Normal Mitochondrial Genomes Research Articles

Identification of a gene responsible for cytoplasmic male-sterility in onions (Allium cepa L.) using comparative analysis of mitochondrial genome sequences of two recently diverged cytoplasms.

Almost identical mitochondrial genome sequences of two recently diverged male-fertile normal and male-sterile CMS-T-like cytoplasms were obtained in onions. A chimeric gene, orf725 , was found to be a CMS-inducing gene. In onions (Allium cepa L.), cytoplasmic male-sterility (CMS) has been widely used in hybrid seed production. Two types of CMS (CMS-S and CMS-T) have been reported in onions. A complete mitochondrial genome sequence of the CMS-S cytoplasm has been reported in our previous study. Draft mitochondrial genome sequences of male-fertile normal and CMS-T-like cytoplasms are reported in this study. Raw reads obtained from normal and CMS-T-like cytoplasms were assembled into eight and nine almost identical contigs, respectively. After connection and reorganization of contigs by PCR amplification and genome walking, four scaffold sequences with total length of 339 and 180bp were produced for the normal cytoplasm. A mitochondrial genome sequence of the CMS-T-like cytoplasm was obtained by mapping trimmed reads of CMS-T onto scaffold sequences of the normal cytoplasm. Compared with the CMS-S mitochondrial genome, the normal mitochondrial genome was highly rearranged with 31 syntenic blocks. A total of 499 single nucleotide polymorphisms (SNPs) or insertions/deletions were identified in these syntenic regions. On the other hand, normal and CMS-T-like mitochondrial genome sequences were almost identical except for orf725, a chimeric gene consisting of cox1 with other sequences. Only three SNPs were identified between normal and CMS-T-like syntenic sequences. These results indicate that orf725 is likely to be the casual gene for CMS induction in onions and that CMS-T-like cytoplasm has recently diverged from the normal cytoplasm by introduction of orf725.

Evolution Along the Mutation Gradient in the Dynamic Mitochondrial Genome of Salamanders

Mitochondria are intracellular organelles where oxidative phosphorylation is carried out to complete ATP synthesis. Mitochondria have their own genome; in metazoans, this is a small, circular molecule encoding 13 electron transport proteins, 22 tRNAs, and 2 rRNAs. In invertebrates, mitochondrial gene rearrangement is common, and it is correlated with increased substitution rates. In vertebrates, mitochondrial gene rearrangement is rare, and its relationship to substitution rate remains unexplored. Mitochondrial genes can also show spatial variation in substitution rates around the genome due to the mechanism of mtDNA replication, which produces a mutation gradient. To date, however, the strength of the mutation gradient and whether movement along the gradient in rearranged (or otherwise modified) genomes impacts genic substitution rates remain unexplored in the majority of vertebrates. Salamanders include both normal mitochondrial genomes and independently derived rearrangements and expansions, providing a rare opportunity to test the effects of large-scale changes to genome architecture on vertebrate mitochondrial gene sequence evolution. We show that: 1) rearranged/expanded genomes have higher substitution rates; 2) most genes in rearranged/expanded genomes maintain their position along the mutation gradient, substitution rates of the genes that do move are unaffected by their new position, and the gradient in salamanders is weak; and 3) genomic rearrangements/expansions occur independent of levels of selective constraint on genes. Together, our results demonstrate that large-scale changes to genome architecture impact mitochondrial gene evolution in predictable ways; however, despite these impacts, the same functional constraints act on mitochondrial protein-coding genes in both modified and normal genomes.

The cytoplasmic male-sterile type and normal type mitochondrial genomes of sugar beet share the same complement of genes of known function but differ in the content of expressed ORFs.

The complete nucleotide sequence (501,020 bp) of the mitochondrial genome from cytoplasmic male-sterile (CMS) sugar beet was determined. This enabled us to compare the sequence with that previously published for the mitochondrial genome of normal, male-fertile sugar beet. The comparison revealed that the two genomes have the same complement of genes of known function. The rRNA and tRNA genes encoded in the CMS mitochondrial genome share 100% sequence identity with their respective counterparts in the normal genome. We found a total of 24 single nucleotide substitutions in 11 protein genes encoded by the CMS mitochondrial genome. However, none of these seems to be responsible for male sterility. In addition, several other ORFs were found to be actively transcribed in sugar beet mitochondria. Among these, Norf246 was observed to be present in the normal mitochondrial genome but absent from the CMS genome. However, it seems unlikely that the loss of Norf246 is causally related to the expression of CMS, because previous studies on mitochondrial translation products failed to detect the product of this ORF. Conversely, the CMS genome contains four transcribed ORFs (Satp6presequence, Scox2-2 , Sorf324 and Sorf119) which are missing from the normal genome. These ORFs, which are potential candidates for CMS genes, were shown to be generated by mitochondrial genome rearrangements.

Aep3p Stabilizes the Mitochondrial Bicistronic mRNA Encoding Subunits 6 and 8 of the H+-translocating ATP Synthase of Saccharomyces cerevisiae

Subunits 6 and 8 of the mitochondrial ATPase in Saccharomyces cerevisiae are encoded by the mitochondrial genome and translated from bicistronic mRNAs containing both reading frames. The stability of the two major species of ATP8/6 mRNA, which differ in the length of the 5'-untranslated region, depends on the expression of several nuclear-encoded factors. In the present study, the product of the gene designated AEP3 (open reading frame YPL005W) is shown to be required for stabilization and/or processing of both ATP8/6 mRNA species. In an aep3-disruptant strain, the shorter ATP8/6 mRNA was undetectable, and the level of the longer mRNA was reduced to approximately 35% that of wild type. Localization of a hemagglutinin-tagged version of Aep3p showed that the protein is an extrinsic constituent of the mitochondrial inner membrane facing the matrix.

New approaches to the treatment of mitochondrial disorders

Mitochondrial disorders are among the most common inherited metabolic diseases and the issue of treatment arises on a regular basis. There is no established treatment for mitochondrial disorders and current management is largely supportive, but recent advances in our understanding of the pathophysiology provide hope for novel treatments. Patients with mitochondrial myopathy due to mutations of mitochondrial DNA (mtDNA) may benefit from treatments that move normal mitochondrial genomes from the muscle satellite cells into skeletal muscle, but there are concerns about the long-term effects of this approach. A greater understanding of the pathophysiology of a number of nuclear genetic mitochondrial disorders suggests new avenues for treatment (such as copper-histidine in children with SCO2 gene mutations, and strategies modifying intra-mitochondrial nucleoside pools in the various disorders of mtDNA maintenance). A number of different strategies are also being explored at the molecular level, including the use of antigenomic molecules to mutated mtDNA and the allotropic expression of mutated mtDNA genes within the cell nucleus. Nuclear transfer techniques also provide hope for women at risk of transmitting pathogenic mtDNA mutations.

Alterations in organization and transcription of the mitochondrial genome of cytoplasmic male sterile sugar beet (Beta vulgaris L.).

We have constructed a physical map of the mitochondrial DNA of a cytoplasmic male sterile (CMS) sugar beet line, TK81-MS, and compared it with that published for normal fertile sugar beet (cv. TK81-O) to clarify the differences between the CMS and normal mitochondrial genomes. The TK81-MS genome is present as a single circular molecule of 481.8 kb, or as two molecules of 184.9 and 296.9 kb. The CMS genome was found to be highly rearranged relative to the normal mitochondrial genome, with at least fifteen rearrangement and/or inversion events being required to align the two DNAs. Analysis of transcription patterns of known mitochondrial genes and rearranged regions revealed six genes, coxI, coxII, atpA, atp6, rps3, and orf324, whose expression is altered in the CMS line relative to the normal line. Of these six, only the coxI transcript pattern differs between male-sterile and fertility-restored genotypes, making it likely that the coxI locus is involved in mediating CMS in sugar beet.

Heteroplasmy and homoplasmy for maize mitochondrial mutants: a rare homoplasmic nad4 deletion mutant plant

The nonchromosomal stripe (NCS) mutants of maize are a set of mitochondrial deletion mutants. In aerobic organisms, deletions of essential mitochondrial genes are lethal. Therefore, most plants carrying deletions of part of essential mitochondrial genes survive only heteroplasmically; that is, the plants contain mixtures of the deleted and normal mitochondrial genomes. The severity of the mutant phenotype depends on which mitochondrial gene is altered, which also influences the extent of homoplasmic tissues on the plants. NCS2 plants carry a partial deletion of the nad4 gene that codes for a subunit of complex I. Unlike animals, plants have additional enzymes that can partially compensate for the loss of complex I function. Kernels that are homoplasmic for the nad4 deletion mutation usually abort, although we report that homoplasmic mutant cultures can be initiated and maintained from immature embryos of such kernels. We also report the first example of a rare homoplasmic mutant NCS2 plant. The plant is very short, pale green, with a reduced number of narrow leaves. It is apparently male and female sterile. The possibility that the kernel that gave rise to this plant had a heteroplasmic endosperm is discussed.

Oxygen Stress Induces an Apoptotic Cell Death Associated with Fragmentation of Mitochondrial Genome

We have investigated the role of mitochondria on an active cell death, a feature of apoptosis, under the oxygen stress. An immortalized human fibroblast cell line (ρ+) carrying normal mitochondrial genome (mtDNA) underwent an active cell death in 95% oxygen; 68% and 84% of the cells died on the 3rd and 4th days, respectively. By contrast, its derivative lacking mtDNA (ρ0) exhibited a marked resistance to cell death. PCR analyses using 180 primer pairs covering the entire regions of the mtDNA revealed extensive fragmentations of the mtDNA; 49 types of deletions increased up to 187 on the 3rd day. These results indicate that mtDNA and its fragmentation are the underlying molecular lesions in a cell death pathway induced by the oxygen stress.

Mitochondrial myopathy: correlation between oxidative defect and mitochondrial DNA deletions at single fiber level.

In situ hybridization combined with immunohistochemical techniques has been applied to study patients affected by mitochondrial myopathies with large mitochondrial (mt)DNA deletions. All patients' muscle biopsies showed ragged red fibers (RRFs) and cytochrome oxidase (COX) deficiency. Two digoxigenin-labeled, polymerase chain reaction (PCR)-amplified DNAs were used as probes. One probe was designed to hybridize only with wild-type mtDNAs, while the other recognized both wild-type and deleted mtDNAs. Concomitant immunocytochemical analysis using antibodies against subunits II, III, (encoded by mtDNA) and IV (encoded by nuclear DNA) of COX was carried out. In our patients deleted mtDNAs are overexpressed in COX-negative RRFs, while wild-type mtDNAs are decreased in the same fibers. Immunohistochemistry studies show that COX IV is overexpressed in RRFs and that COX II and COX III subunits are still present. Deleted mtDNAs are spatially segregated in muscle fibers, where they interfere with the local population of normal mitochondrial genomes, causing a regional deficiency of the mitochondrial respiratory activity.

Wolfram syndrome: a mitochondrial-mediated disorder?

Mitochondrial DNA mutations cause several human diseases, (eg, Leber's hereditary optic neuropathy). Wolfram syndrome (characterised by diabetes insipidus, diabetes mellitus, optic atrophy, and deafness) also has, in some cases, a mitochondrial origin. The disease, often familial, has been well documented as an autosomal recessive disorder, and most of the clinical phenotypes are consistent with an ATP supply defect that is often seen in mitochondrial-mediated disorders. We propose a dual genome defect model for Wolfram syndrome in which nuclear genetic defects or mitochondrial genetic defects can independently lead to the disease. This model suggests that besides a mitochondrial gene defect alone, a nuclear gene defect, which interferes with the normal function of mitochondria (probably with a normal mitochondrial genome), can also be the underlying explanation for the pleiotropic features of Wolfram syndrome. This hypothesis explains how an autosomal recessive disorder can result in mitochondrial dysfunction, and has a general application in the identification of candidate genes for the various important phenotypes (eg, deafness and diabetes mellitus) seen in mitochondrial disorders.

Evidence for intramitochondrial complementation between deleted and normal mitochondrial DNA in some patients with mitochondrial myopathy

Twenty-three patients with mitochondrial myopathies and mitochondrial DNA deletions in muscle were studied by means of deletion mapping and sequencing, histochemistry and polarography. Histochemistry showed significantly less focal cytochrome oxidase deficiency relative to number of ragged red fibres when the deletion did not involve reading frames for cytochrome oxidase subunits. Polarography in such patients showed defects exclusively involving complex I, in contrast to the others with larger deletions who generally had more diffuse respiratory chain defects. Analysis of other published histochemical data showed similar findings to our own. It is concluded that translation of a proportion of deleted mitochondrial DNAs occurs in at least some patients with mitochondrial DNA deletions, implying that deleted and normal mitochondrial genomes share transfer RNAs within mitochondria in such cases.

Mitochondrial DNA in Postmortem Brain from Patients with Parkinson's Disease

Journal of NeurochemistryVolume 56, Issue 5 p. 1819-1819 Free Access Mitochondrial DNA in Postmortem Brain from Patients with Parkinson's Disease P. Lestienne, P. Lestienne U 298 INSERM CHRU Angers Angers, FranceSearch for more papers by this authorI. Nelson, I. Nelson U 298 INSERM CHRU Angers Angers, FranceSearch for more papers by this authorP. Riederer, P. Riederer University of Würzburg Würzburg, F.R.G.Search for more papers by this authorH. Reichmann, H. Reichmann University of Würzburg Würzburg, F.R.G.Search for more papers by this authorK. Jellinger, K. Jellinger LBI of Clinical Neurology Lainz Hospital Vienna, AustriaSearch for more papers by this author P. Lestienne, P. Lestienne U 298 INSERM CHRU Angers Angers, FranceSearch for more papers by this authorI. Nelson, I. Nelson U 298 INSERM CHRU Angers Angers, FranceSearch for more papers by this authorP. Riederer, P. Riederer University of Würzburg Würzburg, F.R.G.Search for more papers by this authorH. Reichmann, H. Reichmann University of Würzburg Würzburg, F.R.G.Search for more papers by this authorK. Jellinger, K. Jellinger LBI of Clinical Neurology Lainz Hospital Vienna, AustriaSearch for more papers by this author First published: May 1991 https://doi.org/10.1111/j.1471-4159.1991.tb02087.xCitations: 38AboutPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL No abstract is available for this article. REFERENCES Ikebe S., Tanaka M., Ohno K., Sato W., Hanori K., Kundo T., Mizuno Y., and Ozawa T. (1990) Increase of deleted mitochondrial DNA in the striatum in Parkinson's disease and senescence. Biochem. Biophys. Res. Commun. 170, 1044– 1048. Lestienne P. and Ponsot G. (1988) Kearns-Sayre syndrome with muscle mitochondrial DNA deletions. Lancet i, 885. Lestienne P., Nelson I., Riederer P., Jellinger K., and Reichmann H. (1990) Normal mitochondrial genome in brain from patients with Parkinson's disease and complex I defect. J. Neurochem. 55, 1810– 1812. Saiki R. K., Scharf S., Faloona F., Mullis K. B., Horn G. T., Erlich H. A., and Arheim N. (1985) Enzymatic amplification of β-globin genomic sequences and restriction site analysis for diagnosis of cell anaemia. Science 230, 1350– 1354. Schon E., Rizzuto R., Moraes C. T., Nakase H., Zeviani M., and DiMauro S. (1989) A direct repeat is a hot spot for large-scale deletion of human mitochondrial DNA. Science 244, 346– 349. Citing Literature Volume56, Issue5May 1991Pages 1819-1819 ReferencesRelatedInformation

Normal Mitochondrial Genome in Brain from Patients with Parkinson's Disease and Complex I Defect

The mitochondrial genome codes for 13 proteins which are located in the respiratory chain. In postmortem brain of patients with Parkinson's disease, decreased activity of complex I of the respiratory chain could be demonstrated. Because seven subunits of complex I are coded by the mitochondrial genome, we analyzed the mitochondrial DNA of human postmortem substantia nigra, putamen, and frontal cortex by the Southern blot technique. No deletions of the mitochondrial genome could be demonstrated, thus indicating that either subunits which are encoded by the nuclear genome are decreased or enzyme activity is diminished by metabolites, toxins, or increase of Fe3+.

A direct repeat is a hotspot for large-scale deletion of human mitochondrial DNA.

Kearns-Sayre syndrome (KSS) and progressive external ophthalmoplegia (PEO) are related neuromuscular disorders characterized by ocular myopathy and ophthalmoplegia. Almost all patients with KSS and about half with PEO harbor large deletions in their mitochondrial genomes. The deletions differ in both size and location, except for one, 5 kilobases long, that is found in more than one-third of all patients examined. This common deletion was found to be flanked by a perfect 13-base pair direct repeat in the normal mitochondrial genome. This result suggests that homologous recombination deleting large regions of intervening mitochondrial DNA, which previously had been observed only in lower eukaryotes and plants, operates in mammalian mitochondrial genomes as well, and is at least one cause of the deletions found in these two related mitochondrial myopathies.

Cloning of mtDNA fragments homologous to mitochondrial S2 plasmid-like DNA in maize

Mitochondria from S-type cytoplasmic male-sterile maize contain two small DNA species, S1 and S2, which are absent from other fertile and male-sterile cytoplasms. These species have been cloned in plasmid pBR322 by the homopolymer extension method. Probes made with these recombinant plasmids have been used to establish the homology between high molecular weight mitochondrial DNAs of fertile and male-sterile cytoplasms, and small mitochondrial plasmid-like molecules. Hybridization and mapping data show that S2 DNA copies are homologuous with sequences of the normal mitochondrial genome. A comparison of physical maps of different isolated mtDNA fragments indicates a heterogeneous arrangement of S2 sequences in the mtDNA population of normal fertile maize cytoplasm. The origin of this heterogeneity is discussed.