higher plants

Mitochondrial DNA (mtDNA) fragments that contain cox2 and atp6 were cloned from a wild soybean (Glycine soja, accession `B09002') and from a cultivated soybean (G. max, `Harosoy'). Comparison of these DNAs revealed that two sets of repeated sequences, namely, 299 bp and 23 bp, were present in the 5′ regions of cox2 and atp6. The 299-bp and 23-bp repeats were present close to each other on the 5′ flanking region and the 5′ part of the coding region of cox2 in both `Harosoy' and `B09002', as well as on the 5′ flanking region of atp6 in `Harosoy', while these two repeats were separated by a 706-bp nucleotide sequence that contained a truncated sequence of nad3 at the 5′ flanking region of atp6 in `B09002'. The mtDNA configurations upstream from atp6 and cox2 found in `Harosoy' appeared to have been generated from configurations of cox2 and atp6 found in `B09002' via recombination across the 299-bp or 23-bp repeated sequences, or vice versa, in the mitochondrial genome of the hypothetical progenitor of these plants. The 299-bp sequence was found to be interspersed in the mitochondrial genome. Eight loci were identified that include mtDNA configurations that are inter-convertible with each other via recombination across this sequence in `B09002'. Various loci on the mitochondrial genomes of higher plants that harbor segments of the 299-bp repeats in Glycine were identified. Received: 16 August / 15 December 1997     

Genic rearrangements in Phytophthora mitochondrial DNA

Summary To assess evolutionary relationships among the oomycetous fungi we have constructed a physical and genic map of the mtDNA of a broad host range strain (695T) of Phytophthora megasperma. While, like other Phytophthora species, this 43.5 kb circular genome lacks the typical oomycete large inverted repeat, a short 0.5–0.9 kb inverted repeat has been identified. Comparison of the relative order of seven genic regions with host-specific Phytophthora strains reveals both a clustering of these loci within one-third of the host-specific genomes, and two genic inversion relative to the broad host range genome. The location of the short inverted repeat suggests that at least one of the inversions is a consequence of intramolecular recombination between repeat elements.Abbreviations atp6, atp9 genes for ATP synthase subunits 6 and 9 - cox1. cox2. cox3 genes for cytochrome c oxidase subunits I, II, and III - cob gene for apocytochrome b - rns, rnl genes for small and large mitochondrial rRNAs - mtDNA mitochondrial DNA - kb kilobase pairs     

The organization of mitochondrial atp6 gene region in male fertile and CMS lines of pepper (Capsicum annuum L.)

The mitochondrial atp6 gene in male fertile (N) and CMS (S) pepper has previously been compared and was found to be present in two copies (Kim et al. in J Kor Soc Hort Sci 42:121–127 2001). In the current study, these atp6 copies were amplified by an inverse PCR technique, and the coding region as well as the 5′ and 3′ flanking regions were sequenced. The atp6 copies in CMS pepper were detected as one intact gene and one pseudogene, truncated at the 3′ coding region. When the atp6 genes in pepper were compared to other plant species, pepper, potato, and petunia all possessed a sequence of 12 identical amino acids at the 3′ extended region, which was considered a hallmark of the Solanaceae family. Northern blot analysis showed differences in mRNA band patterns between CMS and restorer lines, indicating that atp6 gene is one of the candidates for CMS in pepper. GenBank accession number: DQ126682 (atp6-1 genomic sequence common to fertile and CMS pepper), DQ126681 (Fertile atp6-2 genomic sequence), DQ126680 (CMS pseudo-atp6-2 genomic sequence)     

The complete DNA sequence of the mitochondrial genome of the dermatophyte fungus Epidermophyton floccosum

We report here the complete nucleotide sequence of the 30.9-kb mitochondrial genome of the dermatophyte fungus Epidermophyton floccosum. All genes are encoded on the same DNA strand and include seven subunits of the reduced nicotinamide adenine dinucleotide ubiquinone oxireductase (nad1, nad2, nad3, nad4, nad4L, nad5, and nad6), three subunits of cytochrome oxidase (cox1, cox2, and cox3), apocytochrome b (cob), three subunits of ATP synthase (atp6, atp8, and atp9), the small and large ribosomal RNAs (rns and rnl), and 25 tRNAs. A ribosomal protein gene (rps5) is present as an intronic ORF in the large ribosomal subunit. The genes coding for cob and cox1 carry one intron and nad5 carries two introns with ORFs. The mtDNA of E. floccosum has the same gene order as Trichophyton rubrum mtDNA, with the exception of some tRNA genes. Maximum likelihood phylogenetic analysis confirms T. rubrum as a close relative of E. floccosum. This is the first complete mitochondrial sequence of a species of the order Onygenales. This sequence is available under GenBank accession number AY916130.     

Catalog of 605 single-nucleotide polymorphisms (SNPs) among 13 genes encoding human ATP-binding cassette transporters: ABCA4, ABCA7, ABCA8, ABCD1, ABCD3, ABCD4, ABCE1, ABCF1, ABCG1, ABCG2, ABCG4, ABCG5, and ABCG8

 Single-nucleotide polymorphisms (SNPs) at some gene loci are useful as markers of individual risk for adverse drug reactions or susceptibility to complex diseases. We have been focusing on identifying SNPs in and around genes encoding drug-metabolizing enzymes and transporters, and have constructed several high-density SNP maps of such regions. Here we report SNPs at additional loci, specifically 13 genes belonging to the superfamily of ATP-binding cassette transporters (ABCA4, ABCA7, ABCA8, ABCD1, ABCD3, ABCD4, ABCE1, ABCF1, ABCG1, ABCG2, ABCG4, ABCG5, and ABCG8). Sequencing a total of 416 kb of genomic DNA from 48 Japanese volunteers identified 605 SNPs among these 13 loci: 14 in 5′ flanking regions, 5 in 5′ untranslated regions, 37 within coding elements, 529 in introns, 8 in 3′ untranslated regions, and 12 in 3′ flanking regions. By comparing our data with SNPs deposited in the dbSNP database of the National Center for Biotechnology Information (US) and with published reports, we determined that 491 (81%) of the SNPs reported here were novel. We also detected 107 genetic variations of other types among the loci examined (insertion–deletions or mono- di-, or trinucleotide polymorphisms). The high-density SNP maps we constructed on the basis of these data should provide useful information for investigating associations between genetic variations and common diseases or responsiveness to drug therapy. Received: February 26, 2002 / Accepted: March 5, 2002     

High-resolution SNP map in the 55-kb region containing the selectin gene family on chromosome 1q24–q25

 Immunoglobulin A nephropathy (IgAN) is the most common form of glomerulonephritis characterized by predominant IgA deposits in glomerular mesangium. By means of a genome-wide case-control association study, we previously demonstrated that eight single-nucleotide polymorphisms (SNPs) within the selectin gene cluster are significantly associated with IgAN. Here we provide more detailed information of variations corresponding to selectin loci, consisting of 88 SNPs and two insertion–deletion polymorphisms in the Japanese population: 27 in 5′ flanking regions, 1 in 5′ untranslated regions, 6 within coding regions, 46 in introns, 4 within 3′ untranslated regions, and 4 in 3′ flanking regions. The SNP map presented here will be a useful resource not only for examining the relationships between selectin genotypes and susceptibility to the IgAN phenotype, but also for analyzing gene scans of complex diseases mapped to this local segment on chromosome 1. Received: November 18, 2002 / Accepted: November 22, 2002 Correspondence to:Y. Nakamura     

A broad bean mitochondrial atp6 gene with an unusually simple,non-conserved 5′ region

Summary A nucleotide sequence of broad bean mitochondrial DNA (mtDNA) that contains an atp6 gene of 876 ntp is presented. Relative to other plant atp6 genes, this broad bean gene comprises a 90 ntp non-conserved 5 region, a 759 ntp highly conserved central region and a 27 ntp non-conserved 3 region. The non-conserved, 5 region of the broad bean atp6 gene differs from the corresponding regions of most other plant atp6 genes in that it contains only one potential translation initiation codon and, following this codon, a 63 ntp segment that predicts an amino acid sequence with a predominance of alternating leucines.     

Organization of a 117-kb circular mitochondrial chromosome in IR36 rice

The organization of the mitochondrial genome in the IR36 indica variety of rice was analyzed using constant-field and CHEF gel electrophoresis. The minimum complexity of the genome is estimated to be 300 kb. There was no evidence for a master circular molecule of this size. The genome exists in vivo in multiple subgenomic circular molecules or mitochondrial chromosomes. Using several rare-cutting restriction enzymes a restriction map was constructed for a 117-kb circular chromosome. Six genetic loci have been placed on this chromosome; cox1 and atp1 have been assigned precise positions while four additional genes (rrn26, cob2, atp6 and rrn18) were mapped to restriction fragments. Present address: Center for Biotechnology, SPIC Science Foundation, 110 Mount Road, Madras 600032, India     

Chimeric organization of two genes for the soybean mitochondrial ATPase subunit 6

Summary There are two copies of the ATPase subunit 6 (atp6) gene in the soybean mitochondrial genome which differ in their gene organization but share extensive homology with the maize atp6 gene except at their 5 ends. The two soybean genes are chimeric, containing regions with homology to other known mitochondrial genes at their 5 ends. Sequences homologous to the cytochrome oxidase subunit II (coxII) are located in one copy and sequences homologous to the ATPase subunit 9 (atp9) gene are located in the other copy, both of which contain methionine (ATG) codons that are in-frame with the remainder of the atp6 open reading frame. At least the copy of atp6 that contains the coxII sequence at its 5 end is abundantly transcribed to give an RNA of approximately 1,200 nucleotides.     

Nucleotide sequence and genomic organization of a newly identified member of the genus <Emphasis Type="Italic">Carmovirus</Emphasis>, soybean yellow mottle mosaic virus,from soybean

The viral genome of soybean yellow mottle mosaic virus (SYMMV) from infected soybean (Glycine max) in Korea was cloned and sequenced. The complete monopartite single-stranded RNA genome of SYMMV consists of 4009 base pairs with six putative open reading frames and includes 5′- and 3′-untranslated regions of 39 and 229 nucleotides, respectively. The nucleotide and coat protein sequences of SYMMV share the highest sequence identity with those of cowpea mottle virus. Based on its genomic organization, its predicted amino acid sequence, and its phylogenetic relatedness to known carmoviruses, we report that SYMMV is a new member of the genus Carmovirus in the family Tombusviridae.     

Catalog of 320 single nucleotide polymorphisms (SNPs) in 20 quinone oxidoreductase and sulfotransferase genes

Single nucleotide polymorphisms (SNPs) in genes encoding drug-metabolizing enzymes, transporters, receptors, and other drug targets have been widely implicated as contributors to differences among individuals as regards the efficacy and toxicity of many medications, as well as the susceptibility to complex diseases. By combining the polymerase chain reaction (PCR) technique with direct sequencing, we screened genomic DNAs from 48 Japa-nese volunteers for SNPs in genes encoding three quinone oxidoreductases (NQO1, NQO2, and PIG3) and 17 sulfotransferases (SULT1A1, SULT1A2, SULT1A3, SULT1C1, SULT1C2, SULT2A1, SULT2B1, ST1B2, TPST1, TPST2, SULTX3, STE, CST, HNK-1 ST, CHST2, CHST4, and CHST5). In all, we identified 320 SNPs from these 20 loci: 22 within coding elements, 21 in 5′ flanking regions, 10 in 5′ untranslated regions, 223 in introns, 19 in 3′ untranslated regions, and 25 in 3′ flanking regions. The ratio of transitions to transversions was approximately 2.3 to 1. Of the 22 coding SNPs, 6 were nonsynonymous substitutions that resulted in amino-acid substitutions. The high-density SNP maps we constructed from this data for each of the quinone oxidoreductases and sulfotransferases examined here should provide useful information for investigations designed to detect association(s) between genetic variations and common diseases or responsiveness to drug therapy. Received: January 12, 2001 / Accepted: January 19, 2001     

Catalog of 668 SNPs detected among 31 genes encoding potential drug targets on the cell surface

 We have been publishing a series of detailed maps of single-nucleotide polymorphisms (SNPs) detected within the genomic loci of 145 genes encoding drug-metabolizing enzymes and transporters. As an addition to the maps reported earlier, we provide here high-density SNP maps of 31 genes encoding various receptors and adhesion molecules of medical importance. By examining a total of approximately 382 kb of genomic DNA encompassing these 31 genes, we identified 668 SNPs among 48 healthy Japanese individuals: 86 in 5′ flanking regions, 27 in 5′ untranslated regions, 45 in coding regions, 399 in introns, 47 in 3′ untranslated regions, and 64 in 3′ flanking regions. We also discovered 113 variations of other types. Of the 668 SNPs, 371 (55.5%) appeared to be novel, on the basis of comparisons with the dbSNP database of the National Center for Biotechnology Information (US) or with previous publications. The maps constructed in this study will serve as an additional resource for studies of complex genetic diseases and drug-response phenotypes to be mapped by linkage-disequilibrium analyses. Received: November 6, 2002 / Accepted: November 7, 2002 Correspondence to:Y. Nakamura     

Absence of F1-ATPase activity in Kluyveromyces lactis lacking the ɛ subunit

The mitochondrial F₁-ATPase is a multimeric enzyme, comprised of 3α, 3β, γ, δ and ɛ subunits, that is primarily responsible for the synthesis of ATP in eukaryotic cells. Recent work has shown that the F₁ complex of the petite-negative yeast Kluyveromyces lactis, with specific mutations in the α, β or γ subunits, has a novel function that suppresses lethality caused by loss of mtDNA. Previously, genes for the four largest subunits of K. lactis F₁ have been identified and characterised. In this study the gene coding for the ɛ-subunit of F₁, KlATPɛ, has been isolated and found to encode a polypeptide of 61 amino acids with only 32 residues identical to those in the protein from Saccharomyces cerevisiae. Strains carrying a null mutation of KlATPɛ are respiratory deficient while the introduction of ATPɛ from S. cerevisiae restores growth on non-fermentable carbon sources. In contrast to S. cerevisiae, K. lactis disrupted in ATPɛ does not have a detectable F₁-related mitochondrial ATP hydrolysis activity, suggesting that the ɛ-subunit plays a critical role in the formation of the catalytic sector of F₁. With a disrupted KlATPɛ, the ρ^o-lethality suppressor function of F₁ carrying the atp2-1 and atp1-6 alleles is abolished. However, inactivation of the ɛ subunit does not eliminate the ρ^o-viable phenotype of the atp1-1, atp2-9, atp3-2 mutants. It is suggested that the absence of ɛ may effect the assembly or stability of F₁ in the wild-type, atp 2-1 and atp1-6 strains, whereas the defect can be suppressed by the atp1-1, atp2-9 and atp3-2 mutations in the α, β and γ subunits respectively. Received: 9 November 1999 / 11 February 2000     

Mitochondrial DNAs of the fungus Armillaria ostoyae: restriction map and length variation

A restriction-enzyme-site map is presented for the 147-kb mtDNA of North American Armillaria ostoyae. The locations of five structural genes, atp6, atp8, coxI, coxIII, and cob, along with the location and orientation of the large and small ribosomal RNA genes, were determined through Southern hybridizations with cloned genes from other fungal mtDNAs. Based on this map, the variation in mtDNA suggested geographic structure at two different levels. On a large geographic scale, 17 mtDNA types from North America were distinct, with respect to both size and restriction maps, from three mtDNA types from Europe. At the local scale, identical mtDNA types were evident among several different genetic individuals located no more than 1 km apart at a site in Michigan. No mtDNA type occurred more than once among genetic individuals from different regions of North America, although the occurrence of similar mtDNAs in isolates from distant regions suggested that this may occur at a low frequency with large sample sizes. Among the North American mtDNA types, analysis of discrete length variants was inconsistent with the hypothesis that the mtDNA of A. ostoyae evolves as a clonal lineage in which each length mutation represents a unique event. The two remaining hypotheses, that similar mutational events have occurred independently and that genetic exchange and recombination occurs among mtDNAs in natural populations of this species, remain to be tested.     

Genome-wide patterns of histone modifications in fission yeast

We have used oligonucleotide tiling arrays to construct genome-wide high-resolution histone acetylation maps for fission yeast. The maps are corrected for nucleosome density and reveal surprisingly uniform patterns of modifications for five different histone acetylation sites. We found that histone acetylation and methylation patterns are generally polar, i.e. they change as a function of distance from the ATG codon. A typical fission yeast gene shows a distinct peak of histone acetylation around the ATG and gradually decreased acetylation levels in the coding region. The patterns are independent of gene length but dependent on the gene expression levels. H3K9Ac shows a stronger peak near the ATG and is more reduced in the coding regions of genes with high expression compared with genes with low expression levels. H4K16Ac is strongly reduced in coding regions of highly expressed genes. A second microarray platform was used to confirm the 5′ to 3′ polarity effects observed with tiling microarrays. By comparing coding region histone acetylation data in HDAC mutants and wild type, we found that hos2 affects primarily the 5′ regions, sir2 and clr6 affect middle regions, and clr6 affects 3′ regions. Thus, mechanisms involving different HDACs modulate histone acetylation levels to maintain a 5′ to 3′ polarity within the coding regions. Electronic Supplementary Material Supplementary material is available for this article at and accessible for authorised users.     

Identification of sequence polymorphisms in two sulfation-related genes, PAPSS2 and SLC26A2, and an association analysis with knee osteoarthritis

Osteoarthritis (OA) is one of the most common musculoskeletal disorders and is characterized by degeneration of articular cartilage. Sulfation of extracellular ma-trix proteins in articular cartilage is an important step in maintaining normal cartilage metabolism. Two sulfation-related genes have been reported as the causal genes of severe chondrodysplasias: mutations in PAPSS2 (3′-phosphoadenosine 5′-phosphosulfate synthase 2) cause spondylo-epimetaphyseal dysplasia (SEMD), and mutations in SLC26A2 (solute carrier family 26, member 2) cause diastrophic dysplasia. Given their critical roles in cartilage metabolism and the severe phenotypes that result from mutations in these genes, we examined PAPSS2 and SLC26A2 as candidate susceptibility loci for OA. We identified sequence polymorphisms in the coding and core promoter regions of these genes and analyzed their potential association with knee OA within the Japanese population. Ten sequence polymorphisms were detected in PAPSS2 and five in SLC26A2. An association analysis showed suggestive association of one minor polymorphism in the promoter region of SLC26A2. This 4-bp adenine deletion allele, del4A, was over-represented in knee OA (P = 0.043, odds ratio = 3.43) and is thought to confer a minor susceptibility to knee OA within the Japanese population. Haplotype analysis showed no evidence of association with the two genes, however, excluding them as major susceptibility loci for knee OA. Received: May 7, 2001 / Accepted: June 21, 2001     

In vitro DNA fragmentation of mitochondrial DNA caused by single-stranded breakage related to macroplasmodial senescence of the true slime mold, Physarum polycephalum

We found that mitochondrial DNA (mtDNA) isolated from Physarum polycephalum fragmented itself in weak ionic solutions. The mtDNA was dissolved in STE (saline Tris-EDTA: 150 mM NaCl, 10 mM Tris-HCl, 1 mM EDTA), TE (10 mM Tris-HCl, 1 mM EDTA) and DW, and then electrophoresed in an agarose gel. The intact 86-kbp mtDNA band was seen in STE, but several novel bands appeared in TE and DW. In TE, two discrete bands appeared at 6.7-kbp (α-band) and 5.0-kbp (β-band), whereas at least 17 discrete bands were observed in distilled water (DW). These fragmentation patterns were not stoichiometric, as seen when using restriction endonucleases, but were clearly different from the degradation of DNA caused by a physical shearing force or a contaminating nuclease. In this paper, we characterize this in vitro fragmentation of mtDNA from P. polycephalum. We located 19 fragments, including the α and β fragments, on a mtDNA restriction map, and demonstrated that these cleavage sites were S1 nuclease-sensitive regions, which are single-stranded DNA regions such as nicks and gaps in the mtDNA. The α and β fragments are derived from the region encoding ribosomal RNAs (rRNAs) and the ATP synthase (atpA) gene, while the other 17 fragments are not derived from any specific region, but the cleavage sites are located throughout the mtDNA molecule. In P. polycephalum, it is well known that the growth rate of macroplasmodia decreases with aging. Equal amounts of mtDNA from juvenile and aged macroplasmodia were electrophoresed and the frequency of the β fragment in each sample was measured. The ratio of the β band to the total signal including background was estimated to be 3.3–4.0% in juvenile macroplasmodia, whereas it increased to 8.3–28.2% in aged macroplasmodia. This result suggests that the in vitro fragmentation of mtDNA is associated with macroplasmodial senescence. The single-stranded breakage of mtDNA of P. polycephalum may accumulate with age. Received: 16 April / 2 September 1999     

Catalog of 46 single-nucleotide polymorphisms (SNPs) in the microsomal glutathione S-transferase 1 (MGST1) gene

A major goal in our laboratory is to understand the role of common genetic variations among individual patients as regards susceptibility to common diseases and differences in therapeutic efficacy and/or side effects of drugs. As an addition to the high-density SNP (single-nucleotide polymorphism) maps of 12 glutathione S-transferase and related genes reported earlier, we provide here an SNP map of the microsomal glutathione S-transferase 1 (MGST1) gene. Among 48 healthy Japanese volunteers examined, we identified a total of 46 SNPs at this locus, 36 of which had not been reported before: 4 in the promoter region, 34 in introns, 3 in the 3′ untranslated region, and 5 in the 3′ flanking region. No SNP was found in 5′ untranslated or coding regions. The ratio of transition to transversion was approximately 1.2 : 1. Among the 13 insertion–deletion polymorphisms was a 2-bp deletion in the coding region of MGST1 in DNA from one of the volunteers, which resulted in a frame-shift mutation. Since the gene product encoded by this mutant allele would lack the C-terminal half including the MAPEG (membrane-associated proteins in eicosanoid and glutathione metabolism) domain, MGST1 activity is likely to be reduced in the carrier's cells. The SNP map presented here adds to the archive of tools for studying complex genetic diseases, population migration patterns, and a variety of pharmacogenetic possibilities. Received: June 27, 2001 / Accepted: July 16, 2001