Tuesday, 20 March 2018

Age-Associated Impairments in Mitochondrial ADP Sensitivity Contribute to Redox Stress in Senescent Human Skeletal Muscle


Graham P. Holloway, Andrew M. Holwerda, Paula M. Miotto, Marlou L. Dirks, Lex B. Verdijk, and Luc J.C. van Loon

  • The authors sought to determine whether there is an age-associated increase in mitochondrial reactive oxygen species (ROS) in vitro, using permeabilized muscle fibres.
  • They find that the capacity of mitochondrial H2O2 emission does not increase with ageing
  • However, ADP sensitivity does reduce with age. Consequently, H2O2 levels increase with age.
  • Increasing muscle mass, strength, and maximal mitochondrial respiration through exercise in older individuals did not alter H2O2 emission rates, the fraction of electron leak to H2O2 or the redox state of muscle.
  • In summary, reduction in mitochondrial ADP sensitivity increases mitochondrial H2O2 emission, which cannot be rescued through resistance training in later life (although there were other benefits to health of these individuals).

Optimized Mitochondrial Targeting of Proteins Encoded by Modified mRNAs Rescues Cells Harboring Mutations in mtATP6


Randall Marcelo Chin, Tadas Panavas, Jeffrey M. Brown, and Krista K. Johnson

  • Allotopic expression where a gene ordinarily encoded by mitochondrial DNA (mtDNA) is placed inside the nucleus, and modified such that the resultant protein is correctly transported into the mitochondria.
  • It is hoped that allotopic expression may be able to rescue pathologies which arise due to mutations in mitochondrial DNA: indeed, allotopic expression-based gene therapy is in phase 3 clinical trials for the mitochondrial disease LHON. 
  • mtDNA-encoded proteins are highly hydrophobic, causing them to often fold into import-incompetent states, thereby preventing them from entering the mitochondria. 
  • Mitochondrial targeting sequences (MTSs) and 3' untranslated regions (3' UTRs) have been used to target proteins or mRNA to the mitochondria. 
  • In this study, the authors performed a screen of 31MTSs and 15 UTRs in their ability to localize up to 9 allotopically expressed proteins to the mitochondrial DNA (note that mtDNA encodes 13 proteins, 22 tRNAs and 2 rRNAs).
  • Cybrid cells harbouring the 8993T>G point mutation in the mtATP6 gene were transiently transfected with a construct which was able to allotopically express mtATP6 and rescue the mtATP6-deficient cells.

Wednesday, 14 March 2018

Mitochondrial DNA as an inflammatory mediator in cardiovascular diseases

Hiroyuki Nakayama and Kinya Otsu


In this review, the authors discuss the role of mitochondria, and especially mitochondrial DNA (mtDNA) in triggering and maintaining cardiac inflammation. In this blost post we only summarise some parts of the review directly related to mtDNA.

We all know that the immune system provides protection against microorganisms such as bacteria, viruses, and fungi. This is achieved by sensing both pathogen-associated molecular patterns (PAMPs) and damage-associated molecular patterns (DAMPs).

One major mechanism for activating the innate immune system is the sensing of pathogen-derived nucleic acids, and this is where mitochondria come into play. Due to their bacterial origin, mtDNA shares similarities with bacterial DNA (e.g. it contains cardiolipin and a predominantly unmethylated CpG motif). Mitochondria also release other DAMPs which can bind and activate multiple pattern recognition receptors similar to those activated by PAMPs.

MtDNA can leave the mitochondria and enter the cytoplasm or leave the entire cell. Opening of the mitochondrial transition pore plays an important role of mtDNA release from mitochondria, as inhibition of pore opening reduced levels of mtDNA in the cyotosol. MtDNA release is also controlled by other regulatory proteins such as the voltage dependent anion channel, Bax, and Bak.

MtDNA released after cell death functions as a DAMP. The mechanism of releasing mtDNA from non-nectrotic cells remains unclear, though exosomal release is proposed to be involved in this mechanism.  MtDNA enters the endocytic pathway by endocytosis and stimulates pattern recognition receptors which eventually leads to inflammasome formation.

It is important to degrade extracellular mtDNA to inhibit unnecessary inflammatory responses. It could be that mtDNA, like other non-host DNA in circulation, is digested in part by circulating nucleases. It is, however, unclear whether this occurs in physiological conditions, especially when mtDNA exists in microvesicles such as exosomes. Inside cells, DNasell plays an important role in mtDNA degradation

Levels of circulating mtDNA increase with age and correlate with levels of pro-inflammatory cytokines. Therefore, mtDNA-induced inflammatory responses can be involved in age-related cardiovascular disease, heart failure and atherosclerosis.

During heart failure, multiple endogenous DAMPs (including mtDNA) are released and recognized to induce an inflammatory response. However, no associated was found between the severity of heart failure and mtDNA levels in serum of patients (patients do show much higher serum mtDNA levels compared to controls).


Monday, 12 March 2018

Efficient termination of nuclear lncRNA transcription promotes mitochondrial genome maintenance


Dorine Jeanne Mariëtte du Mee, Maxim Ivanov, Joseph Paul Parker, Stephen Buratowski, Sebastian Marquardt

  • Most of the DNA of eukaryotes does not code for protein, yet many such regions of non-coding DNA are still transcribed into RNA (these are called lncRNA). Understanding the biological functions of these large expanses of non-coding DNA is an active area of current research.
  • Here, the authors show that a particular non-coding RNA in budding yeast (CUT60) is required for the proper transcription of its neighbouring gene ATP16. Mutations in CUT60 could result in the fusion of the lncRNA with the RNA of ATP16, and consequently the ATP16 protein could not be produced.
  • Interestingly, this had the consequence of yeast cells losing their mitochondrial DNA. ATP16 constitutes a subunit of ATP-synthase, which perhaps explains this striking phenotype.
  • The authors speculate that loss of mtDNA triggered by controlled, transient, transcription termination efficiency of CUT60 could allow cells to detoxify themselves of deleterious mtDNA 
The authors raise an interesting idea that yeast cells may be able to cleans their mtDNA through reducing CUT60 transcription termination efficiency. They suggest that cells may shed their mtDNA, and then gain healthy copies of molecules through mating. We know from animals that, during development, the mtDNA bottleneck serves to cleanse the developing embryo of deleterious mtDNA mutations. I wonder whether yeast cells could use this mechanism to a gentler extent (not completely losing their mtDNA, just reducing their mtDNA copy number) to bottleneck their mtDNA?

Thursday, 1 March 2018

Hallmarks of Cellular Senescence


Alejandra Hernandez-Segura, Jamil Nehme, Marco Demaria

A senescent cell is one which permanently stops dividing. In vitro, this can be caused by various stimuli, although it is unclear which amongst these cause senescence in vivo. The accumulation of senescent cells is observed through ageing, and a growing body of evidence is pointing towards the removal of senescent cells as a strategy to combat ageing.

Types of senescence currently known:

- DNA damage-induced senescence. This can be induced in vitro through radiation or drugs
- Oncogene-induced senescence. Activation of oncogenes (e.g. Ras or BRAF) or inactivation of tumour suppressors (e.g. PTEN) can induce senescence
- Chemotherapy-induced senescence. Drugs such as bleomycin or doxorubicin induce DNA damage. Drugs such as abemaciclib and palbociclib can inhibit cyclin-dependent kinases which regulate the cell cycle
- Mitochondrial dysfunction-associated senescence. The so-called "senescence associated secretory phenotype" (SASP) appears to be characteristic of this kind of senescence
- Epigenetically induced senescence. Inhibitors of DNA methylases or histone deacetylases can cause senescence
- Paracrine senescence. Senescence can be induced via the SASP produced by primary senescent cells

The senescence phenotype is often characterised by:

- Activation of a chronic DNA damage response
- Engagement of various cyclin-dependent kinase inhibitors
- SASP (which comprises, in part, various proinflamatory and tissue-remodelling factors)
- Induction of anti-apoptotic genes
- Altered metabolic rates
- Endoplasmic reticulum stress
- Consequent to the above, senescent cells are: enlarged and more flattened; have altered plasma membrane composition; accumulate lysosomes and mitochondria.

Current methods which are used to detect senescent cells include:

- DNA damage response: Immunostaining for γ-H2AX, p53
- Cell cycle arrest: Measurement of colony-formation potential or DNA synthesis rate via BrdU/EdU-incorporation. Expression level of the cyclin-dependent kinase inhibitors p16 and p21.
- Secretory phenotype: Cytokines (IL-1a, IL-6 and IL-8) , chemokines (CCL2) and metalloproteinases (MMP-1, MMP-3). However, the SASP is heterogeneous.
- Apoptosis resistance: Upregulation of BCL-proteins, BCL-2, Bcl-w or Bcl-xL.
- Cell size: Enlarged cell body and irregular shape using bright-field microscopy. Immunofluorescence targetting vimentin, actin or other cytoplasmic proteins have been used.
- Increased lysosomal content: e.g. SA-βgal, SSB, GL13, LysoTrackers, orange acridine
- Accumulation of mitochondria: MitoTrackers

Transitional correlation between inner-membrane potential and ATP levels of neuronal mitochondria

In this paper, the authors simultaneously measure mitochondrial membrane potential (Δψ) and mitochondrial ATP production (ATPmito) in dorsal root ganglion neurons from rat embryos. They measure the dynamics of Δψ and ATPmito, as well as their correlation, during physiological neuronal activity and focus on the following questions:

Is there a relation between Δψ, ATPmito and 
  • mitochondrial size?
  • mitochondrial transport velocity?
  • mitochondrial transport direction?
 Furthermore, they ask the question
  • What happens to Δψ and ATPmito during mitochondrial fusion and fission events? 20 fusion events and 20 fission events were investigated.

Some of the Results:
  • Δψ and ATPmito were compared among anterogradely transported, retrogradely transported, and stationary mitochondria in axons. Retrogradely transported mitochondria had slightly lower Δψ compared to anterogradely transported mitochondria.
  • No correlation was found between Δψ, ATPmito and mitochondrial velocity and transported distance.
  • Post-fusion mitochondrial membrane potential Δψ seemed to be higher than the average of the pre-fusion potentials (i.e. Δψfused > 0.5 (Δψpre-1 + Δψpre-2)).*
  • ATPmito was higher in the post-fusion mitochondrion compared to the average of the two pre-fusion mitochondria
  •  The two post-fission mitochondria tended to have different values for
    Δψ and their average was typically lower than the pre-fission potential (again, no information on size was provided as discussed below*).
  • No changes in ATPmito were observed upon fission
  • Mitochondrial density was higher in growth cones (an extension of a developing or regenerating neurite seeking its synaptic target) compared to axons. 
  • Average ATPmito levels were slightly lower in growth cones, though integrated ATP levels (over all mitochondria) were higher. 
  • Average Δψ was higher in growth cones compared to axons
  • Higher ATP levels in growth cones led to faster elongation of the axon, though no correlation between elongation speed and Δψ was found.
  • ATPmito tends to follow a change in Δψ (i.e. the change in Δψ occurs first). 
    ATPmito and Δψ are not necessarily always correlated.
  • Various other results were obtained which you can find by reading the paper!

*We note that no information was provided regarding mitochondrial size. If one of the pre-fusion mitochondria is much larger than the other, we might expect the membrane potential of the former to have more influence on the final potential of the post-fusion mitochondrion. In this case, one would not expect  Δψfused to be the arithmetic average of the two pre-fusion potentials.

Wednesday, 28 February 2018

Circadian Control of DRP1 Activity Regulates Mitochondrial Dynamics and Bioenergetics

Schmitt K, Grimm A, Dallmann R, Oettinghaus B, Restelli LM, Witzig M, Ishihara N, Mihara K, Ripperger JA, Albrecht U, Frank S, Brown SA, Eckert A

  • Circadian regulation of dynamin-related protein 1 (DRP1), a key mitochondrial fission protein, results in daily cycles of fission and fusion which are essential for circadian oscillations in ATP production
  • Genetic and pharmacological abrogation of DRP1 activity abolished circadian network dynamics and eliminated circadian ATP production

Wednesday, 14 February 2018

Myosin VI-Dependent Actin Cages Encapsulate Parkin-Positive Damaged Mitochondria


Antonina J. Kruppa,Chieko Kishi-Itakura, Thomas A. Masters, Joanna E. Rorbach, Guinevere L. Grice, John Kendrick-Jones, James A. Nathan, Michal Minczuk, Folma Buss

  • The authors identify a protein MYO6 which triggers the formation of F-actin cages to form around damaged mitochondria (mitochondria were damaged using a variety of pharmacological means)
  • These cages form a physical barrier, preventing damaged mitochondria from refusing with the network
  • MYO6 interacts with other proteins known to recruit autophagosomes to damaged mitochondria

Friday, 9 February 2018

Affinity purification of cell-specific mitochondria from whole animals resolves patterns of genetic mosaicism


Arnaud Ahier, Chuan-Yang Dai, Andrea Tweedie, Ayenachew Bezawork-Geleta, Ina Kirmes & Steven Zuryn

  • The authors demonstrate a technique called Cell-specific mitochondrial affinity purification (CS-MAP) to yield intact, functional, mitochondrial with >96% enrichment, >96% purity at single-cell and single-animal resolution in C. elegans.
  • CS-MAP consists of tagging the outer mitochondrial membrane protein TOMM-20 with a fluorophore and a particular epitope (which is something that an antibody can bind to) called HA. They placed the fusion protein under the control of a tissue-specific promoter so that e.g. muscle-specific mitochondria could be tagged. Mitochondria can then be purified using magnetic beads coated with anti-HA antibody and performing immunoprecipitation.
  • The authors crossed worms containing the CS-MAP construct with animals containing mitochondrial DNA deletions, and analysed the heteroplasmy in the individual mitochondria purified from different cell types and compared this to homogenate heteroplasmy across the whole animal. The authors found that intestine and neurons had significantly lower heteroplasmy than the homogenate, whereas the germline had significantly higher heteroplasmy than the homogenate.
  • The authors also quantified mtDNA copy number per mitochondrion in a number of different tissues, finding that the germline had ~3.5 mtDNAs per mitochondrion whereas tissues such as neurons and the intestine had ~1.5 mtDNAs per mitochondrion.
  • Using three-dimensional reconstruction from fluorescence images, the authors found that individual germ cells contained 71.2+/-6.5 mtDNAs per cell, whereas neurons contained 14.4+/-0.5 mtDNAs per cell.
  • The authors suggest that mtDNA turnover is higher in germ cells, which may account for their observations of increased copy number and heteroplasmy in the germline

Check out a pre-print from our group with some ideas on how increases in copy number may be related to heteroplasmy, and thoughts about comparisons between homogenate heteroplasmy and cellular heteroplasmy

An energetic view of stress: Focus on mitochondria


Martin Picard, Bruce S McEwen, Elissa S Epel & Carmen Sandi

In this review, the authors discuss the link between mitochondria and mental stress. 

  • Allostasis is defined as the active (i.e. energy-requiring) process of achieving stability, or homeostasis, through physiological or behavioural change. This includes neuroendocrine, autonomic, epigenetic, metabolic and immune changes, and is generally a short-term adaptation when regulated in a healthy setting.
  • When allostatic mediators are not turned off, these same mediators can cause unhealthy changes in the brain and body: these are the pathophysiological consequences of stress. The authors refer to "allostatic load" as the pathophysiological consequences of chronic dysregulation of allostatic mediators.
  • Metabolic intermediates that are the substrates or co-factors for epigenetic modifications are all derived from the Krebs cycle and other metabolic pathways within mitochondria. Some examples are discussed within the review. Hence, both the addition and removal of epigenetic marks are metabolically/mitochondrially regulated.
  • Mitochondria are the site of synthesis for all steroid hormones, including glucocorticoids such as cortisol (the archetypal stress hormone), androgens such as testosterone and estrogens such as estriol. Norepinephrine and epinephrine are also hormones (called catecholamines) which are released in response to certain stressors. Enzymes involved in the degradation of catecholamines (MAO-A and MAO-B) are anchored to the outer mitochondrial membrane.
  • Glucocorticoids (GCs) increase blood glucose levels by acting on the liver, skeletal muscles and adipose tissue by targetting the glucocorticoid receptor (GR). In the liver, GR activation has been shown to induce chromatin remodelling (an epigenetic effect). In skeletal muscle, GCs antagonize several elements of insulin signalling, and inhibits the uptake of pyruvate by mitochondria.
  • Humans with higher circulating levels of cortisol under resting conditions also have higher levels of glucose, triglycerides and higher insulin resistance (essentially a pre-diabetic state). In mice, chronic GC administration results in glucose intolerance, elevated triglycerides, weight gain and depressive behaviour.
  • Under healthy conditions, GCs are associated with the proper maintainance of a diurnal cycle.
  • Some, but not all, synapses in many parts of the cerebral cortex turn over during the diurnal cycle. Interfering with the daily cycle of GCs can impair motor learning in humans.
  • An animal model of shift work caused dendrites to shrink in the prefrontal cortex and the animal to become cognitively rigid, as well as gaining weight and becoming insulin resistant.

Wednesday, 7 February 2018

Mitochondrial levels determine variability in cell death by modulating apoptotic gene expression


Silvia Márquez-Jurado, Juan Díaz-Colunga, Ricardo Pires das Neves, Antonio Martinez-Lorente, Fernando Almazán, Raúl Guantes & Francisco J. Iborra

Chemotherapies often leave a proportion of cancer cells behind. Even genetically identical cells grown in vitro show this effect, suggesting that there exists some level of non-genetic heterogeneity in cancer cells. 

Two well-known pathways are able to induce cell death: the intrinsic and extrinsic pathways. The intrinsic pathway (which does not involve signalling from outside of the cell) directly involves mitochondria. In contrast, the extrinsic pathway may be activated via the binding of specific ligands to cell death receptors on the plasma membrane of the cell and does not directly involve mitochondria. However, several proteins may participate in both the intrinsic and extrinsic pathways, meaning that these pathways have cross-talk. 

TNF-related apoptosis-inducing ligand (TRAIL) is a protein which may induce the extrinsic apoptosis pathway. When cells are treated with TRAIL, the authors observe that the fraction of cells which are killed saturates at 35% with the concentration of TRAIL, and that there is heterogeneity in the time to death at every concentration of TRAIL. The authors also observed (as noted previously by other authors) that sister cells tended to have the same fate and very similar times to death (Pearson correlation >0.8).

The authors investigated how mitochondrial content affected cell death propensity. They found that:
  • Cells with higher mitochondrial mass (as determined by Mitotracker Green with 24 hr live-cell imaging) were more likely to die under TRAIL (as well as other cell death inducing drugs such as CHX and DRB).
  • Cells show a weak correlation between time to death and mitochondrial mass (rho = -0.47) for intermediate TRAIL concentrations.
  • After sorting cells into mito-high and mito-low fractions, (a fold-change of ~x5 between fractions), mito-high fractions had ~x3 more RNA than mito-low.
  • Mitochondrial mass contributed to around 50% of the total variability observed in proteins which participate in apoptosis (both pro-apoptotic and anti-apoptotic).
  • Including the observed correlation between mitochondrial mass and protein levels of apoptosis genes in a pre-existing mathematical model of the extrinsic cell death pathway in HeLa cells was able to recapitulate many of the authors' experimental observations.

The authors also investigated whether these results hold in real tumours, as opposed to cell culture conditions where environmental noise is minimised. The authors stained sections from colon cancer biopsies with antibodies against Aconitase 2 (for mitochondrial mass) and various cell death proteins. Whilst the mitochondrial contribution to variability for some apoptotic proteins was lost, others were retained (where pro-apoptotic proteins tended to have a higher correlation with mitochondrial mass than anti-apoptotic proteins).
Does cell volume confound any of these observations?

Friday, 19 January 2018

Identification of New Activators of Mitochondrial Fusion Reveals a Link between Mitochondrial Morphology and Pyrimidine Metabolism


Laia Miret-Casals, David Sebastián, José Brea, Eva M.Rico-Leo, Manuel Palacín, Pedro M.Fernández-Salguero, M. Isabel Loza, Fernando Albericio, Antonio Zorzano

  • The authors develop a high-throughput drug screen on HeLa cells to identify FDA-approved drugs which modulate the activity of the mitochondrial fusion protein MFN2, allowing the authors to find compounds which are able to upregulate MFN2 expression and induce mitochondrial fusion.
  • The authors identify leflunomide (a drug used for the treatment of arthritis) as the most potent modulator of MFN2 expression, inducing a 67% increase in MFN2 mRNA levels. The compound was also found to increase both MFN1 and MFN2 protein levels by a factor of ~x2. HeLa cells morphologically appeared to have higher fusion and mitochondrial membrane potential.
  • Leflunomide inhibits de novo synthesis of pyrimidines by inhibiting the mitochondrial inner membrane enzyme dihydroorotate dehydrogenase (DHODH).
  • As a consequence, leflunomide had anti-proliferative effects upon cells.
  • Uridine may be added to cells as an external source of pyrimidines. The authors found that addition of uridine to leflunomide-treated cells abolished the ability of leflunomide to induce MFN2 expression.
  • Another drug, brequinar sodium (BRQ), which is an inhibitor of DHODH also has similar properties to leflunomide.
  • Hence, inhibition of pyrimidine nucleotide synthesis may induce mitochondrial elongation via MFN induction.
  • DHODH uses ubiquinone as a substrate, which is converted to ubiquinol. Ubiquinol is substrate of complex III of the respiratory chain.
  • Inhibiting DHODH therefore inhibits the cycling of ubiquinone -> ubiquinol -> ubiquinone, and therefore inhibits the activity of complex III. 
  • The authors found that direct inhibition of complex III via the drug myxothiazol inhibited DHODH activity, reflecting the coupling between pyrimidine synthesis via DHODH and complex III activity.
  • Complex III inhibition via myxothiazol induced MFN induction and also elongation of mitochondria, even in MFN knockout cells. 
Overall, depletion of pyrimidine pools by complex III inhibition causes cell-cycle arrest and promotes mitochondrial elongation as an adaptive response to energetic stress.

Wednesday, 17 January 2018

Pervasive within-Mitochondrion Single-Nucleotide Variant Heteroplasmy as Revealed by Single Mitochondrion Sequencing


Morris J, Na YJ, Zhu H, Lee JH, Giang H, Ulyanova AV, Baltuch GH, Brem S, Chen HI, Kung DK, Lucas TH, O'Rourke DM, Wolf JA, Grady MS, Sul JY, Kim J, Eberwine J

This study looks at the prevalence of mutations in mitochondrial DNA within single mitochondria. The authors do this by collecting single mitochondria from cells with a micropipette, then perform PCR to amplify the copy number of DNA and finally illumina deep sequencing.

The authors collected 118 samples from the brains of lab mice (C57BL/6N strain), and found on average 3.9 single-nucleotide variants per mitochondrion with a standard deviation of 5.71 (although the mtDNA copy number per mitochondrion was not quantified). Some of the mutations observed are thought to be deleterious: for instance, a mutation found at position 9027 (G>A) encoding MT-CO3 (complex III of the respiratory chain) is a missense mutation, annotated to have moderate pathophysiological impact. The authors found 59 samples with this mutation. The intra-mitochondrial heteroplasmy was > 90% for 39 of these mitochondrial samples.

The authors also collected 21 samples of mitochondria from 8 different neurons from the brain of a 63-year-old female using residual tissue removed after surgery. From these samples, the authors found that within-mitochondrion heteroplasmy was ~50% less common in their human sample than in lab mice.  The authors also found that the within-mitochondrion heteroplasmy of different mitochondria in the same cell, and the inter-cellular heteroplasmy between cells, tended to be similar in their human sample but different in mouse.

The authors suggest that the differences between humans and mice are most likely due to the effect of only observing a single individual for their human experiment, but many individuals for mice. The authors found a large effect from the identity of the mother in determining the extent of within-mitochondrion heteroplasmy.

Wednesday, 10 January 2018

How cells adapt to progressive mitochondrial mutation

Mitochondria produce the cell's major energy currency: ATP. If mitochondria become dysfunctional, this can be associated with a variety of devastating diseases, from Parkinson's disease to cancer. Technological advances have allowed us to generate huge volumes of data about these diseases. However, it can be a challenge to turn these large, complicated, datasets into basic understanding of how these diseases work, so that we can come up with rational treatments.

We were interested in a dataset (see here) which measured what happened to cells as their mitochondria became progressively more dysfunctional. A typical cell has roughly 1000 copies of mitochondrial DNA (mtDNA), which contains information on how to build some of the most important parts of the machinery responsible for making ATP in your cells. When mitochondrial DNA becomes mutated, these instructions accumulate errors, preventing the cell's energy machinery from working properly. Since your cells each contain about 1000 copies of mitochondrial DNA, it is interesting to think about what happens to a cell as the fraction of mutated mitochondrial DNA (called 'heteroplasmy') gradually increases. We used maths to try and explain how a cell attempts to cope with increasing levels of heteroplasmy, resulting in a wealth of hypotheses which we hope to explore experimentally in the future.


The central idea arising from our analysis of this large dataset is that cells attempt to maintain the number of normal mtDNAs per cell volume as heteroplasmy initially increases from 0% mutant. We suggest they do this by shrinking their size. By getting smaller, cells are able to reduce their energy demands as the fraction of mutant mtDNA increases, allowing them to balance their energy budget and maintain energy supply = demand. However, cells can only get so small and eventually the cell must change its strategy. At a critical fraction of mutated mtDNA (h* in the cartoon above), we suggest that cells switch on an alternative energy production mode called glycolysis. This causes energy supply to increase, and as a result, cells grow larger in size again. These ideas, as well as experimental proposals to test them, are freely available in our latest publication in Biochemical Journal. Juvid, Iain and Nick.