bims-midmar 2022-03-13 papers

bims-midmar

Biomed News

on Mitochondrial DNA maintenance and replication

Issue of 2022‒03‒13
nine papers selected by
Flavia Söllner
Ludwig-Maximilians University

Ultra-deep whole genome bisulfite sequencing reveals a single methylation hotspot in human brain mitochondrial DNA.
The Complicated Nature of Somatic mtDNA Mutations in Aging.
Mito-FUNCAT-FACS reveals cellular heterogeneity in mitochondrial translation.
ANALYSIS OF MITOCHONDRIAL DNA CYTOCHROME-B (CYB) and ATPase-6 GENE MUTATIONS IN COVID-19 PATIENTS.
Protein methylation in mitochondria.
Sperm mitochondrial DNA copy number in relation to semen quality: A cross-sectional study of 1164 potential sperm donors.
Advanced glycation end products, leukocyte telomere length, and mitochondrial DNA copy number in patients with coronary artery disease and alterations of glucose homeostasis: From the GENOCOR study.
Positive selection over the mitochondrial genome and its role in the diversification of gentoo penguins in response to adaptation in isolation.
mt tRFs, New Players in MELAS Disease.

Epigenetics. 2022 Mar 07. 1-16

Ultra-deep whole genome bisulfite sequencing reveals a single methylation hotspot in human brain mitochondrial DNA.

Romain Guitton, Christian Dölle, Guido Alves, Tysnes Ole-Bjørn, Gonzalo S Nido, Charalampos Tzoulis.

  While DNA methylation is established as a major regulator of gene expression in the nucleus, the existence of mitochondrial DNA (mtDNA) methylation remains controversial. Here, we characterized the mtDNA methylation landscape in the prefrontal cortex of neurological healthy individuals (n=26) and patients with Parkinson's disease (n=27), using a combination of whole-genome bisulphite sequencing (WGBS) and bisulphite-independent methods. Accurate mtDNA mapping from WGBS data required alignment to an mtDNA reference only, to avoid misalignment to nuclear mitochondrial pseudogenes. Once correctly aligned, WGBS data provided ultra-deep mtDNA coverage (16,723 ± 7,711) and revealed overall very low levels of cytosine methylation. The highest methylation levels (5.49 ± 0.97%) were found on CpG position m.545, located in the heavy-strand promoter 1 region. The m.545 methylation was validated using a combination of methylation-sensitive DNA digestion and quantitative PCR analysis. We detected no association between mtDNA methylation profile and Parkinson's disease. Interestingly, m.545 methylation correlated with the levels of mtDNA transcripts, suggesting a putative role in regulating mtDNA gene expression. In addition, we propose a robust framework for methylation analysis of mtDNA from WGBS data, which is less prone to false-positive findings due to misalignment of nuclear mitochondrial pseudogene sequences.

Keywords:  NUMTs; epigenetics; mitochondria; mtDNA; parkinson’s disease

DOI:  https://doi.org/10.1080/15592294.2022.2045754
Front Aging. 2022 ;pii: 805126. [Epub ahead of print]2

The Complicated Nature of Somatic mtDNA Mutations in Aging.

Monica Sanchez-Contreras, Scott R Kennedy.

  Mitochondria are the main source of energy used to maintain cellular homeostasis. This aspect of mitochondrial biology underlies their putative role in age-associated tissue dysfunction. Proper functioning of the electron transport chain (ETC), which is partially encoded by the extra-nuclear mitochondrial genome (mtDNA), is key to maintaining this energy production. The acquisition of de novo somatic mutations that interrupt the function of the ETC have long been associated with aging and common diseases of the elderly. Yet, despite over 30 years of study, the exact role(s) mtDNA mutations play in driving aging and its associated pathologies remains under considerable debate. Furthermore, even fundamental aspects of age-related mtDNA mutagenesis, such as when mutations arise during aging, where and how often they occur across tissues, and the specific mechanisms that give rise to them, remain poorly understood. In this review, we address the current understanding of the somatic mtDNA mutations, with an emphasis of when, where, and how these mutations arise during aging. Additionally, we highlight current limitations in our knowledge and critically evaluate the controversies stemming from these limitations. Lastly, we highlight new and emerging technologies that offer potential ways forward in increasing our understanding of somatic mtDNA mutagenesis in the aging process.

Keywords:  aging; mitochondria; mtDNA; mutagenesis; sequencing; somatic mutations

DOI:  https://doi.org/10.3389/fragi.2021.805126
RNA. 2022 Mar 07. pii: rna.079097.122. [Epub ahead of print]

Mito-FUNCAT-FACS reveals cellular heterogeneity in mitochondrial translation.

Yusuke Kimura, Hironori Saito, Tatsuya Osaki, Yasuhiro Ikegami, Taisei Wakigawa, Yoshiho Ikeuchi, Shintaro Iwasaki.

  Mitochondria possess their own genome that encodes components of oxidative phosphorylation (OXPHOS) complexes, and mitochondrial ribosomes within the organelle translate the mRNAs expressed from the mitochondrial genome. Given the differential OXPHOS activity observed in diverse cell types, cell growth conditions, and other circumstances, cellular heterogeneity in mitochondrial translation can be expected. Although individual protein products translated in mitochondria have been monitored, the lack of techniques that address the variation in overall mitochondrial protein synthesis in cell populations poses analytic challenges. Here, we adapted mitochondrial-specific fluorescent noncanonical amino acid tagging (FUNCAT) for use with fluorescence-activated cell sorting (FACS) and developed mito-FUNCAT-FACS. The click chemistry-compatible methionine analog L-homopropargylglycine (HPG) enabled the metabolic labeling of newly synthesized proteins. In the presence of cytosolic translation inhibitors, HPG was selectively incorporated into mitochondrial nascent proteins and conjugated to fluorophores via the click reaction (mito-FUNCAT). The application of in situ mito-FUNCAT to flow cytometry allowed us to separate changes in net mitochondrial translation activity from those of the organelle mass and detect variations in mitochondrial translation in cancer cells. Our approach provides a useful methodology for examining mitochondrial protein synthesis in individual cells.

Keywords:  FACS; FUNCAT; HPG; Mitochondria; Translation

DOI:  https://doi.org/10.1261/rna.079097.122
J Med Virol. 2022 Mar 08.

ANALYSIS OF MITOCHONDRIAL DNA CYTOCHROME-B (CYB) and ATPase-6 GENE MUTATIONS IN COVID-19 PATIENTS.

Ebubekir Dirican, Şeyda Tuba Savrun, İsmail Erkan Aydın, Gonca Gülbay, Ülkü Karaman.

  BACKGROUND: Coronavirus disease of 2019 (COVID-19) is a pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Mutations of mitochondrial DNA (mtDNA) are becoming increasingly common in various diseases. This study aims to investigate mutations in the cytochrome-b (CYB) and adenosine triphosphatase-6 (ATPase-6) genes of mtDNA in COVID-19 patients. The association between mtDNA mutations and clinical outcomes is investigated.METHODS: In the present study, mutations of the mtDNA genes CYB and ATPase-6 were investigated in COVID-19(+) (n=65) and COVID-19 (-) patients (n=65). First, we isolated DNA from the blood samples. After the PCR analyses, the mutations were defined using Sanger DNA sequencing.
RESULTS: The age, creatinine, ferritin and CRP levels of the COVID 19 (+) patients were higher than those of the COVID-19 (-) patients (p=0.0036, p=0.0383, p=0.0305, p<0.0001, respectively). We also found 16 different mutations in the CYB gene and 14 different mutations in the ATPase-6 gene. The incidences of CYB gene mutations A15326G, T15454C and C15452A were higher in COVID-19 (+) patients than COVID-19 (-) patients (p<0.0001 (OR (95% CI): 4.966 (2.215-10.89)), p=0.0226 and p=0.0226, respectively). In contrast, the incidences of A8860G and G9055A ATPase-6 gene mutations were higher in COVID-19 (+) patients than COVID-19 (-) patients (p<0.0001 (OR (95%CI): 5.333 (2.359-12.16) and p=0.0121 respectively). Yet, no significant relationship was found between mtDNA mutations and patients' age and biochemical parameters (p> 0.05).
CONCLUSIONS: The results showed that the frequency of mtDNA mutations in COVID-19 patients is quite high and it is important to investigate the association of these mutations with other genetic mechanisms in larger patient populations. This article is protected by copyright. All rights reserved.

Keywords:  ATPase-6; COVID-19; CYB; PCR; Sanger sequencing; mtDNA

DOI:  https://doi.org/10.1002/jmv.27704
J Biol Chem. 2022 Mar 02. pii: S0021-9258(22)00231-9. [Epub ahead of print] 101791

Protein methylation in mitochondria.

Jędrzej M Małecki, Erna Davydova, Pål Ø Falnes.

  Many proteins are modified by post-translational methylation, introduced by a number of methyltransferases (MTases). Protein methylation plays important roles in modulating protein function, and thus in optimizing and regulating cellular and physiological processes. Research has mainly focused on nuclear and cytosolic protein methylation, but it has been known for many years that also mitochondrial proteins are methylated. During the last decade, significant progress has been made on identifying the MTases responsible for mitochondrial protein methylation and addressing its functional significance. In particular, several novel human MTases have been uncovered that methylate lysine, arginine, histidine, and glutamine residues in various mitochondrial substrates. Several of these substrates are key components of the bioenergetics machinery, e.g. respiratory Complex I, citrate synthase and the ATP synthase. In the present review we report the status of the field of mitochondrial protein methylation, with a particular emphasis on recently discovered human MTases. We also discuss evolutionary aspects and functional significance of mitochondrial protein methylation, and present an outlook for this emergent research field.

Keywords:  ATP synthase; bioenergetics; electron transport chain; methyltransferase; mitochondria; oxidative phosphorylation; protein methylation

DOI:  https://doi.org/10.1016/j.jbc.2022.101791
BJOG. 2022 Mar 11.

Sperm mitochondrial DNA copy number in relation to semen quality: A cross-sectional study of 1164 potential sperm donors.

Bin Sun, Jian Hou, Yi-Xiang Ye, Heng-Gui Chen, Peng Duan, Ying-Jun Chen, Cheng-Liang Xiong, Yi-Xin Wang, An Pan.

  OBJECTIVE: To investigate the association between mitochondrial DNA copy number (mtDNAcn) and semen quality.DESIGN: A cross-sectional study.
SETTING: Hubei Province Human Sperm Bank of China (from April 2017 to July 2018).
POPULATION: A total of 1164 healthy male sperm donors with 5739 specimens.
MAIN OUTCOME MEASURES: Real-time quantitative polymerase chain reaction (RT-PCR) was used to measure sperm mtDNAcn. We also determined semen volume, concentration, and motility parameters (progressive motility, nonprogressive motility, and immotility).
METHODS: Mixed-effect models and general linear models were performed.
RESULTS: After adjusting for relevant confounding factors, mixed-effect models revealed diminished sperm motility (progressive and total), concentration, and total count across the quartiles of mtDNAcn (all p < 0.05). Compared to men in the lowest quartile, men in the highest quartile of mtDNAcn had lower progressive sperm motility, total motility, concentration, and total count of -8.9% (95% CI: -12.7%, -5.0%), -8.0% (95% CI: -11.6%, -4.4%), -42.8% (95% CI: -47.7%, -37.4%), and -44.3% (95% CI: -50.1%, -37.7%), respectively. These inverse dose-response relationships were further confirmed in the cubic spline models, where mtDNAcn was modeled as continuous variables.
CONCLUSIONS: We found that mtDNAcn was inversely associated with semen quality in a dose-dependent manner. Our results provide novel clues that sperm mtDNAcn may serve as a useful predictor of human semen characteristics.

Keywords:  Mitochondrial DNA copy number; Semen quality; mitochondrial biomarkers

DOI:  https://doi.org/10.1111/1471-0528.17139
Nutr Metab Cardiovasc Dis. 2022 Jan 24. pii: S0939-4753(22)00038-2. [Epub ahead of print]

Advanced glycation end products, leukocyte telomere length, and mitochondrial DNA copy number in patients with coronary artery disease and alterations of glucose homeostasis: From the GENOCOR study.

Cecilia Vecoli, Giuseppina Basta, Andrea Borghini, Melania Gaggini, Serena Del Turco, Antonella Mercuri, Amalia Gastaldelli, Maria Grazia Andreassi.

  BACKGROUND AND AIM: Alterations of glucose homeostasis can increase advanced glycation end products (AGEs) that exacerbate vascular inflammatory disease and may increase vascular senescence and aging. This study examined the relationships between carboxymethyl-lysine (CML) and soluble receptor for AGEs (sRAGE) with leukocyte telomere length (LTL) and mitochondrial DNA copy number (mtDNAcn), as cell aging biomarkers, in patients with established coronary artery disease (CAD).METHODS AND RESULTS: We studied 459 patients with CAD further categorized as having normal glucose homeostasis (NG, n = 253), pre-diabetes (preT2D, n = 85), or diabetes (T2D, n = 121). All patients were followed up for the occurrence of major adverse cardiovascular events (MACEs). Plasma concentrations of sRAGE and CML were measured by ELISA. mtDNAcn and LTL were measured by qRT-PCR. CML levels were significantly higher in patients with preT2D (p < 0.007) or T2D (p < 0.003) compared with those with NG. mtDNAcn resulted lower in T2D vs preT2D (p = 0.04). At multivariate Cox proportional hazard analysis, short LTL (HR: 2.89; 95% CI: 1.11-10.1; p = 0.04) and high levels of sRAGE (HR: 2.20; 95% CI: 1.01-5.14; p = 0.04) were associated with an increased risk for MACEs in patients with preT2D and T2D, respectively. T2D patients with both short LTL and high sRAGE levels had the highest risk of MACEs (HR: 3.11; 95% CI: 1.11-9.92; p = 0.04).
CONCLUSIONS: High levels of sRAGE and short LTL were associated with an increased risk of MACEs, especially in patients with diabetes, supporting the usefulness of both biomarkers of glycemic impairment and aging in predicting cardiovascular outcomes in patients with CAD.

Keywords:  Advanced glycation end products; Coronary artery disease; Diabetes; Glucose homeostasis; Leukocyte telomere length; Major adverse cardiac events; Mitochondrial DNA copy number; Soluble receptor for AGEs

DOI:  https://doi.org/10.1016/j.numecd.2022.01.021
Sci Rep. 2022 Mar 08. 12(1): 3767

Positive selection over the mitochondrial genome and its role in the diversification of gentoo penguins in response to adaptation in isolation.

D Noll, F Leon, D Brandt, P Pistorius, C Le Bohec, F Bonadonna, P N Trathan, A Barbosa, A Raya Rey, G P M Dantas, R C K Bowie, E Poulin, J A Vianna.

Although mitochondrial DNA has been widely used in phylogeography, evidence has emerged that factors such as climate, food availability, and environmental pressures that produce high levels of stress can exert a strong influence on mitochondrial genomes, to the point of promoting the persistence of certain genotypes in order to compensate for the metabolic requirements of the local environment. As recently discovered, the gentoo penguins (Pygoscelis papua) comprise four highly divergent lineages across their distribution spanning the Antarctic and sub-Antarctic regions. Gentoo penguins therefore represent a suitable animal model to study adaptive processes across divergent environments. Based on 62 mitogenomes that we obtained from nine locations spanning all four gentoo penguin lineages, we demonstrated lineage-specific nucleotide substitutions for various genes, but only lineage-specific amino acid replacements for the ND1 and ND5 protein-coding genes. Purifying selection (dN/dS < 1) is the main driving force in the protein-coding genes that shape the diversity of mitogenomes in gentoo penguins. Positive selection (dN/dS > 1) was mostly present in codons of the Complex I (NADH genes), supported by two different codon-based methods at the ND1 and ND4 in the most divergent lineages, the eastern gentoo penguin from Crozet and Marion Islands and the southern gentoo penguin from Antarctica respectively. Additionally, ND5 and ATP6 were under selection in the branches of the phylogeny involving all gentoo penguins except the eastern lineage. Our study suggests that local adaptation of gentoo penguins has emerged as a response to environmental variability promoting the fixation of mitochondrial haplotypes in a non-random manner. Mitogenome adaptation is thus likely to have been associated with gentoo penguin diversification across the Southern Ocean and to have promoted their survival in extreme environments such as Antarctica. Such selective processes on the mitochondrial genome may also be responsible for the discordance detected between nuclear- and mitochondrial-based phylogenies of gentoo penguin lineages.

DOI: https://doi.org/10.1038/s41598-022-07562-0
Front Physiol. 2022 ;13 800171

mt tRFs, New Players in MELAS Disease.

Salvador Meseguer, Mari-Paz Rubio.

  MELAS (mitochondrial encephalomyopathy, lactic acidosis, and stroke-like episodes) is an OXPHOS disease mostly caused by the m.3243A>G mutation in the mitochondrial tRNALeu(UUR) gene. Recently, we have shown that the mutation significantly changes the expression pattern of several mitochondrial tRNA-derived small RNAs (mt tsRNAs or mt tRFs) in a cybrid model of MELAS and in fibroblasts from MELAS patients versus control cells. Among them are those derived from mt tRNA LeuUUR containing or not the m.3243A>G mutation (mt 5'-tRF LeuUUR-m.3243A>G and mt 5'-tRF LeuUUR), whose expression levels are, respectively, increased and decreased in both MELAS cybrids and fibroblasts. Here, we asked whether mt 5'-tRF LeuUUR and mt 5'-tRF LeuUUR-m.3243A>G are biologically relevant and whether these mt tRFs are detected in diverse patient samples. Treatment with a mimic oligonucleotide of mt tRNA LeuUUR fragment (mt 5'-tRF LeuUUR) showed a therapeutic potential since it partially restored mitochondrial respiration in MELAS cybrids. Moreover, these mt tRFs could be detected in biofluids like urine and blood. We also investigated the participation of miRNA pathway components Dicer and Ago2 in the mt tRFs biogenesis process. We found that Dicer and Ago2 localize in the mitochondria of MELAS cybrids and that immunoprecipitation of these proteins in cytoplasm and mitochondria fractions revealed an increased mt tRF/mt tRNA ratio in MELAS condition compared to WT. These preliminary results suggest an involvement of Dicer and Ago2 in the mechanism of mt tRF biogenesis and action.

Keywords:  mitochondrial dysfunction; retrograde signaling; sncRNAs; tRF and tiRNA; tRNA fragment

DOI:  https://doi.org/10.3389/fphys.2022.800171