scholarly journals A Yeast-Based Repurposing Approach for the Treatment of Mitochondrial DNA Depletion Syndromes Led to the Identification of Molecules Able to Modulate the dNTP Pool

2021 ◽  
Vol 22 (22) ◽  
pp. 12223
Author(s):  
Giulia di Punzio ◽  
Micol Gilberti ◽  
Enrico Baruffini ◽  
Tiziana Lodi ◽  
Claudia Donnini ◽  
...  
Keyword(s):  
Mitochondrial Dna ◽  
Physiological Role ◽  
Homologous Gene ◽  
Inner Mitochondrial Membrane ◽  
Dntp Pool ◽  
Recessive Mutations ◽  
Mitochondrial Dna Depletion ◽  
Mtdna Deletion ◽  
Selective Channel ◽  
Deoxyribonucleoside Triphosphate

Mitochondrial DNA depletion syndromes (MDS) are clinically heterogenous and often severe diseases, characterized by a reduction of the number of copies of mitochondrial DNA (mtDNA) in affected tissues. In the context of MDS, yeast has proved to be both an excellent model for the study of the mechanisms underlying mitochondrial pathologies and for the discovery of new therapies via high-throughput assays. Among the several genes involved in MDS, it has been shown that recessive mutations in MPV17 cause a hepatocerebral form of MDS and Navajo neurohepatopathy. MPV17 encodes a non selective channel in the inner mitochondrial membrane, but its physiological role and the nature of its cargo remains elusive. In this study we identify ten drugs active against MPV17 disorder, modelled in yeast using the homologous gene SYM1. All ten of the identified molecules cause a concomitant increase of both the mitochondrial deoxyribonucleoside triphosphate (mtdNTP) pool and mtDNA stability, which suggests that the reduced availability of DNA synthesis precursors is the cause for the mtDNA deletion and depletion associated with Sym1 deficiency. We finally evaluated the effect of these molecules on mtDNA stability in two other MDS yeast models, extending the potential use of these drugs to a wider range of MDS patients.

scholarly journals Ribonucleotides incorporated by the yeast mitochondrial DNA polymerase are not repaired

2017 ◽  
Vol 114 (47) ◽  
pp. 12466-12471 ◽  
Author(s):  
Paulina H. Wanrooij ◽  
Martin K. M. Engqvist ◽  
Josefin M. E. Forslund ◽  
Clara Navarrete ◽  
Anna Karin Nilsson ◽  
...  
Keyword(s):  
Mitochondrial Dna ◽  
Nuclear Dna ◽  
Excision Repair ◽  
Genome Replication ◽  
Dntp Pool ◽  
Frequent Exchange ◽  
Repair Mechanisms ◽  
Ribonucleoside Triphosphate ◽  
Deoxyribonucleoside Triphosphate ◽  
Mitochondrial Dna Polymerase

Incorporation of ribonucleotides into DNA during genome replication is a significant source of genomic instability. The frequency of ribonucleotides in DNA is determined by deoxyribonucleoside triphosphate/ribonucleoside triphosphate (dNTP/rNTP) ratios, by the ability of DNA polymerases to discriminate against ribonucleotides, and by the capacity of repair mechanisms to remove incorporated ribonucleotides. To simultaneously compare how the nuclear and mitochondrial genomes incorporate and remove ribonucleotides, we challenged these processes by changing the balance of cellular dNTPs. Using a collection of yeast strains with altered dNTP pools, we discovered an inverse relationship between the concentration of individual dNTPs and the amount of the corresponding ribonucleotides incorporated in mitochondrial DNA, while in nuclear DNA the ribonucleotide pattern was only altered in the absence of ribonucleotide excision repair. Our analysis uncovers major differences in ribonucleotide repair between the two genomes and provides concrete evidence that yeast mitochondria lack mechanisms for removal of ribonucleotides incorporated by the mtDNA polymerase. Furthermore, as cytosolic dNTP pool imbalances were transmitted equally well into the nucleus and the mitochondria, our results support a view of the cytosolic and mitochondrial dNTP pools in frequent exchange.

The role of the Mpv17 protein mutations of which cause mitochondrial DNA depletion syndrome (MDDS): lessons from homologs in different species

2015 ◽  
Vol 396 (1) ◽  
pp. 13-25 ◽  
Author(s):  
Stefanie Löllgen ◽  
Hans Weiher
Keyword(s):  
Mitochondrial Dna ◽  
Tricarboxylic Acid Cycle ◽  
Clinical Manifestations ◽  
Nuclear Gene ◽  
Genotoxic Stress ◽  
Strand Break ◽  
Temperature Sensitive ◽  
Inner Mitochondrial Membrane ◽  
Knock Out ◽  
Mitochondrial Dna Depletion

Abstract Mitochondrial DNA depletion syndromes (MDDS) are severe pediatric diseases with diverse clinical manifestations. Gene mutations that underlie MDDS have been associated with alterations in the mitochondrial DNA (mtDNA) replication machinery or in mitochondrial deoxyribonucleoside triphosphate pools. However, the nuclear gene MPV17, whose mutated forms are associated with hepatocerebral MDDS in humans, plays a so-far unknown role in mtDNA maintenance. A high degree of conservation has been determined between MPV17 and its mouse (Mpv17), zebrafish (tra) and yeast (SYM1) homologs, respectively, whereby mutants in these cause very different phenotypes. While dysfunction in this gene in humans causes fatal liver disease, kidney pathology is induced in mice. Moreover, in zebrafish inactivation of the Mpv17 homolog was detected as a viable dyscolouration mutant. Knock out of the yeast ortholog results in a temperature-sensitive metabolic growth phenotype. Detailed analyses on common denominators between these different phenotypes strengthen the hypothesis that the Mpv17 protein forms a channel in the inner mitochondrial membrane, allowing small molecules – in vertebrates probably nucleotides, and in yeast probably intermediates of the tricarboxylic acid cycle – to pass. Moreover, a function modifying the pathologic manifestations of MPV17-related disease in mice has been identified. This signaling pathway remarkably involves the non-mitochondrial catalytic subunit of DNA-dependent protein kinase (PRKDC), important in double-strand break repair resistance against reactive oxygen-induced genotoxic stress.

scholarly journals Novel Autosomal Recessivec10orf2Mutations Causing Infantile-Onset Spinocerebellar Ataxia

2012 ◽  
Vol 2012 ◽  
pp. 1-4 ◽  
Author(s):  
Jessica N. Hartley ◽  
Frances A. Booth ◽  
Marc R. Del Bigio ◽  
Aizeddin A. Mhanni
Keyword(s):  
Mitochondrial Dna ◽  
Spinocerebellar Ataxia ◽  
Severe Phenotype ◽  
Phenotype Correlation ◽  
Finnish Population ◽  
Recessive Mutations ◽  
Phenotypic Spectrum ◽  
Helicase Gene ◽  
Mitochondrial Dna Depletion ◽  
Genes Encoding

Recessive mutations in genes encoding mitochondrial DNA replication machinery lead to mitochondrial DNA depletion syndromes. This genetically and phenotypically heterogeneous group includes infantile onset spinocerebellar ataxia (OMIM# 271245) a neurodegenerative disease caused by mutations in the mtDNA helicase gene,c10orf2, with an increased frequency in the Finnish population due to a founder mutation. We describe a child of English descent who presented with a severe phenotype of IOSCA as a result of two-novel mutations in thec10orf2gene. This paper expands the phenotypic spectrum of IOSCA and adds further evidence for the presence of a genotype-phenotype correlation among patients with recessive mutations in this gene.

scholarly journals The Human Mitochondrial DNA Depletion Syndrome GeneMPV17Encodes a Non-selective Channel That Modulates Membrane Potential

2015 ◽  
Vol 290 (22) ◽  
pp. 13840-13861 ◽  
Author(s):  
Vasily D. Antonenkov ◽  
Antti Isomursu ◽  
Daniela Mennerich ◽  
Miia H. Vapola ◽  
Hans Weiher ◽  
...  
Keyword(s):  
Mitochondrial Dna ◽  
Membrane Potential ◽  
Human Mitochondrial Dna ◽  
Mitochondrial Dna Depletion ◽  
Selective Channel ◽  
Mitochondrial Dna Depletion Syndrome

MPV17 encodes an inner mitochondrial membrane protein and is mutated in infantile hepatic mitochondrial DNA depletion

2006 ◽  
Vol 38 (5) ◽  
pp. 570-575 ◽  
Author(s):  
Antonella Spinazzola ◽  
Carlo Viscomi ◽  
Erika Fernandez-Vizarra ◽  
Franco Carrara ◽  
Pio D'Adamo ◽  
...  
Keyword(s):  
Mitochondrial Dna ◽  
Membrane Protein ◽  
Mitochondrial Membrane ◽  
Inner Mitochondrial Membrane ◽  
Mitochondrial Dna Depletion ◽  
Mitochondrial Membrane Protein

Development of a mito-CRISPR system for generating mitochondrial DNA-deleted strain in Saccharomyces cerevisiae

2020 ◽  
Vol 85 (4) ◽  
pp. 895-901
Author(s):  
Takamitsu Amai ◽  
Tomoka Tsuji ◽  
Mitsuyoshi Ueda ◽  
Kouichi Kuroda
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Mitochondrial Dna ◽  
Ethidium Bromide ◽  
Nuclear Genome ◽  
Target Sequence ◽  
Guide Rna ◽  
Crispr System ◽  
Mitochondrial Target ◽  
Gene Treatment ◽  
Mtdna Deletion

ABSTRACT Mitochondrial dysfunction can occur in a variety of ways, most often due to the deletion or mutation of mitochondrial DNA (mtDNA). The easy generation of yeasts with mtDNA deletion is attractive for analyzing the functions of the mtDNA gene. Treatment of yeasts with ethidium bromide is a well-known method for generating ρ° cells with complete deletion of mtDNA from Saccharomyces cerevisiae. However, the mutagenic effects of ethidium bromide on the nuclear genome cannot be excluded. In this study, we developed a “mito-CRISPR system” that specifically generates ρ° cells of yeasts. This system enabled the specific cleavage of mtDNA by introducing Cas9 fused with the mitochondrial target sequence at the N-terminus and guide RNA into mitochondria, resulting in the specific generation of ρ° cells in yeasts. The mito-CRISPR system provides a concise technology for deleting mtDNA in yeasts.

scholarly journals Expression of the Saccharomyces cerevisiae Gene YME1 in the Petite-Negative Yeast Schizosaccharomyces pombe Converts It to Petite-Positive

Genetics ◽  
2000 ◽  
Vol 154 (1) ◽  
pp. 147-154 ◽  
Author(s):  
Douglas J Kominsky ◽  
Peter E Thorsness
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Mitochondrial Dna ◽  
Schizosaccharomyces Pombe ◽  
Atp Synthase ◽  
Kluyveromyces Lactis ◽  
Blot Hybridization ◽  
Inner Mitochondrial Membrane ◽  
Yeast Saccharomyces Cerevisiae ◽  
Mitochondrial Atp Synthase ◽  
Petite Negative Yeast

Abstract Organisms that can grow without mitochondrial DNA are referred to as “petite-positive” and those that are inviable in the absence of mitochondrial DNA are termed “petite-negative.” The petite-positive yeast Saccharomyces cerevisiae can be converted to a petite-negative yeast by inactivation of Yme1p, an ATP- and metal-dependent protease associated with the inner mitochondrial membrane. Suppression of this yme1 phenotype can occur by virtue of dominant mutations in the α- and γ-subunits of mitochondrial ATP synthase. These mutations are similar or identical to those occurring in the same subunits of the same enzyme that converts the petite-negative yeast Kluyveromyces lactis to petite-positive. Expression of YME1 in the petite-negative yeast Schizosaccharomyces pombe converts this yeast to petite-positive. No sequence closely related to YME1 was found by DNA-blot hybridization to S. pombe or K. lactis genomic DNA, and no antigenically related proteins were found in mitochondrial extracts of S. pombe probed with antisera directed against Yme1p. Mutations that block the formation of the F1 component of mitochondrial ATP synthase are also petite-negative. Thus, the F1 complex has an essential activity in cells lacking mitochondrial DNA and Yme1p can mediate that activity, even in heterologous systems.

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