scholarly journals 2-Deoxy-D-glucose couples mitochondrial DNA replication with mitochondrial fitness and promotes the selection of wild-type over mutant mitochondrial DNA

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Boris Pantic ◽  
Daniel Ives ◽  
Mara Mennuni ◽  
Diego Perez-Rodriguez ◽  
Uxoa Fernandez-Pelayo ◽  
...  
Keyword(s):  
Mitochondrial Dna ◽  
Glucose Utilization ◽  
Wild Type ◽  
Human Mitochondrial Dna ◽  
Disease States ◽  
Glutamine Consumption ◽  
Mtdna Replication ◽  
Mitochondrial Dna Replication ◽  
Mutant Cells ◽  
Selection Of

AbstractPathological variants of human mitochondrial DNA (mtDNA) typically co-exist with wild-type molecules, but the factors driving the selection of each are not understood. Because mitochondrial fitness does not favour the propagation of functional mtDNAs in disease states, we sought to create conditions where it would be advantageous. Glucose and glutamine consumption are increased in mtDNA dysfunction, and so we targeted the use of both in cells carrying the pathogenic m.3243A>G variant with 2-Deoxy-D-glucose (2DG), or the related 5-thioglucose. Here, we show that both compounds selected wild-type over mutant mtDNA, restoring mtDNA expression and respiration. Mechanistically, 2DG selectively inhibits the replication of mutant mtDNA; and glutamine is the key target metabolite, as its withdrawal, too, suppresses mtDNA synthesis in mutant cells. Additionally, by restricting glucose utilization, 2DG supports functional mtDNAs, as glucose-fuelled respiration is critical for mtDNA replication in control cells, when glucose and glutamine are scarce. Hence, we demonstrate that mitochondrial fitness dictates metabolite preference for mtDNA replication; consequently, interventions that restrict metabolite availability can suppress pathological mtDNAs, by coupling mitochondrial fitness and replication.

scholarly journals Seleksi Primer LCO – HCO, Primer bird-f1 – HCO Dan Primer bch – bcl Untuk Amplifikasi Gen COI DNA Mitokondria Itik Magelang (Anas javanica)

2014 ◽  
Vol 16 (2) ◽  
pp. 83
Author(s):  
Sonny Abdi Setiyawan ◽  
Anto Budiharjo ◽  
Hermin Pancasakti Kusumaningrum
Keyword(s):  
Mitochondrial Dna ◽  
Key Words ◽  
Coi Gene ◽  
Wild Type ◽  
High Production ◽  
Partial Homology ◽  
Selection Of

Magelang duck is a wild type of local duck from Indonesia. The advantagesof Magelangduckcompare tootherlocalduck from Indonesiaareabilityto livein the highlandsandlowlands and high production of egg and meat. Geneticcharacterization of Magelangduck still not available until now.The aim of the research is selectprimers forampliflying COIgeneof mitochondrialDNAof MagelangduckusingLCO-HCO, bird-f1 -HCO, andbcl-bch primers.The research methodwas DNAisolationfrom Magelangduck. Followed by, selection of primer in silicoto find homologywithin COIsequenceusing ClustalX, Genedoc, and FastPCR programs. Amplification of COIgenewas performedusing PCRwith all primerpairs. Result showed partial homology with all primer in COI sequence. TheamplificationusingtheLCO-HCO primer produced  primerdimer.Primerbirdf1-HCOand bch-bcl primers showed no amplification.   Key words: Magelang duck, COI gene, mitochondrial DNA, primer

scholarly journals Selection of Drug-Tolerant Strains of Pythium sylvaticum Using Sublethal Enrichment

1997 ◽  
Vol 87 (7) ◽  
pp. 685-692 ◽  
Author(s):  
Frank N. Martin ◽  
Charles R. Semer
Keyword(s):  
Mitochondrial Dna ◽  
Growth Rates ◽  
Wild Type ◽  
Maternal Parent ◽  
Pythium Sylvaticum ◽  
Paternal Parent ◽  
High Level ◽  
Selection Of

Sublethal enrichment was used to generate mutants of Pythium sylvaticum tolerant to kanamycin and tetracycline. Kanamycin tolerance was readily generated, and mutants had growth rates similar to wild-type isolates at antibiotic concentrations lethal to wild-type isolates. Based on crosses between wild-type and mutant isolates, two types of inheritance of tolerance were identified. A high level of kanamycin tolerance was inherited in progeny only when the maternal parent was drug tolerant and was correlated with the inheritance of maternal mitochondrial DNA. A lower level of tolerance was observed in some progeny when the paternal parent was tolerant to the antibiotic and, based on the lack of inheritance of paternal mitochondrial DNA, was presumably nuclear-encoded. Selection of mutants tolerant to tetracycline took longer to generate than kanamycin-tolerant mutants. Based on crosses between tolerant and wild-type parents, tolerance to tetracycline was nuclear-encoded. Tolerance to both antibiotics was stable, with cultures retaining tolerance following repeated transfers on nonamended medium and after storage for 7 years.

scholarly journals MITOL-dependent ubiquitylation negatively regulates the entry of PolγA into mitochondria

PLoS Biology ◽  
2021 ◽  
Vol 19 (3) ◽  
pp. e3001139
Author(s):  
Mansoor Hussain ◽  
Aftab Mohammed ◽  
Shabnam Saifi ◽  
Aamir Khan ◽  
Ekjot Kaur ◽  
...  
Keyword(s):  
Mitochondrial Dna ◽  
Outer Membrane ◽  
E3 Ligase ◽  
Insoluble Fraction ◽  
Progressive External Ophthalmoplegia ◽  
Brdu Incorporation ◽  
Mitochondrial Outer Membrane ◽  
Wild Type ◽  
Mtdna Replication ◽  
Enhanced Degradation

Mutations in mitochondrial replicative polymerase PolγA lead to progressive external ophthalmoplegia (PEO). While PolγA is the known central player in mitochondrial DNA (mtDNA) replication, it is unknown whether a regulatory process exists on the mitochondrial outer membrane which controlled its entry into the mitochondria. We now demonstrate that PolγA is ubiquitylated by mitochondrial E3 ligase, MITOL (or MARCH5, RNF153). Ubiquitylation in wild-type (WT) PolγA occurs at Lysine 1060 residue via K6 linkage. Ubiquitylation of PolγA negatively regulates its binding to Tom20 and thereby its mitochondrial entry. While screening different PEO patients for mitochondrial entry, we found that a subset of the PolγA mutants is hyperubiquitylated by MITOL and interact less with Tom20. These PolγA variants cannot enter into mitochondria, instead becomes enriched in the insoluble fraction and undergo enhanced degradation. Hence, mtDNA replication, as observed via BrdU incorporation into the mtDNA, was compromised in these PEO mutants. However, by manipulating their ubiquitylation status by 2 independent techniques, these PEO mutants were reactivated, which allowed the incorporation of BrdU into mtDNA. Thus, regulated entry of non-ubiquitylated PolγA may have beneficial consequences for certain PEO patients.

scholarly journals Design and Use of a Peptide Nucleic Acid for Detection of the Heteroplasmic Low-Frequency Mitochondrial Encephalomyopathy, Lactic Acidosis, and Stroke-like Episodes (MELAS) Mutation in Human Mitochondrial DNA

2002 ◽  
Vol 48 (12) ◽  
pp. 2155-2163 ◽  
Author(s):  
Diane K Hancock ◽  
Frederick P Schwarz ◽  
Fenhong Song ◽  
Lee-Jun C Wong ◽  
Barbara C Levin
Keyword(s):  
Mitochondrial Dna ◽  
Lactic Acidosis ◽  
Low Frequency ◽  
Pcr Amplification ◽  
Mitochondrial Encephalomyopathy ◽  
Nucleotide Position ◽  
Wild Type ◽  
Human Mitochondrial Dna ◽  
Mtdna Mutations ◽  
A3243g Mutation

Abstract Background: Most pathogenic human mitochondrial DNA (mtDNA) mutations are heteroplasmic (i.e., mutant and wild-type mtDNA coexist in the same individual) and are difficult to detect when their concentration is a small proportion of that of wild-type mtDNA molecules. We describe a simple methodology to detect low proportions of the single base pair heteroplasmic mutation, A3243G, that has been associated with the disease mitochondrial encephalomyopathy, lactic acidosis, and stroke-like episodes (MELAS) in total DNA extracted from blood. Methods: Three peptide nucleic acids (PNAs) were designed to bind to the wild-type mtDNA in the region of nucleotide position 3243, thus blocking PCR amplification of the wild-type mtDNA while permitting the mutant DNA to become the dominant product and readily discernable. DNA was obtained from both apparently healthy and MELAS individuals. Optimum PCR temperatures were based on the measured ultraviolet thermal stability of the DNA/PNA duplexes. The presence or absence of the mutation was determined by sequencing. Results: In the absence of PNAs, the heteroplasmic mutation was either difficult to detect or undetectable by PCR and sequencing. Only PNA 3 successfully inhibited amplification of the wild-type mtDNA while allowing the mutant mtDNA to amplify. In the presence of PNA 3, we were able to detect the heteroplasmic mutation when its concentration was as low as 0.1% of the concentration of the wild-type sequence. Conclusion: This methodology permits easy detection of low concentrations of the MELAS A3243G mutation in blood by standard PCR and sequencing methods.

Mitochondrial DNA replication: clinical syndromes

2018 ◽  
Vol 62 (3) ◽  
pp. 297-308 ◽  
Author(s):  
Mohammed Almannai ◽  
Ayman W. El-Hattab ◽  
Fernando Scaglia
Keyword(s):  
Mitochondrial Dna ◽  
Continuous Process ◽  
Nuclear Genes ◽  
Mitochondrial Proteins ◽  
Adult Onset ◽  
Mild Disease ◽  
Mtdna Deletions ◽  
Clinical Syndromes ◽  
Mtdna Replication ◽  
Mitochondrial Dna Replication

Each nucleated cell contains several hundreds of mitochondria, which are unique organelles in being under dual genome control. The mitochondria contain their own DNA, the mtDNA, but most of mitochondrial proteins are encoded by nuclear genes, including all the proteins required for replication, transcription, and repair of mtDNA. MtDNA replication is a continuous process that requires coordinated action of several enzymes that are part of the mtDNA replisome. It also requires constant supply of deoxyribonucleotide triphosphates(dNTPs) and interaction with other mitochondria for mixing and unifying the mitochondrial compartment. MtDNA maintenance defects are a growing list of disorders caused by defects in nuclear genes involved in different aspects of mtDNA replication. As a result of defects in these genes, mtDNA depletion and/or multiple mtDNA deletions develop in affected tissues resulting in variable manifestations that range from adult-onset mild disease to lethal presentation early in life.

scholarly journals A test of the transcription model for biased inheritance of yeast mitochondrial DNA.

1995 ◽  
Vol 15 (9) ◽  
pp. 4803-4809 ◽  
Author(s):  
H E Lorimer ◽  
B J Brewer ◽  
W L Fangman
Keyword(s):  
Mitochondrial Dna ◽  
Nuclear Gene ◽  
Replication Initiation ◽  
High Density ◽  
Mitochondrial Rna ◽  
Wild Type ◽  
Mitochondrial Rna Polymerase ◽  
Origin Region ◽  
Mtdna Replication ◽  
Yeast Mitochondrial Dna

Two strand-specific origins of replication appear to be required for mammalian mitochondrial DNA (mtDNA) replication. Structural equivalents of these origins are found in the rep sequences of Saccharomyces cerevisiae mtDNA. These striking similarities have contributed to a universal model for the initiation of mtDNA replication in which a primer is created by cleavage of an origin region transcript. Consistent with this model are the properties of deletion mutants of yeast mtDNA ([rho-]) with a high density of reps (HS [rho-]). These mutant mtDNAs are preferentially inherited by the progeny resulting from the mating of HS [rho-] cells with cells containing wild-type mtDNA ([rho+]). This bias is presumed to result from a replication advantage conferred on HS [rho-] mtDNA by the high density of rep sequences acting as origins. To test whether transcription is indeed required for the preferential inheritance of HS [rho-] mtDNA, we deleted the nuclear gene (RPO41) for the mitochondrial RNA polymerase, reducing transcripts by at least 1000-fold. Since [rho-] genomes, but not [rho+] genomes, are stable when RPO41 is deleted, we examined matings between HS [rho-] and neutral [rho-] cells. Neutral [rho-] mtDNAs lack rep sequences and are not preferentially inherited in [rho-] x [rho+] crosses. In HS [rho-] x neutral [rho-] matings, the HS [rho-] mtDNA was preferentially inherited whether both parents were wild type or both were deleted for RPO41. Thus, transcription from the rep promoter does not appear to be necessary for biased inheritance. Our results, and analysis of the literature, suggest that priming by transcription is not a universal mechanism for mtDNA replication initiation.

92 Disease mutations in the human mitochondrial DNA polymerase thumb subdomain impart severe defects in mtDNA replication

Mitochondrion ◽  
2010 ◽  
Vol 10 (2) ◽  
pp. 225-226
Author(s):  
Rajesh Kasiviswanathan ◽  
Matthew J. Longley ◽  
Sherine S.L. Chan ◽  
William C. Copeland
Keyword(s):  
Mitochondrial Dna ◽  
Dna Polymerase ◽  
Human Mitochondrial Dna ◽  
Disease Mutations ◽  
Mtdna Replication ◽  
Mitochondrial Dna Polymerase
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