The mitochondrial genome of the aquatic phycomycete Allomyces macrogynus. Physical mapping and mitochondrial DNA instability

Summary A physical map of the mitochondrial genome of the aquatic phycomycete Allomyces macrogynus strain Burma 3–35 (35°C) has previously been published (Borkhardt and Delius 1983). This map has been extended in this study by locating 37 additional recognition sites for five new restriction enzymes in the mitochondrial genome. Homologous regions for the genes coding for cytochrome oxidase subunits 1, 2, and 3, apocytochrome b, ATPase subunits 6 and 9, the small and large ribosomal RNA, URF1, URF5, and perhaps urfa, a presumptive gene hitherto found only in the mitochondrial genome of the fission yeast Schizosaccharomyces pombe, were located in the mitochondrial genome of A. macrogynus by heterologous hybridizations with specific, mitochondria) gene probes from Saccharomyces cerevisiae, Aspergillus nidulans, Neurospora crassa, and S. pombe. The mitochondrial gene order in A. macrogynus was found to be identical to that of A. arbuscula; a gene order hitherto found only among members of the family Blastocladiaceae. Spontaneous insertion mutations have been found to occur quite frequently in the mitochondrial genome of A. macrogynus. In all mutated mitochondrial genomes so far studied, insertions have been located in a specific region located between the genes coding for the ATPase subunit 9 and the large ribosomal RNA. In two of the mutated mitochondrial genomes the insertional event(s) resulted in the presence of mitochondrial DNA molecules differing in size by multiples of approximately 70 base pairs.     

The mitochondrial genome of the basidiomycete Agrocybe aegerita: molecular cloning,physical mapping and gene location

Summary The mtDNA of a wild-type strain of Agrocybe aegerita was purified from mitochondria isolated by cellular fractionation. A representative library was constructed in E. coli by molecular cloning at the HindIII restriction site of pBR322. Southern hybridizations between total DNA of the fungal strain and cloned mitochondrial insert probes were used to establish the restriction map of the mtDNA molecule. Its size was assessed at about 80 500 bp. Four structural genes (for Cox 1, Cox 2, Atp 6, and Atp 8) were located on the map by heterologous hybridizations with oligonucleote probes specific for yeast mitochondrial genes. The location of the genes coding for the large and the small RNAs of the mitochondrial ribosome was determined by hybridization with the E. coli rrnB operon. A comparison of A. aegerita mtDNA organization with that of both phylogenetically close and distant fungi is discussed.     

Restriction map of the mitochondrial DNA of the true slime mould,Physarum polycephalum: linear form and long tandem duplication

Summary The mitochondrial DNA (mtDNA) of the true slime mould, Physarum polycephalum strain CH934xCH938, was isolated and characterized by restriction mapping. Cloned fragments of the mtDNA were assembled and used to construct the restriction map. This map showed that the mtDNA was a linear molecule of 86.0 kb with a tandem duplication of 19.6 kb. The terminal fragments were identified by sensitivity to Bal31 exonuclease. One of the duplications was located at the right end and the other was located 5 kb from the left end. Each duplicated segment contained 26 restriction sites for ten enzymes and these restriction sites were completely conserved in each duplication. Genes for the large and small rRNAs were mapped to positions about 30 kb from the right end of the mtDNA by hybridization with its own rRNAs. With the exception of a probe for the gene for the large rRNA in Tetrahymena pyriformis mtDNA, various probes from the mtDNAs of Saccharomyces cerevisiae and T. pyriformis showed no significant hybridization to any of the restriction fragments of the mtDNA from P. polycephalum.     

Physical mapping and genome organization of mitochondrial DNA from Candida maltosa

Summary Mitochondrial (mt) DNA of the ascomycetous yeast Candida maltosa was isolated and characterized. The mtDNA is circular and the size estimated from restriction analysis performed with 7 endonucleases was 52 kb pairs. A restriction map was constructed, using the cleavage data of four endonucleases. Using mt genes from Saccharomyces cerevisiae, six structural genes (large rRNA, apocytochrome b, cytochrome c oxidase subunit I and subunit 11, ATPase subunit 6 and subunit 9) were located on the C. maltosa chondriome by cross hybridization experiments. The comparison between the mt genomes of C. maltosa and six other yeasts showed differences in the overall genome organization.     

Mitochondrial DNA in the aquatic fungus Allomyces

Summary The mitochondrial DNA from seven species of the aquatic phycomycete Allomyces has been isolated and characterized by restriction enzyme analysis. Comparison of the mitochondrial DNA restriction enzyme fragmentation patterns showed pronounced differences not only among species but also among four isolates of A. arbuscula. The mitochondrial DNAs range in size from 39 kbp in A. neo-moniliformis to 56 kbp in A. macrogynus.A physical map of the mitochondrial DNA of Allomyces arbuscula strain Costa Rica 21 has been constructed. The genome is circular and has a size of 49.2 kbp. The genes coding for the small and large ribosomal RNAs, cytochrome oxidase subunits 1, 2, and 3, apocytochrome b, and ATPase subunits 6 and 9 were localized in the mitochondria) DNA by heterologous hybridization with specific mitochondria) gene probes from Saccaromyces cerevisiae and Neurospora crassa. Comparison of the gene map of the closely related species Blastocladiella emersonii with that of A. arbuscula indicates a similar gene order in the two organisms.     

The maize mitochondrial genome of the normal type and the cytoplasmic male sterile type T have very different organization

Summary The normal type (N) and cytoplasmic male sterile type T (cmsT) maize mitochondrial genomes are 570 kb and 540 kb in length respectively. Detailed hybridization studies have been undertaken to compare the sequence complexity and variation between the two genomes (genotype B37). They have approximately 500 kb of common sequences but there is considerable variation in sequence organization which can be accounted for by structural alterations from large and small permutations. The sequences not shared between the two genomes totaling 70 kb in N and 40 kb in cmsT can be accounted for by the presence of different repeated sequences, the absence of the integrated form of plasmids S1 and S2 in cmsT, the presence or absence of chloroplast sequences, and a number of sequences that are specific to N or to cmsT.     

Linear mitochondrial genome organization in vivo in the genus Pythium

Pulsed-field gel electrophoresis (PFGE) of isolates of Pythium oligandrum with linear mitochondrial genomes revealed a distinct band in ethidium bromide-stained gels similar in size to values estimated by restriction mapping of mitochondrial DNA (mtDNA). Southern analysis confirmed that these bands were mtDNA and indicated that linear genomes were present in unit-length size as well as multimers. Isolates of this species with circular mtDNA restriction maps also had low levels of linear mono- and multimers. visualized by Southern analysis of PFGE gels. Examination of 17 additional species revealed similar results; three species had distinct linear mtDNA bands in ethidium bromide-stained gels while the remainder had linear mono- and multi-mers in lower amounts detected only by Southern analysis. Sequence analysis of an isolate of P. oligandrum with a primarily circular mitochondrial genomic map and a low amount of linear molecules revealed that the small unique region of the circular map (which corresponded to the terminal region of linear genomes) was flanked by palindromic intrastrand complementary sequences separated by a unique 194-bp sequence. Sequences with similarity to ATPase9 coding regions from other organisms were located adjacent to this region. Sequences with similarity to mitochondrial origins of replication and autonomously replicating sequences were also located in this region: their potential involvement in the generation of linear molecules is discussed.     

Characterization of inverted repeats from plasmid-like DNAs and the maize mitochondrial genome

Summary Integrated inverted repeat (IR) sequences similar to those of the S plasmids have been isolated from the genomes of the normal and S type male-sterile cytoplasms of maize mitochondria. The nucleotide sequences of both the IRs and their flanking regions have distinguished and characterized several different types of repeats. The repeats may be involved in the recombinational process that occurs continuously in the mitochondrial genome. One cloned fragment, derived from a fertile revertant and containing sequences similar to S-2, does not appear to act as atypical transposable element during reversion. Several of the flanking regions examined contain a small repeat of 34 base pairs, in which a nonanucleotide segment is found with similarity to the yeast mitochondrial promoter.     

The organization of the genes for ribosomal RNA on mitochondrial DNA of Kluyveromyces lactis

Summary We have constructed a physical map of Kluyveromyces lactis mtDNA using the restriction enzymes HindII and HindIII. In contrast to Saccharomyces, the genes for the large and small ribosomal RNAs are much closer to each other, being separated by a maximal distance of 2,250 base pairs.Abbreviations bp base pair(s) - rRNA ribosomal RNA - SSC 0.15 M NaCl, 0.015 M sodium citrate (pH 7.0)     

Physical mapping of the mitochondrial genome of the cultivated mushroom Agaricus brunnescens (= A. bisporus)

Summary Mitochondrial (mt) DNA from the commercial mushroom Agaricus brunnescens Peck [= A. bisporus (Lange) Imbach] was purified by cesium chloride/bisbenzimide gradient centrifugation. A physical map of the mtDNA fragments produced by BamHI, EcoRl, and PvuII digestion was generated by filter hybridizations with radiolabelled BamHI mtDNA probes. The A. brunnescens mtDNA was a circular molecule 136 kilo-basepairs (kbp) in length and contained an inverted repeat between 4.6 and 9.2 kbp in size. Orientational isomers of the mitochondrial genome were not detected. The positions of six genes were located on the A. brunnescens mtDNA map by heterologous hybridization. No coding function has yet been ascribed to the inverted repeat. The large rRNA gene was located on the smaller single copy region. The genes for cytochrome b, cytochrome oxidase (subunit III), ATPase (subunits 8 and 6) and the small rRNA were located on different regions of the larger single copy region.     

The mitochondrial genome of Schizosaccharomyces pombe

Summary The open reading frame of the first intron of the mitochondrial cox1 gene (cox1I1) was expressed in Escherichia coli. The putative intron-encoded protein stimulated the formation of intra-chromosomal lac ⁺-recombinants about threefold. No stimulation was found when the reading frame was inserted in the opposite direction, or when it was interrupted by a deletion. The intronic open reading frame did not complement recA ^– or recB ^– mutants of E. coli. In S. pombe, elimination of this intron did not abolish homologous recombination in mitochondria. A possible role of the recombinase activity in yeast mitochondria will be discussed.     

MboI,ThaI and HinfI endonuclease cleavage maps of the yeast mitochondrial DNA

Summary MboI, HinfI and ThaI cleavage maps have been constructed for the region of the mitochondrial DNA from S. cerevisiae where transfer RNA genes are principally located. About 40 cleavage sites have been localized between the C and P genetic markers. The MboI map covers about 50% of the total mitochondrial genome. For constructing maps we have used a series of rho^– deletion mutants whose mitochondrial DNAs have a typical single deletion structure as judged by previous genetic and physical analyses. The mutant DNAs carry known transfer RNA genes and genetic markers and, therefore, the comparison between genetic and restriction maps has allowed us to localize individual transfer RNA genes within defined physical segments.Abbreviations bp base pairs - mtDNA mitochondrial DNA - tRNA transfer RNA - rRNA ribosomal RNA - ThaI formerly TacI     

The genes for cytochrome b,ND 4L,ND6 and two tRNAs from the mitochondrial genome of the locust,Locusta migratoria

We have cloned and characterized a 2,778-kb XbaI segment of the mitochondrial genome of the locust, Locusta migratoria. It harbours portions of the ND4 and the ND1 genes, the entire genes for ND6, ND4L and cytochrome b, and the genes for three mitochondrial tRNAs. The genes are arranged in an order which is conserved between orthopteran and dipteran insects. The analysis of the cytochrome b sequence, and its comparison with other systems, supports the current model structure for this polypeptide.     

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.     

The gene for the large (16S) ribosomal RNA from the Locusta migratoria mitochondrial genome

Summary The nucleotide sequence of a segment of the mtDNA molecule of the locust Locusta migratoria containing the complete large rRNA (16S) gene and some nucleotides in its vincinity has been determined. The gene contains 1314 nucleotides, comprising the smallest metazoan gene reported to date. The gene has a low content of GC (21%) and exhibits an extended sequence homology to the corresponding gene of the dipteran insect Drosophila yakuba, suggesting a comparable secondary structure. The gene structure is discussed in an evolutionary and functional context.     

Genetic map of mitochondrial DNA in Podospora anserina

Summary In order to develop an eukaryotic vector with the Podospora plasmid, further characterization is required of the mitochondrial DNA into which this plasmid is integrated, a physical map (restriction sites) of the Podospora chondriome (size 95 kb) has been completed. As prerequisite for the establishment of a genetic (functional) map, 70% of the chondriome was cloned in E. coli vectors. Using mitochondrial genes from Saccharomyces cerevisiae, six structural genes were located on the Podospora chondriome by cross hybridization experiments. There is strong evidence that the plasmid is inserted into the cytochrome b gene. A comparison of the genetic map of the Podospora chondriome with those of Neurospora crassa and Aspergillus nidulans exhibits a rather good accordance with respect to the sequence of genes.     

Molecular analysis of the mitochondrial genome of Phytophthoraa

Summary The organization of the mitochondrial genomes from two morphologically similar Phytophthora isolates, P. megasperma f. sp. glycinea (Pmg) and P. megasperma f. sp. medicaginis (Pmm), and the morphologically different species, P. parasitica var. nicotianae (Ppn), has been studied. The mtDNAs are circular, and their estimated sizes are 45.3 kb, 41 kb, and 39.5 kb for Pmg, Pmm, and Ppn, respectively. Physical maps were constructed for restriction endonuclease sites. Four genes (l-rRNA, s-rRNA, oxi-2, and cob) were found to have the same order in the three mtDNAs.