Dissecting the role of the mitochondrial carrier SLC25A47

Transporters of the SLC25 mitochondrial carrier superfamily bridge cytoplasmic and mitochondrial metabolism by channeling metabolites across mitochondrial membranes and are pivotal for metabolic homeostasis. Despite their physiological relevance as gatekeepers of cellular metabolism, most of the SLC25 family members remain uncharacterized. During my PhD studies, I investigated the function of SLC25A47, an orphan carrier uniquely expressed in hepatocytes. We used a murine loss-of-function (LOF) model to unravel the role of this transporter in mitochondrial and hepatic homeostasis. Slc25a47hep-/- mice displayed a wide variety of metabolic abnormalities, as a result of sustained energy deficiency in the liver originating from impaired mitochondrial respiration in this organ. This mitochondrial phenotype was associated with a robust activation of the mitochondrial stress response (MSR) in the liver, which in turn, induced the secretion of several mitokines, amongst which FGF21 played a preponderant role in the translation of the effects of the MSR on systemic physiology. To dissect the FGF21-dependent and -independent physiological changes induced by the loss of Slc25a47, we generated a double Slc25a47-Fgf21 LOF mouse model, and demonstrated that several aspects of the hypermetabolic state were entirely driven by hepatic secretion of FGF21. On the other hand, the metabolic fuel inflexibility induced by loss of Slc25a47 could not be rescued by genetic removal of Fgf21. Finally, we challenged Slc25a47hep-/- mice with high-fat high-sucrose (HFHS) diet and observed the development of liver fibrosis.Collectively, the data presented in this thesis show that SLC25A47 is a novel determinant of hepatic metabolism and place this carrier at the center of mitochondrial homeostasis.

Mitochondrial stress signalling and its impacts on physiology

Adrienne Joëlle Laurence Mottis

Mitochondria are responsible for respiration and energy harvesting across the eukaryotic kingdom. They also take part in other cellular processes like intermediary metabolism. As a result, mitochondrial function directly impacts organismal features such as metabolic homeostasis, fitness and aging. Moreover, mitochondrial dysfunction is involved in numerous pathologies, such as neurodegenerative disease, obesity, diabetes and cancer. Several surveillance pathways constantly monitor mitochondria to ensure their proper function. Among the pathways triggered by stress, the mitochondrial unfolded protein response (UPRmt) aims to restore proteostasis within this organelle. A better understanding of the cellular response to mitochondrial stress would expand our fundamental knowledge of physiological and pathological processes involving mitochondria, leading to potential new therapeutics that target these organelles. This thesis focuses on the mechanistic and physiological characterization of the mitochondrial stress response in several organisms. I used the model organism Caenorhabtis elegans to screen for novel players of the UPRmt and found the poly(A)-binding protein pab-1. Using transcript profiling, we showed that the induction of immune genes is a common consequence of several mitochondrial stressors. We demonstrated that pab-1 regulates the activation of the mitochondrial stress response and innate immunity as well. On top, pab-1 is required for the survival of C. elegans upon bacterial infection. Transcriptomic data from multiple human tissues suggest that the human pab-1 orthologue, PABPC1, has a conserved role in immunity. In mice, I explored the regulation of the mitochondrial stress response using a combination of bioinformatics and in vivo experimental approaches. Combined transcriptomic and proteomic analyses in the BXD mouse genetic reference population revealed a tight co-regulation of the orthologues of the UPRmt under normal physiological conditions. As a complementary approach, we triggered mitochondrial stress pharmacologically in newly born and adult mice. Due to the bacterial ancestry of mitochondria, the effects of the antibiotic doxycycline (dox) are also deleterious to this organelle. We found that dox impairs mitochondrial proteostasis and oxygen consumption in adult mice. However, mitochondrial stress induced by post-natal dox treatment did not cause long-lasting effects on mouse physiology and longevity, which is very distinct from observations in C. elegans. The mammalian gut flora plays an important role in organismal homeostasis. The use of an antibiotic causes perturbations of the microbiota, with repercussions on metabolism, inflammation, and the nervous system. To eliminate these confounding effects, we also treated germ-free mice with dox to characterize the response of the mouse to a so-called âpureâ mitochondrial stress. Using multi-omics profiling, we found that organs highly dependent on mitochondria display specific transcriptomic and proteomic signatures following dox treatment. In the kidney, we observed an inhibition of translation and of the mTOR pathway, accompanied by an activation of the ATF4 integrated stress response (ISR). In the liver, dox led to a remodelling of lipid metabolism. In both organs, target transcripts of type I interferon anti-viral response were induced. This thesis demonstrates that the response to mitochondrial stress is a multi-faceted process with conserved aspects acr

EPFL2018

Mitochondrial unfolded protein response, its molecular mechanism and physiological impact

Virginija Jovaisaite

Mitochondria are the main producers of ATP, the energy currency of the cell, and they perform a wide array of other fundamental functions. These organelles are essential not only for cellular metabolism, but also for organismal physiology and lifespan. A recently discovered mitochondrial unfolded protein response (UPRmt) maintains the mitochondrial proteostasis by managing protein quality control (PQC) network during proteotoxic stress. Recent findings show that activation of this repair pathway improves mitochondrial function and is associated with increased longevity. My thesis aims to expand the knowledge about the UPRmt by exploring its molecular mechanism and impact on mitochondrial stress induced longevity. We performed three studies: Identification of two epigenetic UPRmt regulators (chapter 2). In collaboration with Prof. Dillinâs laboratory (UCB, USA), we identified the conserved histone demethylases jmjd-1.2/PHF8 and jmjd-3.1/JMJD3 as positive regulators of UPRmt and longevity in response to mitochondrial electron transport chain (ETC) dysfunction across species. A systems genetics analysis of data from the BXD mouse genetic reference population (GRP) further indicates conserved roles of the mammalian orthologs of these demethylases in longevity and UPRmt signaling. These findings illustrate an evolutionary conserved epigenetic mechanism that determines the rate of aging, downstream of mitochondrial perturbations. Relation between UPRmt and TCA cycle (chapter 3). We investigated the effects of genetic perturbations of the TCA cycle on the UPRmt and lifespan. We found that downregulation of several TCA cycle enzymes induces UPRmt, but they have divergent effects on lifespan. Restricting the knockdown of these genes to early larval development stages had positive effects on longevity, which was dependent on UPRmt. We analysed the transcriptomes of C. elegans with knockdown of these TCA cycle genes and compared them with those obtained after the knockdown of other mitochondrial genes, which are known to induce the UPRmt. We identified a core set of transcripts, which change in common and which likely represent a signature of mitochondrial dysfunction associated with UPRmt induction. Bioinformatic analysis of UPRmt (chapter 4). In two collaborative studies, we investigated the UPRmt using newly collected data from the BXD mouse GRP. In the first study, we quantified the transcriptome and proteome from 40 strains of the BXD mouse GRP on two different diets. Integrated molecular profiles were used to characterize the UPRmt, which shows strikingly variant responses at the transcript and protein level that are conserved between C.elegans, mice and humans. In the second study, whole genomes of BXD GRP strains were sequenced to detect sequence variants, which were then used to find novel associations within a high coverage phenome of various traits. One of the novel associations included a missense mutation in fumarate hydratase that controls variation in the UPRmt in both mouse and C. elegans. In summary, our work characterized novel epigenetic regulators of UPRmt and associated mitochondrial stress induced longevity, described the connection between the UPRmt and the TCA cycle, and illustrated the conservation of UPRmt in a mouse GRP. These results expand the knowledge about the UPRmt and promote further investigation of its role in longevity and mitochondrial diseases.

EPFL2016

Calorie Restriction-like Effects of 30 Days of Resveratrol Supplementation on Energy Metabolism and Metabolic Profile in Obese Humans

Johan Auwerx, Richardus Houtkooper, Dongryeol Ryu

Resveratrol is a natural compound that affects energy metabolism and mitochondrial function and serves as a calorie restriction mimetic, at least in animal models of obesity. Here, we treated 11 healthy, obese men with placebo and 150 mg/day resveratrol (resVida) in a randomized double-blind crossover study for 30 days. Resveratrol significantly reduced sleeping and resting metabolic rate. In muscle, resveratrol activated AMPK, increased SIRT1 and PGC-1α protein levels, increased citrate synthase activity without change in mitochondrial content, and improved muscle mitochondrial respiration on a fatty acid-derived substrate. Furthermore, resveratrol elevated intramyocellular lipid levels and decreased intrahepatic lipid content, circulating glucose, triglycerides, alanine-aminotransferase, and inflammation markers. Systolic blood pressure dropped and HOMA index improved after resveratrol. In the postprandial state, adipose tissue lipolysis and plasma fatty acid and glycerol decreased. In conclusion, we demonstrate that 30 days of resveratrol supplementation induces metabolic changes in obese humans, mimicking the effects of calorie restriction.