scholarly journals Molecular Basis of the Pathogenic Mechanism Induced by the m.9191T>C Mutation in Mitochondrial ATP6 Gene

2020 ◽  
Vol 21 (14) ◽  
pp. 5083 ◽  
Author(s):  
Xin Su ◽  
Alain Dautant ◽  
François Godard ◽  
Marine Bouhier ◽  
Teresa Zoladek ◽  
...  
Keyword(s):  
Atp Synthase ◽  
Atp Synthesis ◽  
Pathogenic Mutation ◽  
Loss Of Function ◽  
Subunit A ◽  
Mtdna Mutations ◽  
Leucine Residue ◽  
Atp6 Gene ◽  
Functional Consequences ◽  
Α Helix

Probing the pathogenicity and functional consequences of mitochondrial DNA (mtDNA) mutations from patient’s cells and tissues is difficult due to genetic heteroplasmy (co-existence of wild type and mutated mtDNA in cells), occurrence of numerous mtDNA polymorphisms, and absence of methods for genetically transforming human mitochondria. Owing to its good fermenting capacity that enables survival to loss-of-function mtDNA mutations, its amenability to mitochondrial genome manipulation, and lack of heteroplasmy, Saccharomyces cerevisiae is an excellent model for studying and resolving the molecular bases of human diseases linked to mtDNA in a controlled genetic background. Using this model, we previously showed that a pathogenic mutation in mitochondrial ATP6 gene (m.9191T>C), that converts a highly conserved leucine residue into proline in human ATP synthase subunit a (aL222P), severely compromises the assembly of yeast ATP synthase and reduces by 90% the rate of mitochondrial ATP synthesis. Herein, we report the isolation of intragenic suppressors of this mutation. In light of recently described high resolution structures of ATP synthase, the results indicate that the m.9191T>C mutation disrupts a four α-helix bundle in subunit a and that the leucine residue it targets indirectly optimizes proton conduction through the membrane domain of ATP synthase.

scholarly journals Case Report: Identification of a Novel Variant (m.8909T>C) of Human Mitochondrial ATP6 Gene and Its Functional Consequences on Yeast ATP Synthase

Life ◽  
2020 ◽  
Vol 10 (9) ◽  
pp. 215
Author(s):  
Qiuju Ding ◽  
Róża Kucharczyk ◽  
Weiwei Zhao ◽  
Alain Dautant ◽  
Shutian Xu ◽  
...  
Keyword(s):  
Mitochondrial Dna ◽  
Atp Synthase ◽  
Atp Synthesis ◽  
Single Individual ◽  
Loss Of Function ◽  
Mtdna Mutations ◽  
Atp6 Gene ◽  
Novel Variant ◽  
Functional Consequences ◽  
Mtdna Variants

With the advent of next generation sequencing, the list of mitochondrial DNA (mtDNA) mutations identified in patients rapidly and continuously expands. They are frequently found in a limited number of cases, sometimes a single individual (as with the case herein reported) and in heterogeneous genetic backgrounds (heteroplasmy), which makes it difficult to conclude about their pathogenicity and functional consequences. As an organism amenable to mitochondrial DNA manipulation, able to survive by fermentation to loss-of-function mtDNA mutations, and where heteroplasmy is unstable, Saccharomyces cerevisiae is an excellent model for investigating novel human mtDNA variants, in isolation and in a controlled genetic context. We herein report the identification of a novel variant in mitochondrial ATP6 gene, m.8909T>C. It was found in combination with the well-known pathogenic m.3243A>G mutation in mt-tRNALeu. We show that an equivalent of the m.8909T>C mutation compromises yeast adenosine tri-phosphate (ATP) synthase assembly/stability and reduces the rate of mitochondrial ATP synthesis by 20–30% compared to wild type yeast. Other previously reported ATP6 mutations with a well-established pathogenicity (like m.8993T>C and m.9176T>C) were shown to have similar effects on yeast ATP synthase. It can be inferred that alone the m.8909T>C variant has the potential to compromise human health.

scholarly journals ATP synthesis without R210 of subunit a in the Escherichia coli ATP synthase

2008 ◽  
Vol 1777 (1) ◽  
pp. 32-38 ◽  
Author(s):  
Robert R. Ishmukhametov ◽  
J. Blake Pond ◽  
Asma Al-Huqail ◽  
Mikhail A. Galkin ◽  
Steven B. Vik
Keyword(s):  
Escherichia Coli ◽  
Atp Synthase ◽  
Atp Synthesis ◽  
Subunit A

Deregulating mitochondrial metabolite and ion transport has beneficial effects in yeast and human cellular models for NARP syndrome

2019 ◽  
Vol 28 (22) ◽  
pp. 3792-3804 ◽  
Author(s):  
Xin Su ◽  
Malgorzata Rak ◽  
Emmanuel Tetaud ◽  
François Godard ◽  
Elodie Sardin ◽  
...  
Keyword(s):  
Atp Synthase ◽  
Leigh Syndrome ◽  
Yeast Cells ◽  
Complex Iv ◽  
Beneficial Effects ◽  
Cellular Models ◽  
Mtdna Mutations ◽  
Atp6 Gene ◽  
Dependent Growth ◽  
Stress Pathway

AbstractThe m.8993T>G mutation of the mitochondrial MT-ATP6 gene has been associated with numerous cases of neuropathy, ataxia and retinitis pigmentosa and maternally inherited Leigh syndrome, which are diseases known to result from abnormalities affecting mitochondrial energy transduction. We previously reported that an equivalent point mutation severely compromised proton transport through the ATP synthase membrane domain (FO) in Saccharomyces cerevisiae and reduced the content of cytochrome c oxidase (Complex IV or COX) by 80%. Herein, we report that overexpression of the mitochondrial oxodicarboxylate carrier (Odc1p) considerably increases Complex IV abundance and tricarboxylic acid-mediated substrate-level phosphorylation of ADP coupled to conversion of α-ketoglutarate into succinate in m.8993T>G yeast. Consistently in m.8993T>G yeast cells, the retrograde signaling pathway was found to be strongly induced in order to preserve α-ketoglutarate production; when Odc1p was overexpressed, this stress pathway returned to an almost basal activity. Similar beneficial effects were induced by a partial uncoupling of the mitochondrial membrane with the proton ionophore, cyanide m-chlorophenyl hydrazone. This chemical considerably improved the glutamine-based, respiration-dependent growth of human cytoplasmic hybrid cells that are homoplasmic for the m.8993T>G mutation. These findings shed light on the interdependence between ATP synthase and Complex IV biogenesis, which could lay the groundwork for the creation of nutritional or metabolic interventions for attenuating the effects of mtDNA mutations.

scholarly journals Respiratory complex and tissue lineage drive mutational patterns in the tumor mitochondrial genome

2020 ◽  
Author(s):  
Alexander N. Gorelick ◽  
Minsoo Kim ◽  
Walid K. Chatila ◽  
Konnor La ◽  
A. Ari Hakimi ◽  
...  
Keyword(s):  
Mitochondrial Genome ◽  
Atp Synthesis ◽  
Loss Of Function ◽  
Sequencing Data ◽  
Mtdna Mutation ◽  
Driver Genes ◽  
Cancer Driver ◽  
Translational Machinery ◽  
Synonymous Mutations ◽  
Mtdna Mutations

AbstractMitochondrial DNA (mtDNA) encodes essential protein subunits and translational machinery for four distinct complexes of oxidative phosphorylation (OXPHOS). Using repurposed whole-exome sequencing data, we demonstrate that pathogenic mtDNA mutations arise in tumors at a rate comparable to the most common cancer driver genes. We identify OXPHOS complexes as critical determinants shaping somatic mtDNA mutation patterns across tumor lineages. Loss-of-function mutations accumulate at an elevated rate specifically in Complex I, and often arise at specific homopolymeric hotspots. In contrast, Complex V is depleted of all non-synonymous mutations, suggesting that mutations directly impacting ATP synthesis are under negative selection. Both common truncating mutations and rarer missense alleles are associated with a pan-lineage transcriptional program, even in cancer types where mtDNA mutations are comparatively rare. Pathogenic mutations of mtDNA are associated with substantial increases in overall survival of colorectal adenocarcinoma patients, demonstrating a clear functional relationship between genotype and phenotype. The mitochondrial genome is therefore frequently and functionally disrupted across many cancers, with significant implications for patient stratification, prognosis and therapeutic development.

scholarly journals Functional investigation of an universally conserved leucine residue in subunit a of ATP synthase targeted by the pathogenic m.9176 T>G mutation

2019 ◽  
Vol 1860 (1) ◽  
pp. 52-59 ◽  
Author(s):  
Roza Kucharczyk ◽  
Alain Dautant ◽  
François Godard ◽  
Déborah Tribouillard-Tanvier ◽  
Jean-Paul di Rago
Keyword(s):  
Atp Synthase ◽  
Subunit A ◽  
Leucine Residue ◽  
Functional Investigation

scholarly journals Rotating with the brakes on and other unresolved features of the vacuolar ATPase

2016 ◽  
Vol 44 (3) ◽  
pp. 851-855 ◽  
Author(s):  
Shaun Rawson ◽  
Michael A. Harrison ◽  
Stephen P. Muench
Keyword(s):  
Secondary Structure ◽  
Atp Synthase ◽  
Atp Synthesis ◽  
Vacuolar Atpase ◽  
Proton Gradient ◽  
Subunit A ◽  
Secondary Transport

The rotary ATPase family comprises the ATP synthase (F-ATPase), vacuolar ATPase (V-ATPase) and archaeal ATPase (A-ATPase). These either predominantly utilize a proton gradient for ATP synthesis or use ATP to produce a proton gradient, driving secondary transport and acidifying organelles. With advances in EM has come a significant increase in our understanding of the rotary ATPase family. Following the sub nm resolution reconstructions of both the F- and V-ATPases, the secondary structure organization of the elusive subunit a has now been resolved, revealing a novel helical arrangement. Despite these significant developments in our understanding of the rotary ATPases, there are still a number of unresolved questions about the mechanism, regulation and overall architecture, which this mini-review aims to highlight and discuss.

scholarly journals pH-dependent 11° F1FO ATP synthase sub-steps reveal insight into the FO torque generating mechanism

eLife ◽  
2021 ◽  
Vol 10 ◽  
Author(s):  
Seiga Yanagisawa ◽  
Wayne D Frasch
Keyword(s):  
Atp Synthase ◽  
Single Molecule ◽  
Atp Hydrolysis ◽  
Proton Translocation ◽  
Atp Synthesis ◽  
Proton Pumping ◽  
Subunit A ◽  
Pka Values ◽  
C Subunit ◽  
Ph Dependent

Most cellular ATP is made by rotary F1FO ATP synthases using proton translocation-generated clockwise torque on the FO c-ring rotor, while F1-ATP hydrolysis can force counterclockwise rotation and proton pumping. The FO torque-generating mechanism remains elusive even though the FO interface of stator subunit-a, which contains the transmembrane proton half-channels, and the c-ring is known from recent F1FO structures. Here, single-molecule F1FO rotation studies determined that the pKa values of the half-channels differ, show that mutations of residues in these channels change the pKa values of both half-channels, and reveal the ability of FO to undergo single c-subunit rotational stepping. These experiments provide evidence to support the hypothesis that proton translocation through FO operates via a Grotthuss mechanism involving a column of single water molecules in each half-channel linked by proton translocation-dependent c-ring rotation. We also observed pH-dependent 11° ATP synthase-direction sub-steps of the E. coli c10-ring of F1FO against the torque of F1-ATPase-dependent rotation that result from H+ transfer events from FO subunit-a groups with a low pKa to one c-subunit in the c-ring, and from an adjacent c-subunit to stator groups with a high pKa. These results support a mechanism in which alternating proton translocation-dependent 11° and 25° synthase-direction rotational sub-steps of the c10-ring occur to sustain F1FO ATP synthesis.

scholarly journals The pathogenic m.8993 T > G mutation in mitochondrial ATP6 gene prevents proton release from the subunit c-ring rotor of ATP synthase

2021 ◽  
Author(s):  
Xin Su ◽  
Alain Dautant ◽  
Malgorzata Rak ◽  
François Godard ◽  
Nahia Ezkurdia ◽  
...  
Keyword(s):  
Atp Synthase ◽  
Electrostatic Interactions ◽  
Neuromuscular Diseases ◽  
Atp Synthesis ◽  
Biochemical Properties ◽  
Gene Encoding ◽  
Subunit C ◽  
Subunit A ◽  
C Subunit ◽  
Respiratory Growth

Abstract The human ATP synthase is an assembly of 29 subunits of 18 different types, of which only two (a and 8) are encoded in the mitochondrial genome. Subunit a, together with an oligomeric ring of c-subunit (c-ring), forms the proton pathway responsible for the transport of protons through the mitochondrial inner membrane, coupled to rotation of the c-ring and ATP synthesis. Neuromuscular diseases have been associated to a number of mutations in the gene encoding subunit a, ATP6. The most common, m.8993 T > G, leads to replacement of a strictly conserved leucine residue with arginine (aL156R). We previously showed that the equivalent mutation (aL173R) dramatically compromises respiratory growth of Saccharomyces cerevisiae and causes a 90% drop in the rate of mitochondrial ATP synthesis. Here we isolated revertants from the aL173R strain that show improved respiratory growth. Four first-site reversions at codon 173 (aL173M, aL173S, aL173K, and aL173W) and five second-site reversions at another codon (aR169M, aR169S, aA170P, aA170G, and aI216S) were identified. Based on the atomic structures of yeast ATP synthase and the biochemical properties of the revertant strains, we propose that the aL173R mutation is responsible for unfavorable electrostatic interactions that prevent the release of protons from the c-ring into a channel from which protons move from the c-ring to the mitochondrial matrix. The results provide further evidence that yeast aL173 (and thus human aL156) optimizes the exit of protons from ATP synthase, but is not essential despite its strict evolutionnary conservation.

scholarly journals A Theoretical Model of Mitochondrial ATP Synthase Deficiencies. The Role of Mitochondrial Carriers

Processes ◽  
2021 ◽  
Vol 9 (8) ◽  
pp. 1424
Author(s):  
Jean-Pierre Mazat ◽  
Anne Devin ◽  
Edgar Yoboue ◽  
Stéphane Ransac
Keyword(s):  
Atp Synthase ◽  
Point Mutations ◽  
Atp Synthesis ◽  
Yeast Saccharomyces Cerevisiae ◽  
Complex Iv ◽  
Equivalent Point ◽  
Mitochondrial Atp Synthase ◽  
Atp6 Gene ◽  
Substrate Level

The m.8993T>G mutation of the mitochondrial MT-ATP6 gene is associated with NARP syndrome (neuropathy, ataxia and retinitis pigmentosa). The equivalent point mutation introduced in yeast Saccharomyces cerevisiae mitochondrial DNA considerably reduced the activity of ATP synthase and of cytochrome-c-oxidase, preventing yeast growth on oxidative substrates. The overexpression of the mitochondrial oxodicarboxylate carrier (Odc1p) was able to rescue the growth on the oxidative substrate by increasing the substrate-level phosphorylation of ADP coupled to the conversion of α-ketoglutarate (AKG) into succinate with an increase in Complex IV activity. Previous studies showed that equivalent point mutations in ATP synthase behave similarly and can be rescued by Odc1p overexpression and/or the uncoupling of OXPHOS from ATP synthesis. In order to better understand the mechanism of the ATP synthase mutation bypass, we developed a core model of mitochondrial metabolism based on AKG as a respiratory substrate. We describe the different possible metabolite outputs and the ATP/O ratio values as a function of ATP synthase inhibition.

scholarly journals Diminished synthesis of subunit a (ATP6) and altered function of ATP synthase and cytochrome c oxidase due to the mtDNA 2 bp microdeletion of TA at positions 9205 and 9206

2004 ◽  
Vol 383 (3) ◽  
pp. 561-571 ◽  
Author(s):  
Pavel JEŠINA ◽  
Markéta TESAŘOVÁ ◽  
Daniela FORNŮSKOVÁ ◽  
Alena VOJTÍŠKOVÁ ◽  
Petr PECINA ◽  
...  
Keyword(s):  
Cytochrome C ◽  
Cytochrome C Oxidase ◽  
Atp Synthase ◽  
Stop Codon ◽  
Atp Synthesis ◽  
Pcr Analysis ◽  
Mitochondrial Atpase ◽  
Atpase Subunit ◽  
Mitochondrial Encephalomyopathies ◽  
Subunit A

Dysfunction of mitochondrial ATPase (F1Fo-ATP synthase) due to missense mutations in ATP6 [mtDNA (mitochondrial DNA)-encoded subunit a] is a frequent cause of severe mitochondrial encephalomyopathies. We have investigated a rare mtDNA mutation, i.e. a 2 bp deletion of TA at positions 9205 and 9206 (9205ΔTA), which affects the STOP codon of the ATP6 gene and the cleavage site between the RNAs for ATP6 and COX3 (cytochrome c oxidase 3). The mutation was present at increasing load in a three-generation family (in blood: 16%/82%/>98%). In the affected boy with severe encephalopathy, a homoplasmic mutation was present in blood, fibroblasts and muscle. The fibroblasts from the patient showed normal aurovertin-sensitive ATPase hydrolytic activity, a 70% decrease in ATP synthesis and an 85% decrease in COX activity. ADP-stimulated respiration and the ADP-induced decrease in the mitochondrial membrane potential at state 4 were decreased by 50%. The content of subunit a was decreased 10-fold compared with other ATPase subunits, and [35S]-methionine labelling showed a 9-fold decrease in subunit a biosynthesis. The content of COX subunits 1, 4 and 6c was decreased by 30–60%. Northern Blot and quantitative real-time reverse transcription–PCR analysis further demonstrated that the primary ATP6 – COX3 transcript is cleaved to the ATP6 and COX3 mRNAs 2–3-fold less efficiently. Structural studies by Blue-Native and two-dimensional electrophoresis revealed an altered pattern of COX assembly and instability of the ATPase complex, which dissociated into subcomplexes. The results indicate that the 9205ΔTA mutation prevents the synthesis of ATPase subunit a, and causes the formation of incomplete ATPase complexes that are capable of ATP hydrolysis but not ATP synthesis. The mutation also affects the biogenesis of COX, which is present in a decreased amount in cells from affected individuals.

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