Mitochondrial DNA Dynamics Research Articles

Big jaw, small genome: first description of the mitochondrial genome of Odontomachus (Formicidae, Ponerinae): evolutionary implications for Ponerinae ants

Mitochondrial DNA is a valuable tool for population genetics and evolutionary studies in a wide range of organisms. With advancements in sequencing techniques, it’s now possible to gain deeper insights into this molecule. By understanding how many genes there are, how they’re organized within the molecule, identifying the presence of spacers, and analyzing the composition of the D-Loop, we can better grasp the rearrangements that play a crucial role in the evolutionary dynamics of mitochondrial DNA. Additionally, phylogenetic analyses benefit significantly from having access to a larger pool of mtDNA genes. This wealth of genetic information allows for the establishment of evolutionary relationships with greater accuracy than ever before, providing a more robust framework than analyses based on a limited number of genes. Studies on mitogenomes belonging to the family Formicidae have proven promising, enabling the identification of gene rearrangements and enhancing our understanding of the internal relationships within the group. Despite this, the number of mitogenomes available for the subfamily Ponerinae is still limited, and here we present for the first time the complete mitogenome of Odontomachus. Our data reveal a gene duplication event in Formicidae, the first involving trnV, and new gene arrangements involving the trnM–trnI–trnQ and trnW–trnC–trnY clusters, suggesting a possible synapomorphy for the genus. Our phylogenetic analysis using the PCGs available for Formicidae supports the monophyly of the subfamily Ponerinae and sheds light on the relationship between Odontomachus and Pachycondyla.

Unraveling the mitochondrial phylogenetic landscape of Thailand reveals complex admixture and demographic dynamics

The evolutionary dynamics of mitochondrial DNA within the Thai population were comprehensively explored with a specific focus on the influence of South Asian admixture. A total of 166 samples were collected through randomized sampling, ensuring a diverse representation. Our findings unveil substantial genetic and haplogroup diversity within the Thai population. We have identified 164 haplotypes categorized into 97 haplogroups, with a notable inclusion of 20 novel haplogroups. The distribution of haplogroups exhibited variations across different populations and countries. The central Thai population displayed a high diversity of haplogroups from both the M and N clades. Maternal lineage affinities were discerned between several Mainland Southeast Asia (MSEA) and South Asian populations, implying ancestral genetic connections and a substantial influence of South Asian women in establishing these relationships. f4-statistics indicates the presence of a Tibeto-Burman genetic component within the Mon population from Thailand. New findings demonstrate two phases of population expansion occurring 22,000–26,000 and 2500–3800 years ago, coinciding with the Last Glacial Maximum, and Neolithic demographic transition, respectively. This research significantly enhances our understanding of the maternal genetic history of Thailand and MSEA, emphasizing the influence of South Asian admixture. Moreover, it underscores the critical role of prior information, such as mutation rates, within the Bayesian framework for accurate estimation of coalescence times and inferring demographic history.

The effect of vitrification on blastocyst mitochondrial DNA dynamics and gene expression profiles.

Does vitrification/warming affect the mitochondrial DNA (mtDNA) content and the gene expression profile of blastocysts? Prospective cohort study in which 89 blastocysts were obtained from 50 patients between July 2017 and August 2018. mtDNA was measured in a total of 71 aneuploid blastocysts by means of real-time polymerase chain reaction (RT-PCR). Transcriptomic analysis was performed by RNA sequencing (RNA-seq) in an additional 8 aneuploid blastocysts cultured for 0h after warming, and 10 aneuploid blastocysts cultured for 4-5h after warming. A significant decrease in mtDNA content just during the first hour after the warming process in blastocysts was found (P < 0.05). However, mtDNA content experimented a significantly increased along the later culture hours achieving the original mtDNA levels before vitrification after 4-5h of culture (P < 0.05). Gene expression analysis and functional enrichment analysis revealed that such recovery was accompanied by upregulation of pathways associated with embryo developmental capacity and uterine embryo development. Interestingly, the significant increase in mtDNA content observed in blastocysts just after warming also coincided with the differential expression of several cellular stress response-related pathways, such as apoptosis, DNA damage, humoral immune responses, and cancer. To our knowledge, this is the first study demonstrating in humans, a modulation in blastocysts mtDNA content in response to vitrification and warming. These results will be useful in understanding which pathways and mechanisms may be activated in human blastocysts following vitrification and warming before a transfer.

Exposure to unmethylated CpG oligonucleotides disrupts blood pressure circadian rhythms and placental clock gene network in pregnant rats.

Bacterial infections and impaired circulating mitochondrial DNA dynamics are associated with adverse pregnancy outcomes. Unmethylated cytosine-guanine dinucleotide (CpG) motifs are common in bacterial and mitochondrial DNA and act as potent immunostimulators. We tested the hypothesis that exposure to CpG oligonucleotides (ODN) during pregnancy would disrupt blood pressure circadian rhythms and placental molecular clock network, mediating aberrant fetoplacental growth dynamics. Rats were repeatedly treated with CpG ODN in the third trimester [gestational days (GD) 14, 16, and 18] and euthanized on GD20 (near term) or treated with a single dose of CpG ODN on GD14 and euthanized 4 h after treatment. Hemodynamic circadian rhythms were analyzed via Lomb-Scargle periodogram analysis on 24-h raw data collected continuously via radiotelemetry. A P value ≥ 0.05 indicates the absence of a circadian rhythm. Following the first treatment with CpG ODN, maternal systolic and diastolic blood pressure circadian rhythms were lost (P ≥ 0.05). Blood pressure circadian rhythm was restored by GD16 and remained unaffected after the second treatment with CpG ODN (P < 0.0001). Diastolic blood pressure circadian rhythm was again lost after the last treatment on GD18 (P ≥ 0.05). CpG ODN increased placental expression of Per2, Per3, and Tnfα (P ≤ 0.05) and affected fetoplacental growth dynamics. Reduced fetal and placental weights were disproportionately associated with increases in the number of resorptions in ODN-treated dams compared with controls. In conclusion, gestational exposure to unmethylated CpG ODN dysregulates the placental molecular clock network and fetoplacental growth dynamics and disrupts blood pressure circadian rhythms.NEW & NOTEWORTHY Gestational exposure to unmethylated CpG ODN dysregulates placental molecular clock network and fetoplacental growth dynamics and disrupts blood pressure circadian rhythms. These findings provide novel insights into the relationship between circadian rhythms and immune responses in pregnancy and propose new mechanisms by which maternal responses to immune triggers could dictate circadian rhythms of cardiovascular processes and placental clock machinery function to determine fetal growth trajectories.

Visualize the Distribution and Dynamics of Mitochondrial DNA (mtDNA) Nucleoids with Multiple Labeling Strategies.

Mitochondrial DNA (mtDNA) encodes a variety of rRNAs, tRNAs, and respiratory chain complex proteins. The integrity of mtDNA supports the mitochondrial functions and plays an essential role in numerous physiological and pathological processes. Mutations in mtDNA cause metabolic diseases and aging. The mtDNA within the human cells are packaged into hundreds of nucleoids within the mitochondrial matrix. Knowledge of how the nucleoids are dynamically distributed and organized within mitochondria is key to understanding mtDNA structure and functions. Therefore, visualizing the distribution and dynamics of mtDNA within mitochondria is a powerful approach to gain insights into the regulation of mtDNA replication and transcription. In this chapter, we describe the methods of observing mtDNA and its replication with fluorescence microscopy in both fixed and live cells using different labeling strategies.

Mitochondrial DNA Dynamics in Reprogramming to Pluripotency.

Mammalian cells, with the exception of erythrocytes, harbor mitochondria, which are organelles that provide energy, intermediate metabolites, and additional activities to sustain cell viability, replication, and function. Mitochondria contain multiple copies of a circular genome called mitochondrial DNA (mtDNA), whose individual sequences are rarely identical (homoplasmy) because of inherited or sporadic mutations that result in multiple mtDNA genotypes (heteroplasmy). Here, we examine potential mechanisms for maintenance or shifts in heteroplasmy that occur in induced pluripotent stem cells (iPSCs) generated by cellular reprogramming, and further discuss manipulations that can alter heteroplasmy to impact stem and differentiated cell performance. This additional insight will assist in developing more robust iPSC-based models of disease and differentiated cell therapies.

MTRIP, an Imaging Tool to Investigate Mitochondrial DNA Dynamics in Physiology and Disease at the Single-Cell Resolution.

Mitochondrial physiology and metabolism are closely linked to replication and transcription of mitochondrial DNA (mtDNA). However, the characterization of mtDNA processing is poorly defined at the single-cell level. We developed mTRIP (mitochondrial Transcription and Replication Imaging Protocol), an imaging approach based on modified fluorescence in situ hybridization (FISH), which simultaneously reveals mitochondrial structures committed to mtDNA initiation of replication as well as the mitochondrial RNA (mtRNA) content at the single-cell level in human cells. Also specific RNA regions, rather than global RNA, can be tracked with mTRIP. In addition, mTRIP can be coupled to immunofluorescence for in situ protein tracking, or to MitoTracker, thereby allowing for simultaneous labeling of mtDNA, mtRNA, and proteins or mitochondria, respectively. Altogether, qualitative and quantitative alterations of the dynamics of mtDNA processing are detected by mTRIP in human cells undergoing physiological changes, as well as stress and dysfunction. mTRIP helped elucidating mtDNA processing alterations in cancer cells, and has a potential for diagnostic of mitochondrial diseases.

Mitochondrial DNA Parameters in Blood of Infants Receiving Lopinavir/Ritonavir or Lamivudine Prophylaxis to Prevent Breastfeeding Transmission of HIV-1.

Children who are human immunodeficiency virus (HIV)-exposed but uninfected (CHEU) accumulate maternal HIV and antiretroviral exposures through pregnancy, postnatal prophylaxis, and breastfeeding. Here, we compared the dynamics of mitochondrial DNA (mtDNA) parameters in African breastfed CHEU receiving lopinavir/ritonavir (LPV/r) or lamivudine (3TC) pre-exposure prophylaxis during the first year of life. The number of mtDNA copies per cell (MCN) and the proportion of deleted mtDNA (MDD) were assessed at day 7 and at week 50 post-delivery (PrEP group). mtDNA depletion was defined as a 50% or more decrease from the initial value, and mtDNA deletions was the detection of mtDNA molecules with large DNA fragment loss. We also performed a sub-analysis with CHEU who did not receive a prophylactic treatment in South Africa (control group). From day seven to week 50, MCN decreased with a median of 41.7% (interquartile range, IQR: 12.1; 64.4) in the PrEP group. The proportion of children with mtDNA depletion was not significantly different between the two prophylactic regimens. Poisson regressions showed that LPV/r and 3TC were associated with mtDNA depletion (reference: control group; LPV/r: PR = 1.75 (CI95%: 1.15–2.68), p < 0.01; 3TC: PR = 1.54 (CI95%: 1.00–2.37), p = 0.05). Moreover, the proportion of children with MDD was unexpectedly high before randomisation in both groups. Long-term health impacts of these mitochondrial DNA parameters should be investigated further for both CHEU and HIV-infected children receiving LPV/r- or 3TC- based regimens.

A nanoscale, multi-parametric flow cytometry-based platform to study mitochondrial heterogeneity and mitochondrial DNA dynamics

Mitochondria are well-characterized regarding their function in both energy production and regulation of cell death; however, the heterogeneity that exists within mitochondrial populations is poorly understood. Typically analyzed as pooled samples comprised of millions of individual mitochondria, there is little information regarding potentially different functionality across subpopulations of mitochondria. Herein we present a new methodology to analyze mitochondria as individual components of a complex and heterogeneous network, using a nanoscale and multi–parametric flow cytometry-based platform. We validate the platform using multiple downstream assays, including electron microscopy, ATP generation, quantitative mass-spectrometry proteomic profiling, and mtDNA analysis at the level of single organelles. These strategies allow robust analysis and isolation of mitochondrial subpopulations to more broadly elucidate the underlying complexities of mitochondria as these organelles function collectively within a cell.

Mitoepigenetics and Neurodegenerative Diseases.

Mitochondrial impairment and increased oxidative stress are common features in neurodegenerative disorders, leading researchers to speculate that epigenetic changes in the mitochondrial DNA (mitoepigenetics) could contribute to neurodegeneration. The few studies performed so far to address this issue revealed impaired methylation levels of the mitochondrial regulatory region (D-loop region) in both animal models, postmortem brain regions, or circulating blood cells of patients with Alzheimer's disease, Parkinson's disease, and amyotrophic lateral sclerosis. Those studies also revealed that mtDNA D-loop methylation levels are subjected to a dynamic regulation within the progression of the neurodegenerative process, could be affected by certain neurodegenerative disease-causative mutations, and are inversely correlated with the mtDNA copy number. The methylation levels of other mtDNA regions than the D-loop have been scarcely investigated in human specimens from patients with neurodegenerative disorders or in animal models of the disease, and evidence of impaired methylation levels is often limited to a single study, making it difficult to clarify their correlation with mitochondrial dynamics and gene expression levels in these disorders. Overall, the preliminary results of the studies performed so far are encouraging making mitoepigenetics a timely and attractive field of investigation, but additional research is warranted to clarify the connections among epigenetic changes occurring in the mitochondrial genome, mitochondrial DNA dynamics and gene expression, and the neurodegenerative process.

Mitochondrial DNA dynamics during fruiting body formation in Grifola frondosa

Abstract In the present study, the copy number of mitochondrial DNA in Grifola frondosa was estimated at several developmental stages by quantitative real-time PCR, revealing that it decreases gradually during fruiting body development. We investigated the relationships between copy number of mitochondrial DNA and transcription levels of genes that are related to activity and number of mitochondria. The lowest expression levels were observed in expression of mitochondrial DNA encoded cytochrome C oxidase subunit 1 gene and NADH dehydrogenase subunit 4 gene at the mature fruit body stage. These results indicate that G. frondosa may deactivate the respiratory chain by decreasing the number of mitochondria during fruiting body development.

Perioperative plasma mitochondrial DNA dynamics and correlation with inflammation during infantile cardiopulmonary bypass

ObjectiveNumerous studies in animals and humans have demonstrated that inflammatory mediators such as tumor necrosis factor (TNF)-α, interleukin (IL)-6, and IL-8 play a role in cardiopulmonary bypass (CPB), which might affect surgical outcomes. Plasma mitochondrial DNA (mtDNA), a recently discovered pro-inflammatory agent, is released by cells upon insult. This study aimed to detect changes in plasma mtDNA levels at different time points after infantile CPB and explore its potential association with inflammatory mediators. MethodsIn the present study, we analyzed the perioperative plasma mtDNA and inflammatory cytokine levels of 48 infants undergoing ventricular septal defect closure. Blood samples were collected before aortic cross-clamping (T1), at the end of CPB (T2), and 6h (T3), 12h (T4), and 24h (T5) post-CPB. Reverse transcription–polymerase chain reaction and specific enzyme-linked immunosorbent assay were used to quantify the plasma mtDNA and inflammatory cytokines, respectively. Bivariate correlation analysis was used to determine the correlations between plasma mtDNA and inflammatory cytokines. ResultsPlasma mtDNA levels increased at T2 and peaked at T3. Significant positive correlations were found between peak plasma mtDNA (at T3) and several inflammatory biomarkers, including IL-6 (at T3) (r=0.62, P<0.001), IL-8 (at T2) (r=0.53, P<0.001), and TNF-α (at T3) (r=0.61, P<0.001). ConclusionHere we report that mtDNA may participate in a systemic inflammatory response to CPB.

High-mobility group box 1 is associated with neurological outcome in patients with post-cardiac arrest syndrome after out-of-hospital cardiac arrest.

BackgroundAlarmins, including high-mobility group box 1 (HMGB-1), can be released from damaged tissues and activated cells as inflammatory mediators. We aimed to evaluate HMGB-1 and mitochondrial DNA dynamics and estimate the prognostic value for neurological outcome in patients with post-cardiac arrest syndrome after out-of-hospital cardiac arrest.MethodsWe evaluated the dynamics of HMGB-1, mitochondrial DNA, and other variables in patients with return of spontaneous circulation after out-of-hospital cardiac arrest. Patients were divided into two groups according to the cerebral performance category at 30 days: the favourable outcome group (cerebral performance categories 1 and 2) and unfavourable group (≥3).ResultsTwenty-one patients were included, and 11 demonstrated favourable outcomes. HMGB-1 levels and mitochondrial DNA on day 1 were significantly higher than on days 2, 3, 5, and 7. Plasma levels of HMGB-1 on day 1 correlated with prognostic parameters (estimated interval to return of spontaneous circulation, lactate, and NH3), tissue damage, systemic inflammation, and disease severity. HMGB-1 on day 1 in the unfavourable group was significantly higher than in the favourable group (median [interquartile range] 15.5 [6.65–18.7], 39.4 [17–69.5], P = 0.009). These findings were not observed regarding mitochondrial DNA. Regarding HMGB-1 prediction accuracy for a good neurological outcome, the area under the receiver operating characteristic curve was 0.864 (95 % confidence interval 0.702, 1.000).ConclusionsHMGB-1 may be involved in acute-phase post-cardiac arrest syndrome pathophysiology, and an increase in plasma levels may be associated with a poor neurological outcome. The study was registered with the University Hospital Medical Information Network Clinical Trials Registry ID: UMIN000006714.Electronic supplementary materialThe online version of this article (doi:10.1186/s40560-016-0161-4) contains supplementary material, which is available to authorized users.

Closed-form stochastic solutions for non-equilibrium dynamics and inheritance of cellular components over many cell divisions.

Stochastic dynamics govern many important processes in cellular biology, and an underlying theoretical approach describing these dynamics is desirable to address a wealth of questions in biology and medicine. Mathematical tools exist for treating several important examples of these stochastic processes, most notably gene expression and random partitioning at single-cell divisions or after a steady state has been reached. Comparatively little work exists exploring different and specific ways that repeated cell divisions can lead to stochastic inheritance of unequilibrated cellular populations. Here we introduce a mathematical formalism to describe cellular agents that are subject to random creation, replication and/or degradation, and are inherited according to a range of random dynamics at cell divisions. We obtain closed-form generating functions describing systems at any time after any number of cell divisions for binomial partitioning and divisions provoking a deterministic or random, subtractive or additive change in copy number, and show that these solutions agree exactly with stochastic simulation. We apply this general formalism to several example problems involving the dynamics of mitochondrial DNA during development and organismal lifetimes.

Mitochondrial DNA dynamics during in vitro culture and pluripotency induction of a bovine Rho0 cell line.

Large number of cellular changes and diseases are related to mutations in the mitochondrial DNA copy number. Cell culture in the presence of ethidium bromide is a known way of depleting mitochondrial DNA and is a useful model for studying such conditions. Interestingly, the morphology of these depleted cells resembles that of pluripotent cells, as they present larger and fragmented mitochondria with poorly developed cristae. Herein, we aimed to study the mechanisms responsible for the control of mitochondrial DNA replication during mitochondrial DNA depletion mediated by ethidium bromide and during the in vitro induction of cellular pluripotency with exogenous transcription factor expression in a bovine model. This article reports the generation of a bovine Rho0 mesenchymal cell line and describes the analysis of mitochondrial DNA copy number in a time-dependent manner. The expression of apoptosis and mitochondrial-related genes in the cells during mitochondrial DNA repletion were also analyzed. The dynamics of mitochondrial DNA during both the depletion process and in vitro reprogramming are discussed. It was possible to obtain bovine mesenchymal cells almost completely depleted of their mitochondrial DNA content (over 90%). However, the production of induced pluripotent stem cells from the transduction of both control and Rho0 bovine mesenchymal cells with human reprograming factors was not successful.

MTRIP: an imaging tool to investigate mitochondrial DNA dynamics in physiology and disease at the single-cell resolution.

Mitochondrial physiology and metabolism are closely linked to replication and transcription of the genome of the organelle, the mitochondrial DNA (mtDNA). However, the characterization of mtDNA processing is poorly defined at the single-cell level. Here, we describe mTRIP (mitochondrial transcription and replication imaging protocol), an imaging approach based on modified fluorescence in situ hybridization (FISH), which simultaneously reveals mitochondrial structures engaged in mtDNA initiation of replication and global mitochondrial RNA (mtRNA) content at the single-cell level in human cells. In addition, mTRIP can be coupled to immunofluorescence for in situ protein tracking, or to MitoTracker, thereby allowing simultaneous labelling of mtDNA, mtRNA, and proteins or mitochondria, respectively. Altogether, qualitative and quantitative alterations of the dynamics of mtDNA processing are detected by mTRIP in human cells undergoing physiological changes, as well as stress and dysfunction, with a potential for diagnostic of mitochondrial diseases.

Direct Quantification of Mitochondria and Mitochondrial DNA Dynamics

Mitochondria are known to be one of major organelles within a cell and to play a crucial role in many cellular functions. These organelles show the dynamic behaviors such as fusion, fission and the movement along cytoskeletal tracks. Besides mitochondria, mitochondrial DNA is also highly motile. Molecular analysis revealed that several proteins are involved in mitochondria and mitochondrial DNA dynamics. In addition to the degeneration of specific nerves with high energy requirement, mutation of genes coding these proteins results in metabolic diseases. During the last few years, a significant amount of relevant data has been obtained on molecular basis of these diseases but mitochondrial dynamics in cells derived from the patients is poorly understood. So far time-lapse fluorescence microscopy, fluorescence recovery after photo bleaching and image correlation methods have been used to study organellar motion. Especially, image correlation method has possibility to evaluate diffusion coefficient of mitochondria and mitochondrial DNA simultaneously and directly. When we search candidates for compounds that modulate mitochondrial dynamics by high throughput screening, image correlation method may be useful although the careful interpretation is required for crowded and heterogeneous environment within a cell.

Image Processing Methods for Quantitative Analysis of Mitochondrial DNA Dynamics

Mitochondria and mitochondrial DNA are known to be highly motile. Molecular analysis revealed that several proteins are bound between mitochondrial DNA and mitochondrial inner membrane. However, nucleoid formed by these proteins is involved in the replication and duplication of mitochondrial DNA and it remains unclear whether mitochondrial DNA keeps binding the mitochondrial inner membrane or not. When nucleoid is necessary during cell cycle, it would be formed probably. When nucleoid is formed in the manner dependent on mitochondrial DNA sequence, the changes in the sequence would result in the equilibrium shifted to dissociation. This implies that the important information on changes in mtDNA accompanied with various symptoms is obtained by quantitative analysis of mtDNA dynamics. In this review, theoretical background of two typical methods, i.e., particle trajectory and image correlation spectroscopy, and several results of the pilot experiment are introduced.

3P325 Direct quantification of mitochondria and mitochondrial DNA dynamics in living cells(Bioimaging,The 48th Annual Meeting of the Biophysical Society of Japan)

3P325 Direct quantification of mitochondria and mitochondrial DNA dynamics in living cells(Bioimaging,The 48th Annual Meeting of the Biophysical Society of Japan)

Mitochondrial DNA ligase function in Saccharomyces cerevisiae.

The Saccharomyces cerevisiae CDC9 gene encodes a DNA ligase protein that is targeted to both the nucleus and the mitochondria. While nuclear Cdc9p is known to play an essential role in nuclear DNA replication and repair, its role in mitochondrial DNA dynamics has not been defined. It is also unclear whether additional DNA ligase proteins are present in yeast mitochondria. To address these issues, mitochondrial DNA ligase function in S.cerevisiae was analyzed. Biochemical analysis of mitochondrial protein extracts supported the conclusion that Cdc9p was the sole DNA ligase protein present in this organelle. Inactivation of mitochondrial Cdc9p function led to a rapid decline in cellular mitochondrial DNA content in both dividing and stationary yeast cultures. In contrast, there was no apparent defect in mitochondrial DNA dynamics in a yeast strain deficient in Dnl4p (Deltadnl4). The Escherichia coli ECO:RI endonuclease was targeted to yeast mitochondria. Transient expression of this recombinant ECO:RI endonuclease led to the formation of mitochondrial DNA double-strand breaks. While wild-type and Deltadnl4 yeast were able to rapidly recover from this mitochondrial DNA damage, clones deficient in mitochondrial Cdc9p were not. These results support the conclusion that yeast rely upon a single DNA ligase, Cdc9p, to carry out mitochondrial DNA replication and recovery from both spontaneous and induced mitochondrial DNA damage.