Roles of sirtuins in the regulation of antioxidant defense and bioenergetic function of mitochondria under oxidative stress

Abstract

In addition to serving as the power house of mammalian cells, mitochondria are crucial for the maintenance of cellular homeostasis in response to physiological or environmental changes. Several lines of evidence suggest that posttranslational modification (PTM) of proteins plays a pivotal role in the regulation of the bioenergetic function of mitochondria. Among them, reversible lysine acetylation of mitochondrial proteins has been established as one of the key mechanisms in cellular response to energy demand by modulating the flux of a number of key metabolic pathways. In this article, we focus on the role of Sirt3-mediated deacetylation in: (1) flexibility of energy metabolism, (2) activation of antioxidant defense, and (3) maintenance of cellular redox status in response to dietary challenge and oxidative stress. We suggest that oxidative stress-elicited down-regulation of Sirt3 plays a role in the pathophysiology of diabetes, cardiac hypotrophy, mitochondrial diseases, and age-related diseases. Besides, the physiological role of newly identified lysine acylation mediated by Sirt5 and its biochemical effects on oxidative metabolism are also discussed. Moreover, we have integrated the regulatory function of several protein kinases that are involved in the phosphorylation of mitochondrial enzymes during oxidative stress. Finally, the functional consequence of the synergistic regulation through diverse protein modifications is emphasized on the maintenance of the bioenergetic homeostasis and metabolic adaptation of the animal and human cells. Together, we have provided an updated review of PTM in mitochondrial biology and their implications in aging and human diseases through an intricate regulation of energy metabolism under oxidative stress.     

Something worth dyeing for: Molecular tools for the dissection of lipid metabolism in Caenorhabditis elegans

The nematode Caenorhabditis elegans (C. elegans) has during the last decade emerged as an invaluable eukaryotic model organism to understand the metabolic and neuro-endocrine regulation of lipid accumulation. The fundamental pathways of food intake, digestion, metabolism, and signalling are evolutionary conserved between mammals and worms making C. elegans a genetically and metabolically extremely tractable model to decipher new regulatory mechanisms of lipid storage and to understand how nutritional and genetic perturbations can lead to obesity and other metabolic diseases. Besides providing an overview of the most important regulatory mechanisms of lipid accumulation in C. elegans, we also critically assess the current methodologies to monitor lipid storage and content as various methods differ in their applicability, consistency, and simplicity.     

Neural melanocortin receptors in obesity and related metabolic disorders

Obesity is a global health issue, as it is associated with increased risk of developing chronic conditions associated with disorders of metabolism such as type 2 diabetes and cardiovascular disease. A better understanding of how excessive fat accumulation develops and causes diseases of the metabolic syndrome is urgently needed. The hypothalamic melanocortin system is an important point of convergence connecting signals of metabolic status with the neural circuitry that governs appetite and the autonomic and neuroendocrine system controling metabolism. This system has a critical role in the defense of body weight and maintenance of homeostasis. Two neural melanocortin receptors, melanocortin 3 and 4 receptors (MC3R and MC4R), play crucial roles in the regulation of energy balance. Mutations in the MC4R gene are the most common cause of monogenic obesity in humans, and a large literature indicates a role in regulating both energy intake through the control of satiety and energy expenditure. In contrast, MC3Rs have a more subtle role in energy homeostasis. Results from our lab indicate an important role for MC3Rs in synchronizing rhythms in foraging behavior with caloric cues and maintaining metabolic homeostasis during periods of nutrient scarcity. However, while deletion of the Mc3r gene in mice alters nutrient partitioning to favor accumulation of fat mass no obvious role for MC3R haploinsufficiency in human obesity has been reported. This article is part of a Special Issue entitled: Modulation of Adipose Tissue in Health and Disease.     

Whole-genome RNAi screen identifies methylation-related genes influencing lipid metabolism in Caenorhabditis elegans

Lipid droplets (LDs) are highly conserved multifunctional cellular organelles and aberrant lipid storage in LDs can lead to many metabolic diseases. However, the molecular mechanisms governing lipid dynamic changes remain elusive, and the high-throughput screen of genes influencing LD morphology was limited by lacking specific LD marker proteins in the powerful genetic tool Caenorhabditis elegans. In this study, we established a new method to conduct whole-genome RNAi screen using LD resident protein DHS-3 as a LD marker, and identified 78 genes involved in significant LD morphologic changes. Among them, mthf-1, as well as a series of methylation-related genes, was found dramatically influencing lipid metabolism. SREBP-1 and SCD1 homologs in C. elegans were involved in the lipid metabolic change of mthf-1(RNAi) worms, and the regulation of ATGL-1 also contributed to it by decreasing triacylglycerol (TAG) hydrolysis. Overall, this study not only identified important genes involved in LD dynamics, but also provided a new tool for LD study using C. elegans, with implications for the study of lipid metabolic diseases.     

Proteomic analysis of mitochondria from Caenorhabditis elegans

Mitochondria play essential roles in cell physiological processes including energy production, metabolism, ion homeostasis, cell growth, aging and apoptosis. Proteomic strategies have been applied to the study of mitochondria since 1998; these studies have yielded decisive information about the diverse physiological functions of the organelle. As an ideal model biological system, the nematode Caenorhabditis elegans has been widely used in the study of several diseases, such as metabolic diseases and cancer. However, the mitochondrial proteome of C. elegans remains elusive. In this study, we purified mitochondria from C. elegans and performed a comprehensive proteomic analysis using the shotgun proteomic approach. A total of 1117 proteins have been identified with at least two unique peptides. Their physicochemical and functional characteristics, subcellular locations, related biological processes, and associations with human diseases, especially Parkinson's disease, are discussed. An orthology comparison was also performed between C. elegans and four other model organisms for a general depiction of the conservation of mitochondrial proteins during evolution. This study will provide new clues for understanding the role of mitochondria in the physiological and pathological processes of C. elegans.     

Caenorhabditis elegans ucp-4 regulates fat metabolism: suppression of ucp-4 expression induced obese phenotype and caused impairment of insulin like pathway

Uncoupling proteins, a family of proton carriers located in the inner mitochondrial membrane, have important functions in energy metabolism and free radical generation that are relevant to mitochondrial function. Five family members have been identified, UCP1-5, that have distinct tissue distributions, and differences and similarities in physiological function. Uncoupling protein 4 (UCP4) is highly expressed and has a unique function in brain. UCP4 appears to be involved with metabolism in neurons and adipocytes, but conclusions on this protein have been controversial. Here, we used Caenorhabditis elegans to explore the functions of ucp-4, particularly in fat metabolism. Our results showed that UCP4 knockdown induced an obese phenotype and impaired the insulin-like pathway, possibly via oxidative stress in C. elegans. This highlights the importance of studying the role of ucp-4 in fat metabolism.     

Haploinsufficiency of Akt1 Prolongs the Lifespan of Mice

There is increasing evidence that nutrient-sensing machinery is critically involved in the regulation of aging. The insulin/insulin-like growth factor-1 signaling pathway is the best-characterized pathway with an influence on longevity in a variety of organisms, ranging from yeast to rodents. Reduced expression of the receptor for this pathway has been reported to prolong the lifespan; however, the underlying mechanisms are largely unknown. Here we show that haploinsufficiency of Akt1 leads to an increase of the lifespan in mice. Akt1 ^+/– mice had a lower body weight than their littermates with less fat mass and normal glucose metabolism. Ribosomal biogenesis and the mitochondrial DNA content were significantly reduced in these mice, along with a decrease of oxidative stress. Consistent with the results obtained in mice, inhibition of Akt-1 promoted longevity in nematodes (Caenorhabditis elegans), whereas activation of Akt-1 shortened the lifespan. Inhibition of Akt-1 led to a decrease of ribosomal gene expression and the mitochondrial DNA content in both human cells and nematodes. Moreover, deletion of ribosomal gene expression resulted in a decrease of the mitochondrial DNA content and normalized the lifespan shortened by Akt-1 activation in nematodes. These results suggest that an increase of mitochondrial amount and energy expenditure associated with enhanced protein synthesis accelerates both aging and the onset of age-associated diseases.     

Cell cycle regulation during development and dormancy in embryos of the annual killifish Austrofundulus limnaeus

Embryos of the annual killifish Austrofundulus limnaeus can enter into a state of metabolic dormancy, termed diapause, as a normal part of their development. In addition, these embryos can also survive for prolonged sojourns in the complete absence of oxygen. Dormant embryos support their metabolism using anaerobic metabolic pathways, regardless of oxygen availability. Dormancy in diapause is associated with high ATP and a positive cellular energy status, while anoxia causes a severe reduction in ATP content and large reductions in adenylate energy charge and ATP/ADP ratios. Most cells are arrested in the G₁/G₀ phase of the cell cycle during diapause and in response to oxygen deprivation. In this paper, we review what is known about the physiological and biochemical mechanisms that support metabolic dormancy in this species. We also highlight the great potential that this model holds for identifying novel therapies for human diseases such as heart attack, stroke and cancer.     

Methionine supplementation stimulates mitochondrial respiration

Mitochondria play essential metabolic functions in eukaryotes. Although their major role is the generation of energy in the form of ATP, they are also involved in maintenance of cellular redox state, conversion and biosynthesis of metabolites and signal transduction. Most mitochondrial functions are conserved in eukaryotic systems and mitochondrial dysfunctions trigger several human diseases.By using multi-omics approach, we investigate the effect of methionine supplementation on yeast cellular metabolism, considering its role in the regulation of key cellular processes. Methionine supplementation induces an up-regulation of proteins related to mitochondrial functions such as TCA cycle, electron transport chain and respiration, combined with an enhancement of mitochondrial pyruvate uptake and TCA cycle activity. This metabolic signature is more noticeable in cells lacking Snf1/AMPK, the conserved signalling regulator of energy homeostasis. Remarkably, snf1Δ cells strongly depend on mitochondrial respiration and suppression of pyruvate transport is detrimental for this mutant in methionine condition, indicating that respiration mostly relies on pyruvate flux into mitochondrial pathways.These data provide new insights into the regulation of mitochondrial metabolism and extends our understanding on the role of methionine in regulating energy signalling pathways.     

RNA-interference in the nematode Caenorhabditis elegans is effective using paraformaldehyde-inactivated E. coli HT115 bacteria as a food source

The nematode Caenorhabditis elegans is a widely used research model for the investigation of metabolism, aging and age-associated diseases. However, when investigating the impact of natural compounds or drugs on those topics, a major confounder is the metabolism of these test substances by live E. coli bacteria, the standard food source of C. elegans. Using paraformaldehyde instead of heat to inactivate E. coli, which allows for high-throughput technologies and better food availability, it is shown here that RNA-interference works equally well, thus demonstrating the absence of considerable interfering modifications of paraformaldehyde with nucleic acids.     

Pyruvate imbalance mediates metabolic reprogramming and mimics lifespan extension by dietary restriction in Caenorhabditis elegans

Dietary restriction (DR) is the most universal intervention known to extend animal lifespan. DR also prevents tumor development in mammals, and this effect requires the tumor suppressor PTEN. However, the metabolic and cellular processes that underly the beneficial effects of DR are poorly understood. We identified slcf‐1 in an RNAi screen for genes that extend Caenorhabditis elegans lifespan in a PTEN/daf‐18‐dependent manner. We showed that slcf‐1 mutation, which increases average lifespan by 40%, mimics DR in worms fed ad libitum. An NMR‐based metabolomic characterization of slcf‐1 mutants revealed lower lipid levels compared to wild‐type animals, as expected for dietary‐restricted animals, but also higher pyruvate content. Epistasis experiments and metabolic measurements support a model in which the long lifespan of slcf‐1 mutants relies on increased mitochondrial pyruvate metabolism coupled to an adaptive response to oxidative stress. This response requires DAF‐18/PTEN and the previously identified DR effectors PHA‐4/FOXA, HSF‐1/HSF1, SIR‐2.1/SIRT‐1, and AMPK/AAK‐2. Overall, our data show that pyruvate homeostasis plays a central role in lifespan control in C. elegans and that the beneficial effects of DR results from a hormetic mechanism involving the mitochondria. Analysis of the SLCF‐1 protein sequence predicts that slcf‐1 encodes a plasma membrane transporter belonging to the conserved monocarboxylate transporter family. These findings suggest that inhibition of this transporter homolog in mammals might also promote a DR response.