Recent preclinical studies showed the potential of nicotinamide adenine dinucleotide : NAD+ : precursors to increase oxidative phosphorylation and improve metabolic health, but human data is lacking. Here, we hypothesized that the nicotinic acid derivative Acipimox, a NAD(+) precursor, would directly affect mitochondrial function, independent of reductions in non-esterified fatty acid (NEFA) concentrations. In a multi-center randomized cross-over trial, 21 patients with type 2 diabetes (age 57.7±1.1 years, BMI, 33.4±0.8 kg/m(2)) received either placebo or 250 mg Acipimox thrice daily for 2 weeks. Acipimox treatment increased plasma NEFA (759±44 vs. 1135±97 µmol/L, p<0.01 for placebo vs. Acipimox), due to a previously described rebound effect. As a result, skeletal muscle lipid content increased and insulin sensitivity decreased. Despite the elevated plasma NEFA levels, ex vivo mitochondrial respiration in skeletal muscle increased. Subsequently, we showed that Acipimox treatment resulted in a robust elevation in expression of nuclear-encoded mitochondrial gene-sets and by presence of a mitonuclear protein imbalance, which may indicate activation of the mitochondrial unfolded protein response (UPR(mt)). Further studies in C2C12 myotubes confirmed a direct effect of Acipimox on NAD+ levels, mitonuclear protein imbalance and mitochondrial oxidative capacity. To the best of our knowledge, this is the first demonstration that NAD+ boosters can also directly impact skeletal muscle mitochondrial function in humans.
The BXD genetic reference population is a recombinant inbred panel descended from crosses between the C57BL/6 (B6) and DBA/2 (D2) strains of mice, which segregate for about 5 million sequence variants. Recently, some of these variants have been established with effects on general metabolic phenotypes such as glucose response and bone strength. Here we phenotype 43 BXD strains and observe they have large variation (-5-fold) in their spontaneous activity during waking hours. QTL analyses indicate that -40% of this variance is attributable to a narrow locus containing the aryl hydrocarbon receptor (Ahr), a basic helix-loop-helix transcription factor with well-established roles in development and xenobiotic metabolism. Strains with the D2 allele of Ahr have reduced gene expression compared to those with the B6 allele, and have significantly higher spontaneous activity. This effect was also observed in B6 mice with a congenic D2 Ahr interval, and in B6 mice with a humanized AHR allele which, like the D2 allele, is expressed much less and has less enzymatic activity than the B6 allele. Ahr is highly conserved in invertebrates, and strikingly inhibition of its orthologs in D. melanogaster and C. elegans (spineless and ahr-1) leads to marked increases in basal activity. In mammals, Ahr has numerous ligands, but most are either non-selective (e.g. resveratrol) or highly toxic (e.g., 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD)). Thus, we chose to examine a major environmental influence--long term feeding with high fat diet (HFD)--to see if the effects of Ahr are dependent on major metabolic differences. Interestingly, while HFD robustly halved movement across all strains, the QTL position and effects of Ahr remained unchanged, indicating that the effects are independent. The highly consistent effects of Ahr on movement indicate that changes in its constitutive activity have a role on spontaneous movement and may influence human behavior.
The ability to respond to various intracellular and/or extracellular stresses allows the organism to adapt to changing environmental conditions and drives evolution. It is now well accepted that a progressive decline of the efficiency of stress response pathways occurs with aging. In this context, a correct proteostasis is essential for the functionality of the cell, and its dysfunction has been associated with protein aggregation and age-related degenerative diseases. Complex response mechanisms have evolved to deal with unfolded protein stress in different subcellular compartments and their moderate activation translates into positive effects on health. In this review, we focus on the mitochondrial unfolded protein response (UPR(mt)), a response to proteotoxic stress specifically in mitochondria, an organelle with a wide array of fundamental functions, most notably the harvesting of energy from food and the control of cell death. We compare UPR(mt) with the extensively characterized cytosolic heat shock response (HSR) and the unfolded protein response in endoplasmic reticulum (UPR(ER)), and discuss the current knowledge about UPR(mt) signaling pathways as well as their potential involvement in physiology.
Immune tolerance breakdown during UC involves the peroxisome proliferator-activated receptor-? (PPAR?), a key factor in mucosal homoeostasis and the therapeutic target of 5-aminosalycilates, which expression is impaired during UC. Here we assess the impact of glucocorticoids (GCs) on PPAR? expression, focusing especially on extra-adrenal cortisol production by colonic epithelial cells (CECs).
No therapy for non-alcoholic steatohepatitis (NASH) has been approved so far. Roux-en-y gastric bypass (RYGB) is emerging as a therapeutic option, although its effect on NASH and related hepatic molecular pathways is unclear from human studies. We studied the effect of RYGB on pre-existent NASH and hepatic mitochondrial dysfunction-a key player in NASH pathogenesis-in a novel diet-induced mouse model nicely mimicking human disease.
The manner by which genotype and environment affect complex phenotypes is one of the fundamental questions in biology. In this study, we quantified the transcriptome--a subset of the metabolome--and, using targeted proteomics, quantified a subset of the liver proteome from 40 strains of the BXD mouse genetic reference population on two diverse diets. We discovered dozens of transcript, protein, and metabolite QTLs, several of which linked to metabolic phenotypes. Most prominently, Dhtkd1 was identified as a primary regulator of 2-aminoadipate, explaining variance in fasted glucose and diabetes status in both mice and humans. These integrated molecular profiles also allowed further characterization of complex pathways, particularly the mitochondrial unfolded protein response (UPR(mt)). UPR(mt) shows strikingly variant responses at the transcript and protein level that are remarkably conserved among C. elegans, mice, and humans. Overall, these examples demonstrate the value of an integrated multilayered omics approach to characterize complex metabolic phenotypes.
Macrophages are important for maintaining intestinal immune homeostasis. Here, we show that PPAR?/? (peroxisome proliferator-activated receptor ?/?) directly regulates CD300a in macrophages that express the immunoreceptor tyrosine based-inhibitory motif (ITIM)-containing receptor. In mice lacking CD300a, high-fat diet (HFD) causes chronic intestinal inflammation with low numbers of intestinal lymph capillaries and dramatically expanded mesenteric lymph nodes. As a result, these mice exhibit triglyceride malabsorption and reduced body weight gain on HFD. Peritoneal macrophages from Cd300a-/- mice on HFD are classically M1 activated. Activation of toll-like receptor 4 (TLR4)/MyD88 signaling by lipopolysaccharide (LPS) results in prolonged IL-6 secretion in Cd300a-/- macrophages. Bone marrow transplantation confirmed that the phenotype originates from CD300a deficiency in leucocytes. These results identify CD300a-mediated inhibitory signaling in macrophages as a critical regulator of intestinal immune homeostasis.
Amyotrophic lateral sclerosis (ALS) is a fatal, neurodegenerative disease that causes death of motor neurons. ALS patients and mouse models of familial ALS display organismal level metabolic dysfunction, which includes increased energy expenditure despite decreased lean mass. The pathophysiological relevance of abnormal energy homeostasis to motor neuron disease remains unclear. Leptin is an adipocyte-derived hormone that regulates whole-animal energy expenditure. Here, we report that placing mutant superoxide dismutase 1 (SOD1) mice in a leptin-deficient background improves energy homeostasis and slows disease progression. Leptin-deficient mutant SOD1 mice possess increased bodyweight and fat mass, as well as decreased energy expenditure. These observations coincide with enhanced survival, improved strength and decreased motor neuron loss. These results suggest that altering whole-body energy metabolism in mutant SOD1 mice can mitigate disease progression. We propose that manipulations that increase fat mass and reduce energy expenditure will be beneficial in the setting of motor neuron disease.
Mitochondria are semi-autonomous organelles regulated by a complex network of proteins that are vital for many cellular functions. Because mitochondrial modulators can impact many aspects of cellular homeostasis, their identification and validation has proven challenging. It requires the measurement of multiple parameters in parallel to understand the exact nature of the changes induced by such compounds. We developed a platform of assays scoring for mitochondrial function in two complementary models systems, mammalian cells and C. elegans. We first optimized cell culture conditions and established the mitochondrial signature of 1,200 FDA-approved drugs in liver cells. Using cell-based and C. elegans assays, we further defined the metabolic effects of two pharmacological classes that emerged from our hit list, i.e. imidazoles and statins. We found that these two drug classes affect respiration through different and cholesterol-independent mechanisms in both models. Our screening strategy enabled us to unequivocally identify compounds that have toxic or beneficial effects on mitochondrial activity. Furthermore, the cross-species approach provided novel mechanistic insight and allowed early validation of hits that act on mitochondrial function.
Mitochondrial activity is controlled by proteins encoded by both nuclear and mitochondrial DNA. Here, we identify Sirt7 as a crucial regulator of mitochondrial homeostasis. Sirt7 deficiency in mice induces multisystemic mitochondrial dysfunction, which is reflected by increased blood lactate levels, reduced exercise performance, cardiac dysfunction, hepatic microvesicular steatosis, and age-related hearing loss. This link between SIRT7 and mitochondrial function is translatable in humans, where SIRT7 overexpression rescues the mitochondrial functional defect in fibroblasts with a mutation in NDUFSI. These wide-ranging effects of SIRT7 on mitochondrial homeostasis are the consequence of the deacetylation of distinct lysine residues located in the hetero- and homodimerization domains of GABP?1, a master regulator of nuclear-encoded mitochondrial genes. SIRT7-mediated deacetylation of GABP?1 facilitates complex formation with GABP? and the transcriptional activation of the GABP?/GABP? heterotetramer. Altogether, these data suggest that SIRT7 is a dynamic nuclear regulator of mitochondrial function through its impact on GABP?1 function.
Nutrient availability is the major regulator of life and reproduction, and a complex cellular signaling network has evolved to adapt organisms to fasting. These sensor pathways monitor cellular energy metabolism, especially mitochondrial ATP production and NAD(+)/NADH ratio, as major signals for nutritional state. We hypothesized that these signals would be modified by mitochondrial respiratory chain disease, because of inefficient NADH utilization and ATP production. Oral administration of nicotinamide riboside (NR), a vitamin B3 and NAD(+) precursor, was previously shown to boost NAD(+) levels in mice and to induce mitochondrial biogenesis. Here, we treated mitochondrial myopathy mice with NR. This vitamin effectively delayed early- and late-stage disease progression, by robustly inducing mitochondrial biogenesis in skeletal muscle and brown adipose tissue, preventing mitochondrial ultrastructure abnormalities and mtDNA deletion formation. NR further stimulated mitochondrial unfolded protein response, suggesting its protective role in mitochondrial disease. These results indicate that NR and strategies boosting NAD(+) levels are a promising treatment strategy for mitochondrial myopathy.
The deacetylase sirtuin 1 (Sirt1) exerts beneficial effects on lipid metabolism, but its roles in plasma LDL-cholesterol regulation and atherosclerosis are controversial. Thus, we applied the pharmacological Sirt1 activator SRT3025 in a mouse model of atherosclerosis and in hepatocyte culture.
Mitochondria, the main site of cellular energy harvesting, are derived from proteobacteria that evolved within our cells in endosymbiosis. Mitochondria retained vestiges of their proteobacterial genome, the circular mitochondrial DNA, which encodes 13 subunits of the oxidative phosphorylation multiprotein complexes in the electron transport chain (ETC), while the remaining ~80 ETC components are encoded in the nuclear DNA (nDNA). A further ~1,400 proteins, which are essential for mitochondrial function are also encoded in nDNA. Thus, a majority of mitochondrial proteins are translated in the cytoplasm, then imported, processed, and assembled in the mitochondria. An intricate protein quality control (PQC) network, constituted of chaperones and proteases that refold or degrade defective proteins, maintains mitochondrial proteostasis and ensures the cell and organism health. The mitochondrial unfolded protein response is a relatively recently discovered PQC pathway, which senses the proteostatic disturbances specifically in the mitochondria and resolves the stress by retrograde signaling to the nucleus and consequent transcriptional activation of protective genes. This PQC system does not only transiently resolve the local stress but also can have long-lasting effects on whole body metabolism, fitness, and longevity. A delicate tuning of its activation levels might constitute a treatment of various diseases, such as metabolic diseases, cancer, and neurodegenerative disorders.
Metabolic homeostasis requires that cellular energy levels are adapted to environmental cues. This adaptation is largely regulated at the transcriptional level, through the interaction between transcription factors, coregulators, and the basal transcriptional machinery. Coregulators, which function as both metabolic sensors and transcriptional effectors, are ideally positioned to synchronize metabolic pathways to environmental stimuli. The balance between inhibitory actions of corepressors and stimulatory effects of coactivators enables the fine-tuning of metabolic processes. This tight regulation opens therapeutic opportunities to manage metabolic dysfunction by directing the activity of cofactors toward specific transcription factors, pathways, or cells/tissues, thereby restoring whole-body metabolic homeostasis.
We previously demonstrated that the deletion of the poly(ADP-ribose)polymerase (Parp)-1 gene in mice enhances oxidative metabolism, thereby protecting against diet-induced obesity. However, the therapeutic use of PARP inhibitors to enhance mitochondrial function remains to be explored. Here, we show tight negative correlation between Parp-1 expression and energy expenditure in heterogeneous mouse populations, indicating that variations in PARP-1 activity have an impact on metabolic homeostasis. Notably, these genetic correlations can be translated into pharmacological applications. Long-term treatment with PARP inhibitors enhances fitness in mice by increasing the abundance of mitochondrial respiratory complexes and boosting mitochondrial respiratory capacity. Furthermore, PARP inhibitors reverse mitochondrial defects in primary myotubes of obese humans and attenuate genetic defects of mitochondrial metabolism in human fibroblasts and C. elegans. Overall, our work validates in worm, mouse, and human models that PARP inhibition may be used to treat both genetic and acquired muscle dysfunction linked to defective mitochondrial function.
Mitochondrial disorders are highly heterogeneous conditions characterized by defects of the mitochondrial respiratory chain. Pharmacological activation of mitochondrial biogenesis has been proposed as an effective means to correct the biochemical defects and ameliorate the clinical phenotype in these severely disabling, often fatal, disorders. Pathways related to mitochondrial biogenesis are targets of Sirtuin1, a NAD(+)-dependent protein deacetylase. As NAD(+) boosts the activity of Sirtuin1 and other sirtuins, intracellular levels of NAD(+) play a key role in the homeostatic control of mitochondrial function by the metabolic status of the cell. We show here that supplementation with nicotinamide riboside, a natural NAD(+) precursor, or reduction of NAD(+) consumption by inhibiting the poly(ADP-ribose) polymerases, leads to marked improvement of the respiratory chain defect and exercise intolerance of the Sco2 knockout/knockin mouse, a mitochondrial disease model characterized by impaired cytochrome c oxidase biogenesis. This strategy is potentially translatable into therapy of mitochondrial disorders in humans.
Nonalcoholic fatty liver disease (NAFLD) affects up to 30% of the adult population in Western societies, yet the underlying molecular pathways remain poorly understood. Here, we identify the dimeric Activator Protein 1 as a regulator of NAFLD. Fos-related antigen 1 (Fra-1) and Fos-related antigen 2 (Fra-2) prevent dietary NAFLD by inhibiting prosteatotic PPAR? signaling. Moreover, established NAFLD and the associated liver damage can be efficiently reversed by hepatocyte-specific Fra-1 expression. In contrast, c-Fos promotes PPAR? expression, while c-Jun exerts opposing, dimer-dependent functions. Interestingly, JunD was found to be essential for PPAR? signaling and NAFLD development. This unique antagonistic regulation of PPAR? by distinct AP-1 dimers occurs at the transcriptional level and establishes AP-1 as a link between obesity, hepatic lipid metabolism, and NAFLD.
SIRT2 belongs to a highly conserved family of NAD+-dependent deacylases, consisting of seven members (SIRT1-SIRT7), which vary in subcellular localizations and have substrates ranging from histones to transcription factors and enzymes. Recently SIRT2 was revealed to play an important role in inflammation, directly binding, deacetylating, and inhibiting the p65 subunit of NF-?B.
Dysfunction of Paneth and goblet cells in the intestine contributes to inflammatory bowel disease (IBD) and colitis-associated colorectal cancer (CAC). Here, we report a role for the NAD+-dependent histone deacetylase SIRT1 in the control of anti-bacterial defense. Mice with an intestinal specific Sirt1 deficiency (Sirt1int-/-) have more Paneth and goblet cells with a consequent rearrangement of the gut microbiota. From a mechanistic point of view, the effects on mouse intestinal cell maturation are mediated by SIRT1-dependent changes in the acetylation status of SPDEF, a master regulator of Paneth and goblet cells. Our results suggest that targeting SIRT1 may be of interest in the management of IBD and CAC.
The estrogen-related receptor-? (ERR?) regulates mitochondrial biogenesis and glucose and fatty acid oxidation during differentiation in skeletal myocytes. However, whether ERR? controls metabolic remodeling during skeletal muscle regeneration in vivo is unknown. We characterized the time course of skeletal muscle regeneration in wild-type (M-ERR?WT) and muscle-specific ERR?(-/-) (M-ERR?(-/-)) mice after injury by intramuscular cardiotoxin injection. M-ERR?(-/-) mice exhibited impaired regeneration characterized by smaller myofibers with increased centrally localized nuclei and reduced mitochondrial density and cytochrome oxidase and citrate synthase activities relative to M-ERR?WT. Transcript levels of mitochondrial transcription factor A, nuclear respiratory factor-2a, and peroxisome proliferator-activated receptor (PPAR)-? coactivator (PGC)-1?, were downregulated in the M-ERR?(-/-) muscles at the onset of myogenesis. Furthermore, coincident with delayed myofiber recovery, we observed reduced muscle ATP content (-45% vs. M-ERR?WT) and enhanced AMP-activated protein kinase (AMPK) activation in M-ERR?(-/-) muscle. We subsequently demonstrated that pharmacologic postinjury AMPK activation was sufficient to delay muscle regeneration in WT mice. AMPK activation induced ERR? transcript expression in M-ERR?WT muscle and in C2C12 myotubes through induction of the Esrra promoter, indicating that ERR? may control gene regulation downstream of the AMPK pathway. Collectively, these results suggest that ERR? deficiency during muscle regeneration impairs recovery of mitochondrial energetic capacity and perturbs AMPK activity, resulting in delayed myofiber repair.-LaBarge, S., McDonald, M., Smith-Powell, L., Auwerx, J., Huss, J. M. Estrogen-related receptor-? (ERR?) deficiency in skeletal muscle impairs regeneration in response to injury.
The vitamin D endocrine system has many extraskeletal targets, including adipose tissue. 1,25-Dihydroxyvitamin D3, the active form of vitamin D, not only increases adipogenesis and the expression of typical adipocyte genes but also decreases the expression of uncoupling proteins. Mice with disrupted vitamin D action-owing to gene deletion of the nuclear receptor vitamin D receptor (Vdr) or the gene encoding 1?-hydroxylase (Cyp27b1)-lose fat mass over time owing to an increase in energy expenditure, whereas mice with increased Vdr-mediated signalling in adipose tissue become obese. The resistance to diet-induced obesity in mice with disrupted Vdr signalling is caused at least partially by increased expression of uncoupling proteins in white adipose tissue. However, the bile acid pool is also increased in these animals, and bile acids are known to be potent inducers of energy expenditure through activation of several nuclear receptors, including Vdr, and G-protein-coupled receptors, such as GPBAR1 (also known as TGR5). By contrast, in humans, obesity is strongly associated with poor vitamin D status. A causal link has not been firmly proven, but most intervention studies have failed to demonstrate a beneficial effect of vitamin D supplementation on body weight. The reasons for the major discrepancy between mouse and human data are unclear, but understanding the link between vitamin D status and energy homeostasis could potentially be very important for the human epidemic of obesity and the metabolic syndrome.
Copper is an essential trace element, the imbalances of which are associated with various pathological conditions, including cancer, albeit via largely undefined molecular and cellular mechanisms. Here we provide evidence that levels of bioavailable copper modulate tumor growth. Chronic exposure to elevated levels of copper in drinking water, corresponding to the maximum allowed in public water supplies, stimulated proliferation of cancer cells and de novo pancreatic tumor growth in mice. Conversely, reducing systemic copper levels with a chelating drug, clinically used to treat copper disorders, impaired both. Under such copper limitation, tumors displayed decreased activity of the copper-binding mitochondrial enzyme cytochrome c oxidase and reduced ATP levels, despite enhanced glycolysis, which was not accompanied by increased invasiveness of tumors. The antiproliferative effect of copper chelation was enhanced when combined with inhibitors of glycolysis. Interestingly, larger tumors contained less copper than smaller tumors and exhibited comparatively lower activity of cytochrome c oxidase and increased glucose uptake. These results establish copper as a tumor promoter and reveal that varying levels of copper serves to regulate oxidative phosphorylation in rapidly proliferating cancer cells inside solid tumors. Thus, activation of glycolysis in tumors may in part reflect insufficient copper bioavailability in the tumor microenvironment.
Apolipoprotein E (ApoE) is found in three different forms in humans (ApoE2, ApoE3 and ApoE4), and ApoE polymorphism is recognized as a major risk factor for Alzheimers disease (AD). ApoE is involved in lipid and cholesterol transport, cell repair, and amyloid-? deposition and certain studies suggest potential implications in neurogenesis. In this regard, we investigated the possible impact of the three different human ApoE isoforms on neurogenesis. We used ApoE knock-in mice of different ages and sex, and quantified newborn cells in the hippocampus by flow cytometry. Young adult ApoE4 mice (10-12 week-old) from both sexes displayed reduced neurogenesis compared with wild-types and the other genotypes. In addition, young adult ApoE2 female mice showed improved hippocampal progenitor cell proliferation. In older mice (1 year), hippocampal neurogenesis was globally decreased, particularly in females, and the difference between ApoE4 and the other genotypes observed in young animals disappeared for the two sexes, except for aged ApoE3 females. Indeed, a surprising protective effect of the ApoE3 genotype was observed in aged females. Our study highlights the role of ApoE in neurogenesis, and shows for the first time an early inequality between the ApoE genotypes. The reduced neurogenesis observed for the ApoE4 genotype and the improved results obtained in young ApoE2 females support the idea of a difference in the balance between neuronal birth and death modulated by the ApoE polymorphism in young animals. The maintenance of this balance and its modulation can influence pathophysiological mechanisms predisposing to neurodegenerative diseases like AD.
Bile reflux contributes to development of Barretts esophagus (BE) and could be involved in its progression to esophageal adenocarcinoma (EAC). We investigated whether bile acids affect levels or functions of microRNAs (MIRs) 221 and 222, which bind to the 3-UTR of p27Kip1 messenger RNA to inhibit its translation. Reduced p27Kip1 increases degradation of the transcription factor CDX2; levels of CDX2 have been reported to decrease during progression of BE to EAC.
Altered adipose tissue formation is a well-known effectors of obesity and T2D. Here, we describe the role of Lrh1 and its co-repressor Shp in the control of adipocyte formation. Expression of Lrh1 in the pre-adipocyte containing SVF is induced in obese mice models and humans while Shp expression is reduced. We demonstrate, that Lrh1 is an inhibitor of adipogenesis while Shp acts functions as an activator through repression of Lrh1 activity. This regulation is at least in part modulated by estradiol conversion through the regulation of Cyp19a1 gene expression. In vivo, loss of Lrh1 leads to induced adipogenesis, while loss of Shp causes uncontrolled activation of Lrh1 and reduced adipogenesis. As Shp expression has been linked to the development of obesity and metabolic disorders, it is possible that alterations of the Shp/Lrh1 network lead to changes in adipocyte formation, which might contribute to the development of obesity associated T2D.
Macrophage-mediated inflammation is a major contributor to obesity-associated insulin resistance. The corepressor NCoR interacts with inflammatory pathway genes in macrophages, suggesting that its removal would result in increased activity of inflammatory responses. Surprisingly, we find that macrophage-specific deletion of NCoR instead results in an anti-inflammatory phenotype along with robust systemic insulin sensitization in obese mice. We present evidence that derepression of LXRs contributes to this paradoxical anti-inflammatory phenotype by causing increased expression of genes that direct biosynthesis of palmitoleic acid and ?3 fatty acids. Remarkably, the increased ?3 fatty acid levels primarily inhibit NF-?B-dependent inflammatory responses by uncoupling NF-?B binding and enhancer/promoter histone acetylation from subsequent steps required for proinflammatory gene activation. This provides a mechanism for the in vivo anti-inflammatory insulin-sensitive phenotype observed in mice with macrophage-specific deletion of NCoR. Therapeutic methods to harness this mechanism could lead to a new approach to insulin-sensitizing therapies.
Nicotinamide adenine dinucleotide (NAD) is a central metabolic cofactor by virtue of its redox capacity, and as such regulates a wealth of metabolic transformations. However, the identification of the longevity protein silent regulator 2 (Sir2), the founding member of the sirtuin protein family, as being NAD?-dependent reignited interest in this metabolite. The sirtuins (SIRT1-7 in mammals) utilize NAD? to deacetylate proteins in different subcellular compartments with a variety of functions, but with a strong convergence on optimizing mitochondrial function. Since cellular NAD? levels are limiting for sirtuin activity, boosting its levels is a powerful means to activate sirtuins as a potential therapy for mitochondrial, often age-related, diseases. Indeed, supplying excess precursors, or blocking its utilization by poly(ADP-ribose) polymerase (PARP) enzymes or CD38/CD157, boosts NAD? levels, activates sirtuins and promotes healthy aging. Here, we discuss the current state of knowledge of NAD? metabolism, primarily in relation to sirtuin function. We highlight how NAD? levels change in diverse physiological conditions, and how this can be employed as a pharmacological strategy.
Sirt5, localized in the mitochondria, is a member of sirtuin family of NAD?-dependent deacetylases. Sirt5 was shown to deacetylate and activate carbamoyl phosphate synthase 1. Most recently, Sirt5 was reported to be the predominant protein desuccinylase and demalonylase in the mitochondria because the ablation of Sirt5 enhanced the global succinylation and malonylation of mitochondrial proteins, including many metabolic enzymes. In order to determine the physiological role of Sirt5 in metabolic homeostasis, we generated a germline Sirt5 deficient (Sirt5?/?) mouse model and performed a thorough metabolic characterization of this mouse line. Although a global protein hypersuccinylation and elevated serum ammonia during fasting were observed in our Sirt5?/? mouse model, Sirt5 deficiency did not lead to any overt metabolic abnormalities under either chow or high fat diet conditions. These observations suggest that Sirt5 is likely to be dispensable for the metabolic homeostasis under the basal conditions.
Sirt3 is a mitochondrial NAD(+)-dependent deacetylase that governs mitochondrial metabolism and reactive oxygen species homeostasis. Sirt3 deficiency has been reported to accelerate the development of the metabolic syndrome. However, the role of Sirt3 in atherosclerosis remains enigmatic. We aimed to investigate whether Sirt3 deficiency affects atherosclerosis, plaque vulnerability, and metabolic homeostasis. Low-density lipoprotein receptor knockout (LDLR (-/-)) and LDLR/Sirt3 double-knockout (Sirt3 (-/-) LDLR (-/-)) mice were fed a high-cholesterol diet (1.25 % w/w) for 12 weeks. Atherosclerosis was assessed en face in thoraco-abdominal aortae and in cross sections of aortic roots. Sirt3 deletion led to hepatic mitochondrial protein hyperacetylation. Unexpectedly, though plasma malondialdehyde levels were elevated in Sirt3-deficient mice, Sirt3 deletion affected neither plaque burden nor features of plaque vulnerability (i.e., fibrous cap thickness and necrotic core diameter). Likewise, plaque macrophage and T cell infiltration as well as endothelial activation remained unaltered. Electron microscopy of aortic walls revealed no difference in mitochondrial microarchitecture between both groups. Interestingly, loss of Sirt3 was associated with accelerated weight gain and an impaired capacity to cope with rapid changes in nutrient supply as assessed by indirect calorimetry. Serum lipid levels and glucose tolerance were unaffected by Sirt3 deletion in LDLR (-/-) mice. Sirt3 deficiency does not affect atherosclerosis in LDLR (-/-) mice. However, Sirt3 controls systemic levels of oxidative stress, limits expedited weight gain, and allows rapid metabolic adaptation. Thus, Sirt3 may contribute to postponing cardiovascular risk factor development.
Mitochondrial dysfunction is not only a hallmark of rare inherited mitochondrial disorders but also implicated in age-related diseases, including those that affect the metabolic and nervous system, such as type 2 diabetes and Parkinsons disease. Numerous pathways maintain and/or restore proper mitochondrial function, including mitochondrial biogenesis, mitochondrial dynamics, mitophagy and the mitochondrial unfolded protein response. New and powerful phenotypic assays in cell-based models as well as multicellular organisms have been developed to explore these different aspects of mitochondrial function. Modulating mitochondrial function has therefore emerged as an attractive therapeutic strategy for several diseases, which has spurred active drug discovery efforts in this area.
Epigenetic regulation of gene expression is strongly influenced by the accessibility of nucleosomal DNA or the state of chromatin compaction. In this context, coregulators, including both coactivators and corepressors, are pivotal intermediates that bridge chromatin-modifying enzymes and transcription factors. NCoR1 (nuclear receptor corepressor) and SMRT (silencing mediator of retinoic acid and thyroid hormone receptor) are among the best-characterized corepressors from a molecular point of view. These coregulators have conserved orthologs in lower organisms, which underscores their functional importance. Here we summarize the results from recent in vivo studies that reveal the wide-ranging roles of NCoR1 and SMRT in developmental as well as homeostatic processes, including metabolism, inflammation, and circadian rhythms. We also discuss the potential implications of NCoR1 and SMRT regulation of pathways ranging from genomic stability and carcinogenesis to metabolic diseases such as type 2 diabetes.
NAD(+) is an important cofactor regulating metabolic homeostasis and a rate-limiting substrate for sirtuin deacylases. We show that NAD(+) levels are reduced in aged mice and Caenorhabditis elegans and that decreasing NAD(+) levels results in a further reduction in worm lifespan. Conversely, genetic or pharmacological restoration of NAD(+) prevents age-associated metabolic decline and promotes longevity in worms. These effects are dependent upon the protein deacetylase sir-2.1 and involve the induction of mitonuclear protein imbalance as well as activation of stress signaling via the mitochondrial unfolded protein response (UPR(mt)) and the nuclear translocation and activation of FOXO transcription factor DAF-16. Our data suggest that augmenting mitochondrial stress signaling through the modulation of NAD(+) levels may be a target to improve mitochondrial function and prevent or treat age-associated decline.
Longevity is regulated by a network of closely linked metabolic systems. We used a combination of mouse population genetics and RNA interference in Caenorhabditis elegans to identify mitochondrial ribosomal protein S5 (Mrps5) and other mitochondrial ribosomal proteins as metabolic and longevity regulators. MRP knockdown triggers mitonuclear protein imbalance, reducing mitochondrial respiration and activating the mitochondrial unfolded protein response. Specific antibiotics targeting mitochondrial translation and ethidium bromide (which impairs mitochondrial DNA transcription) pharmacologically mimic mrp knockdown and extend worm lifespan by inducing mitonuclear protein imbalance, a stoichiometric imbalance between nuclear and mitochondrially encoded proteins. This mechanism was also conserved in mammalian cells. In addition, resveratrol and rapamycin, longevity compounds acting on different molecular targets, similarly induced mitonuclear protein imbalance, the mitochondrial unfolded protein response and lifespan extension in C. elegans. Collectively these data demonstrate that MRPs represent an evolutionarily conserved protein family that ties the mitochondrial ribosome and mitonuclear protein imbalance to the mitochondrial unfolded protein response, an overarching longevity pathway across many species.
For reasons that remain unclear, whether type 5 adenylyl cyclase (AC5), 1 of 2 major AC isoforms in heart, is protective or deleterious in response to cardiac stress is controversial. To reconcile this controversy we examined the cardiomyopathy induced by chronic isoproterenol in AC5 transgenic (Tg) mice and the signaling mechanisms involved.
The mechanisms involved in the coordinate regulation of the metabolic and structural programs controlling muscle fitness and endurance are unknown. Recently, the nuclear receptor PPAR?/? was shown to activate muscle endurance programs in transgenic mice. In contrast, muscle-specific transgenic overexpression of the related nuclear receptor, PPAR?, results in reduced capacity for endurance exercise. We took advantage of the divergent actions of PPAR?/? and PPAR? to explore the downstream regulatory circuitry that orchestrates the programs linking muscle fiber type with energy metabolism. Our results indicate that, in addition to the well-established role in transcriptional control of muscle metabolic genes, PPAR?/? and PPAR? participate in programs that exert opposing actions upon the type I fiber program through a distinct muscle microRNA (miRNA) network, dependent on the actions of another nuclear receptor, estrogen-related receptor ? (ERR?). Gain-of-function and loss-of-function strategies in mice, together with assessment of muscle biopsies from humans, demonstrated that type I muscle fiber proportion is increased via the stimulatory actions of ERR? on the expression of miR-499 and miR-208b. This nuclear receptor/miRNA regulatory circuit shows promise for the identification of therapeutic targets aimed at maintaining muscle fitness in a variety of chronic disease states, such as obesity, skeletal myopathies, and heart failure.
We identified a mutation in the Diet1 gene in a mouse strain that is resistant to hyperlipidemia and atherosclerosis. Diet1 encodes a 236 kD protein consisting of tandem low-density lipoprotein receptor and MAM (meprin-A5-protein tyrosine phosphatase mu) domains and is expressed in the enterocytes of the small intestine. Diet1-deficient mice exhibited an elevated bile acid pool size and impaired feedback regulation of hepatic Cyp7a1, which encodes the rate-limiting enzyme in bile acid synthesis. In mouse intestine and in cultured human intestinal cells, Diet1 expression levels influenced the production of fibroblast growth factor 15/19 (FGF15/19), a hormone that signals from the intestine to liver to regulate Cyp7a1. Transgenic expression of Diet1, or adenoviral-mediated Fgf15 expression, restored normal Cyp7a1 regulation in Diet-1-deficient mice. Diet1 and FGF19 proteins exhibited overlapping subcellular localization in cultured intestinal cells. These results establish Diet1 as a control point in enterohepatic bile acid signaling and lipid homeostasis.
Successful mammalian reproduction depends on proper synthesis of the pituitary-derived glycoprotein hormones, luteinizing hormone (LH) and follicle-stimulating hormone (FSH). Several transcription factors cooperate to activate cell-specific and hormone-regulated expression of the gonadotropin beta subunits (Lhb and Fshb). Among these, NR5A1 (steroidogenic factor 1; SF1) has been shown to directly bind to the Lhb promoter, mediate basal and gonadotropin-releasing hormone (GnRH)-stimulated Lhb transcription, and possibly directly regulate Fshb expression. Recently, the closely-related NR5A2 was shown to activate the rat Lhb promoter in vitro. Here, we further characterized the role of NR5A2 in regulating gonadotropin synthesis. Ectopically expressed NR5A2 directly activated the murine Lhb promoter in a manner identical to that of NR5A1, whereas neither factor activated the murine Fshb promoter. In L?T2 gonadotrope-like cells, depletion of endogenous NR5A1 or NR5A2 impaired basal and GnRH-stimulated Lhb and Fshb transcription. To analyze the physiological role of NR5A2 in gonadotropes in vivo, we generated mice with a gonadotrope-specific deletion of Nr5a2. In contrast with our in vitro data, these mice had normal pituitary Lhb and Fshb expression and intact fertility. Together, our data establish that NR5A2 can act in a non-redundant manner to regulate Lhb and Fshb transcription in vitro, but is dispensable in vivo.
Peroxisome proliferator-activated receptors (PPARs) have been revealed as key regulators of several skin disorders. This has led to a growing interest in the development of drugs targeting PPARs as therapeutics for skin diseases. To evaluate skin PPAR activity, we developed peroxisome proliferator responsive element-luciferase (PPRE-Luc) mice, a mouse model in which the luciferase gene expression is under the control of a PPAR-inducible promoter in all organs. Our aim was to define and validate experimental conditions to establish PPRE-Luc mice as a valuable tool for in vivo non-invasive evaluation of PPARs activation in the skin. We demonstrated by optical imaging that topical application of 40 mm of Luciferin for 10 min was enough to reveal the optimal luciferase activity in mice skin. The treatment of mice skin with the PPAR? and PPAR? agonists, pioglitazone and WY14643, was associated with significant increase in photons emission reaching maximal signalling at 6 h. We have performed dose response studies by testing a large range of pioglitazone and WY14643 concentrations on mouse skin. The specificity of bioluminescence signal induced by pioglitazone and WY14643 was assessed using PPAR? and PPAR? antagonists, GW9662 and GW6471, respectively. This approach revealed that the isoform specificity of PPARs agonists decreased when high ligand concentrations were applied on mouse skin. These results were further confirmed by in vitro measurement of luciferase activity in skin extracts. Overall, our results demonstrated that PPRE-Luc mice represent a valuable reporter mouse model for the in vivo pharmacological profiling of drugs targeting PPARs in the skin.
Defects in axonal transport are thought to contribute to the pathogenesis of neurodegenerative disease. Because ?-tubulin acetylation facilitates axonal transport, inhibition of the ?-tubulin deacetylating enzymes, histone deacetylase 6 (Hdac6) and silent information regulator 2 (Sirt2), is thought to be an interesting therapeutic strategy for these conditions. Amyotrophic lateral sclerosis (ALS) is a one such rapidly progressive and fatal neurodegenerative disorder, in which axonal transport defects have been found in vitro and in vivo. To establish whether the inhibition of Hdac6 or Sirt2 may be of interest for ALS treatment, we investigated whether deleting Hdac6 or Sirt2 from the superoxide dismutase 1, SOD1(G93A) mouse affects the motor neuron degeneration in this ALS model. Deletion of Hdac6 significantly extended the survival of SOD1(G93A) mice without affecting disease onset, and maintained motor axon integrity. This protective effect was associated with increased ?-tubulin acetylation. Deletion of Sirt2 failed to affect the disease course, but also did not modify ?-tubulin acetylation. These findings show that Hdac6, rather than Sirt2, is a therapeutic target for the treatment of ALS. Moreover, Sirt2 appears not to be a major ?-tubulin deacetylase in the nervous system.
Macrophages act as the primary effector cells during Leishmania infection through production of reactive oxygen species (ROS) and interleukin-1? (IL-1?). However, how macrophage-killing mechanisms are activated during Leishmania-macrophage interactions is poorly understood. Here, we report that the macrophage response against Leishmania infantum in vivo is characterized by an M2b-like phenotype and C-type lectin receptors (CLRs) signature composed of Dectin-1, mannose receptor (MR), and the DC-SIGN homolog SIGNR3 expression. Dectin-1 and MR were crucial for the microbicidal response as indicated by the fact that they activated Syk-p47phox and arachidonic acid (AA)-NADPH oxidase signaling pathways, respectively, needed for ROS production and also triggered Syk-coupled signaling for caspase-1-induced IL-1? secretion. In contrast, SIGNR3 has divergent functions during Leishmania infantum pathogenesis; this CLR favored parasite resilience through inhibition of the LTB4-IL-1? axis. These pathways also operated during infection of primary human macrophages. Therefore, our study promotes CLRs as potential targets for treatment, diagnosis, and prevention of visceral leishmaniasis.
Glucose is an important metabolic substrate of the retina and diabetic patients have to maintain a strict normoglycemia to avoid diabetes secondary effects, including cardiovascular disease, nephropathy, neuropathy and retinopathy. Others and we recently demonstrated the potential role of hypoglycemia in diabetic retinopathy. We showed acute hypoglycemia to induce retinal cell death both in vivo during an hyperinsulinemic/hypoglycemic clamp and in vitro in 661W photoreceptor cells cultured at low glucose concentration. In the present study, we showed low glucose to induce a decrease of BCL2 and BCL-XL anti-apoptotic proteins expression, leading to an increase of free pro-apoptotic BAX. In parallel, we showed that, in retinal cells, low glucose-induced apoptosis is involved in the process of autophagosomes formation through the AMPK/RAPTOR/mTOR pathway. Moreover, the decrease of LAMP2a expression led to a defect in the autophagosome/lysosome fusion process. Specific inhibition of autophagy, either by 3-methyladenine or by down-regulation of ATG5 or ATG7 proteins expression, increased caspase 3 activation and 661W cell death. We show that low glucose modifies the delicate equilibrium between apoptosis and autophagy. Cells struggled against low nutrient condition-induced apoptosis by starting an autophagic process, which led to cell death when inhibited. We conclude that autophagy defect is associated with low glucose-induced 661W cells death that could play a role in diabetic retinopathy. These results could modify the way of addressing negative effects of hypoglycemia. Short-term modulation of autophagy could be envisioned to treat diabetic patients in order to avoid secondary complications of the disease.
In recent years, the role of acetylation has gained ground as an essential modulator of intermediary metabolism in skeletal muscle. Imbalance in energy homeostasis or chronic cellular stress, due to diet, aging, or disease, translate into alterations in the acetylation levels of key proteins which govern bioenergetics, cellular substrate use, and/or changes in mitochondrial content and function. For example, cellular stress induced by exercise or caloric restriction can alter the coordinated activity of acetyltransferases and deacetylases to increase mitochondrial biogenesis and function in order to adapt to low energetic levels. The natural duality of these enzymes, as metabolic sensors and effector proteins, has helped biologists to understand how the body can integrate seemingly distinct signaling pathways to control mitochondrial biogenesis, insulin sensitivity, glucose transport, reactive oxygen species handling, angiogenesis, and muscle satellite cell proliferation/differentiation. Our review will summarize the recent developments related to acetylation-dependent responses following metabolic stress in skeletal muscle.
We present a consolidated view of the complexity and challenges of designing studies for measurement of energy metabolism in mouse models, including a practical guide to the assessment of energy expenditure, energy intake and body composition and statistical analysis thereof. We hope this guide will facilitate comparisons across studies and minimize spurious interpretations of data. We recommend that division of energy expenditure data by either body weight or lean body weight and that presentation of group effects as histograms should be replaced by plotting individual data and analyzing both group and body-composition effects using analysis of covariance (ANCOVA).
Sirtuin 1 (SIRT1) is an evolutionarily conserved NAD(+)-dependent deacetylase that is at the pinnacle of metabolic control, all the way from yeast to humans. SIRT1 senses changes in intracellular NAD(+) levels, which reflect energy level, and uses this information to adapt the cellular energy output such that it matches cellular energy requirements. The changes induced by SIRT1 activation are generally (but not exclusively) transcriptional in nature and are related to an increase in mitochondrial metabolism and antioxidant protection. These attractive features have validated SIRT1 as a therapeutic target in the management of metabolic disease and prompted an intensive search to identify pharmacological SIRT1 activators. In this review, we first give an overview of the SIRT1 biology with a particular focus on its role in metabolic control. We then analyze the pros and cons of the current strategies used to activate SIRT1 and explore the emerging evidence indicating that modulation of NAD(+) levels could provide an effective way to achieve such goals.
Silent information regulator 2 (Sir2) proteins (sirtuins) are nicotinamide adenine dinucleotide-dependent deacetylases that regulate important biological processes. Mammals have seven sirtuins, Sirt1 to Sirt7. Four of them (Sirt4 to Sirt7) have no detectable or very weak deacetylase activity. We found that Sirt5 is an efficient protein lysine desuccinylase and demalonylase in vitro. The preference for succinyl and malonyl groups was explained by the presence of an arginine residue (Arg(105)) and tyrosine residue (Tyr(102)) in the acyl pocket of Sirt5. Several mammalian proteins were identified with mass spectrometry to have succinyl or malonyl lysine modifications. Deletion of Sirt5 in mice appeared to increase the level of succinylation on carbamoyl phosphate synthase 1, which is a known target of Sirt5. Thus, protein lysine succinylation may represent a posttranslational modification that can be reversed by Sirt5 in vivo.
The formation of myelin by Schwann cells (SCs) occurs via a series of orchestrated molecular events. We previously used global expression profiling to examine peripheral nerve myelination and identified the NAD(+)-dependent deacetylase Sir-two-homolog 2 (Sirt2) as a protein likely to be involved in myelination. Here, we show that Sirt2 expression in SCs is correlated with that of structural myelin components during both developmental myelination and remyelination after nerve injury. Transgenic mice lacking or overexpressing Sirt2 specifically in SCs show delays in myelin formation. In SCs, we found that Sirt2 deacetylates Par-3, a master regulator of cell polarity. The deacetylation of Par-3 by Sirt2 decreases the activity of the polarity complex signaling component aPKC, thereby regulating myelin formation. These results demonstrate that Sirt2 controls an essential polarity pathway in SCs during myelin assembly and provide insights into the association between intracellular metabolism and SC plasticity.
Aging is characterized by a general decline in cellular function, which ultimately will affect whole body homeostasis. Although DNA damage and oxidative stress all contribute to aging, metabolic dysfunction is a common hallmark of aging at least in invertebrates. Since a comprehensive overview of metabolic changes in otherwise healthy aging mammals is lacking, we here compared metabolic parameters of young and 2 year old mice. We systemically integrated in vivo phenotyping with gene expression, biochemical analysis, and metabolomics, thereby identifying a distinguishing metabolic footprint of aging. Among the affected pathways in both liver and muscle we found glucose and fatty acid metabolism, and redox homeostasis. These alterations translated in decreased long chain acylcarnitines and increased free fatty acid levels and a marked reduction in various amino acids in the plasma of aged mice. As such, these metabolites serve as biomarkers for aging and healthspan.
Dietary restriction can extend life span in most organisms tested to date, suggesting that mechanisms sensing nutrient and energy availability might regulate longevity. The AMP-activated protein kinase (AMPK) has emerged as a key energy sensor with the ability to transcriptionally reprogram the cell and metabolically adapt to external cues. In this review, we will discuss the possible role of AMPK in the beneficial effects of calorie restriction on health and life span.
Statins protect against cardiovascular-related mortality but induce skeletal muscle toxicity. To investigate mechanisms of statins, we tested the hypothesis that statins optimized cardiac mitochondrial function but impaired vulnerable skeletal muscle by inducing different level of reactive oxygen species (ROS).
Sirt1 is an NAD(+)-dependent deacetylase that extends lifespan in lower organisms and improves metabolism and delays the onset of age-related diseases in mammals. Here we show that SRT1720, a synthetic compound that was identified for its ability to activate Sirt1 in vitro, extends both mean and maximum lifespan of adult mice fed a high-fat diet. This lifespan extension is accompanied by health benefits including reduced liver steatosis, increased insulin sensitivity, enhanced locomotor activity and normalization of gene expression profiles and markers of inflammation and apoptosis, all in the absence of any observable toxicity. Using a conditional SIRT1 knockout mouse and specific gene knockdowns we show SRT1720 affects mitochondrial respiration in a Sirt1- and PGC-1?-dependent manner. These findings indicate that SRT1720 has long-term benefits and demonstrate for the first time the feasibility of designing novel molecules that are safe and effective in promoting longevity and preventing multiple age-related diseases in mammals.
Bile acids (BAs) are amphipathic molecules that facilitate the uptake of lipids, and their levels fluctuate in the intestines as well as in the circulation depending on food intake. Besides their role in dietary lipid absorption, BAs function as signaling molecules that activate specific BA receptors and trigger downstream signaling cascades. The BA receptors and the signaling pathways they control are not only important in the regulation of BA synthesis and their metabolism, but they also regulate glucose homeostasis, lipid metabolism and energy expenditure - processes relevant in the context of the metabolic syndrome. In addition to the function of the nuclear receptor FXR? in regulating local effects of BAs in the organs of the enterohepatic axis, increasing evidence points to a crucial role of the G-protein-coupled receptor TGR5 in mediating systemic actions of BAs. Here we review the current knowledge on BA receptors, with a strong focus on the cell membrane receptor TGR5, which has emerged as a promising target for intervention in metabolic diseases.
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-1? 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.
We evaluated the metabolic impact of farnesoid X receptor (FXR) activation by administering a synthetic FXR agonist (GW4064) to mice in which obesity was induced by a high fat diet. Administration of GW4064 accentuated body weight gain and glucose intolerance induced by the high fat diet and led to a pronounced worsening of the changes in liver and adipose tissue. Mechanistically, treatment with GW4064 decreased bile acid (BA) biosynthesis, BA pool size, and energy expenditure, whereas reconstitution of the BA pool in these GW4064-treated animals by BA administration dose-dependently reverted the metabolic abnormalities. Our data therefore suggest that activation of FXR with synthetic agonists is not useful for long term management of the metabolic syndrome, as it reduces the BA pool size and subsequently decreases energy expenditure, translating as weight gain and insulin resistance. In contrast, expansion of the BA pool size, which can be achieved by BA administration, could be an interesting strategy to manage the metabolic syndrome.
Poly(ADP-ribosyl) polymerases (PARPs) have traditionally been linked to chromosome maintenance and DNA repair. Recent findings identify PARPs as key modulators of metabolism through their influence on SIRT1 activity, hinting to a possible role of PARPs as longevity regulators.
Cells respond to stress by coordinating proliferative and metabolic pathways. Starvation restricts cell proliferative (glycolytic) and activates energy productive (oxidative) pathways. Conversely, cell growth and proliferation require increased glycolytic and decreased oxidative metabolism levels. E2F transcription factors regulate both proliferative and metabolic genes. E2Fs have been implicated in the G1/S cell-cycle transition, DNA repair, apoptosis, development and differentiation. In pancreatic ?-cells, E2F1 gene regulation facilitated glucose-stimulated insulin secretion. Moreover, mice lacking E2F1 (E2f1(-/-)) were resistant to diet-induced obesity. Here, we show that E2F1 coordinates cellular responses by acting as a regulatory switch between cell proliferation and metabolism. In basal conditions, E2F1 repressed key genes that regulate energy homeostasis and mitochondrial functions in muscle and brown adipose tissue. Consequently, E2f1(-/-) mice had a marked oxidative phenotype. An association between E2F1 and pRB was required for repression of genes implicated in oxidative metabolism. This repression was alleviated in a constitutively active CDK4 (CDK4(R24C)) mouse model or when adaptation to energy demand was required. Thus, E2F1 represents a metabolic switch from oxidative to glycolytic metabolism that responds to stressful conditions.
The G protein-coupled receptor TGR5 has been identified as an important component of the bile acid signaling network, and its activation has been linked to enhanced energy expenditure and improved glycemic control. Here, we demonstrate that activation of TGR5 in macrophages by 6?-ethyl-23(S)-methylcholic acid (6-EMCA, INT-777), a semisynthetic BA, inhibits proinflammatory cytokine production, an effect mediated by TGR5-induced cAMP signaling and subsequent NF-?B inhibition. TGR5 activation attenuated atherosclerosis in Ldlr(-/-)Tgr5(+/+) mice but not in Ldlr(-/-)Tgr5(-/-) double-knockout mice. The inhibition of lesion formation was associated with decreased intraplaque inflammation and less plaque macrophage content. Furthermore, Ldlr(-/-) animals transplanted with Tgr5(-/-) bone marrow did not show an inhibition of atherosclerosis by INT-777, further establishing an important role of leukocytes in INT-777-mediated inhibition of vascular lesion formation. Taken together, these data attribute a significant immune modulating function to TGR5 activation in the prevention of atherosclerosis, an important facet of the metabolic syndrome.
SIRT1 is a NAD(+)-dependent enzyme that affects metabolism by deacetylating key transcriptional regulators of energy expenditure. Here, we tested whether deletion of PARP-2, an alternative NAD(+)-consuming enzyme, impacts on NAD(+) bioavailability and SIRT1 activity. Our results indicate that PARP-2 deficiency increases SIRT1 activity in cultured myotubes. However, this increase was not due to changes in NAD(+) levels, but to an increase in SIRT1 expression, as PARP-2 acts as a direct negative regulator of the SIRT1 promoter. PARP-2 deletion in mice increases SIRT1 levels, promotes energy expenditure, and increases mitochondrial content. Furthermore, PARP-2(-/-) mice were protected against diet-induced obesity. Despite being insulin sensitized, PARP-2(-/-) mice were glucose intolerant due to a defective pancreatic function. Hence, while inhibition of PARP activity promotes oxidative metabolism through SIRT1 activation, the use of PARP inhibitors for metabolic purposes will require further understanding of the specific functions of different PARP family members.
The aim of the present work was to decipher the cellular basis of the immunoregulatory role of peroxisome proliferator-activated receptor (PPAR)-? in cutaneous hypersensitivity reactions. After challenge with a contact allergen, we observed augmented hypersensitivity reactions with increased numbers of activated T lymphocytes in the skin of PPAR-?-/- mice. Furthermore, following antigen challenge, the percentages of Tregs in the blood, draining lymph nodes and skin were decreased in these mice. PPAR-? deficiency impaired the production of IL-2 in lymph nodes, whereas TGF-? levels remained unchanged. Injection of IL-2 into PPAR-?-/- mice restored the Treg population in the skin-draining lymph nodes of allergen-challenged mice. In vivo induction of Tregs from WT CD4+ CD25- T cells was impaired when adoptively transferred into PPAR-?-/- mice as compared with transfer into WT mice, and reversed by injection of IL-2 into PPAR-?-/- mice. Furthermore, the suppressive capacity of PPAR-?-/- Tregs was impaired when compared to WT Tregs in vitro and in co-adoptive transfer experiments. Finally, injection of IL-2 to PPAR-?-/- mice decreased skin inflammation to a level similar to WT mice. In conclusion, the pro-inflammatory skin phenotype of PPAR-?-/- mice is due to lack of IL-2-mediated Treg induction in these mice.
SIRT1 regulates energy homeostasis by controlling the acetylation status and activity of a number of enzymes and transcriptional regulators. The fact that NAD(+) levels control SIRT1 activity confers a hypothetical basis for the design of new strategies to activate SIRT1 by increasing NAD(+) availability. Here we show that the deletion of the poly(ADP-ribose) polymerase-1 (PARP-1) gene, encoding a major NAD(+)-consuming enzyme, increases NAD(+) content and SIRT1 activity in brown adipose tissue and muscle. PARP-1(-/-) mice phenocopied many aspects of SIRT1 activation, such as a higher mitochondrial content, increased energy expenditure, and protection against metabolic disease. Also, the pharmacologic inhibition of PARP in vitro and in vivo increased NAD(+) content and SIRT1 activity and enhanced oxidative metabolism. These data show how PARP-1 inhibition has strong metabolic implications through the modulation of SIRT1 activity, a property that could be useful in the management not only of metabolic diseases, but also of cancer.
Mechanisms responsible for energy management in the cell and in the whole organism require a complex network of transcription factors and cofactors. Peroxisome proliferator-activated receptor ? coactivator 1? (PGC-1?) has emerged as a master regulator of mitochondrial biogenesis and function, thus becoming a crucial metabolic node. We present an overview of the mechanisms by which PGC-1? is regulated, including the transcriptional regulation of PGC-1? expression and the fine-tuning of its final activity via posttranslational modifications.
Transcriptional coregulators control the activity of many transcription factors and are thought to have wide-ranging effects on gene expression patterns. We show here that muscle-specific loss of nuclear receptor corepressor 1 (NCoR1) in mice leads to enhanced exercise endurance due to an increase of both muscle mass and of mitochondrial number and activity. The activation of selected transcription factors that control muscle function, such as MEF2, PPAR?/?, and ERRs, underpins these phenotypic alterations. NCoR1 levels are decreased in conditions that require fat oxidation, resetting transcriptional programs to boost oxidative metabolism. Knockdown of gei-8, the sole C. elegans NCoR homolog, also robustly increased muscle mitochondria and respiration, suggesting conservation of NCoR1 function. Collectively, our data suggest that NCoR1 plays an adaptive role in muscle physiology and that interference with NCoR1 action could be used to improve muscle function.
The nicotinamide adenine dinucleotide (NAD(+))-dependent deacetylase SIRT1 is a major metabolic regulator activated by energy stresses such as fasting or calorie restriction. SIRT1 activation during fasting not only relies on the increase in the NAD(+)/NADH ratio caused by energy deprivation but also involves an upregulation of SIRT1 mRNA and protein levels in various metabolic tissues. We demonstrate that SIRT1 expression is controlled systemically by the activation of the cyclic AMP response-element-binding protein upon low nutrient availability. Conversely, in the absence of energetic stress, the carbohydrate response-element-binding protein represses the expression of SIRT1. Altogether, these results demonstrate that SIRT1 expression is tightly controlled at the transcriptional level by nutrient availability and further underscore that SIRT1 is a crucial metabolic checkpoint connecting the energetic status with transcriptional programmes.
Insulin resistance, tissue inflammation, and adipose tissue dysfunction are features of obesity and Type 2 diabetes. We generated adipocyte-specific Nuclear Receptor Corepressor (NCoR) knockout (AKO) mice to investigate the function of NCoR in adipocyte biology, glucose and insulin homeostasis. Despite increased obesity, glucose tolerance was improved in AKO mice, and clamp studies demonstrated enhanced insulin sensitivity in liver, muscle, and fat. Adipose tissue macrophage infiltration and inflammation were also decreased. PPAR? response genes were upregulated in adipose tissue from AKO mice and CDK5-mediated PPAR? ser-273 phosphorylation was reduced, creating a constitutively active PPAR? state. This identifies NCoR as an adaptor protein that enhances the ability of CDK5 to associate with and phosphorylate PPAR?. The dominant function of adipocyte NCoR is to transrepress PPAR? and promote PPAR? ser-273 phosphorylation, such that NCoR deletion leads to adipogenesis, reduced inflammation, and enhanced systemic insulin sensitivity, phenocopying the TZD-treated state.
Liver receptor homolog-1 (NR5A2) is a nuclear receptor originally identified in the liver and mostly known for its regulatory role in cholesterol and bile acid homeostasis. More recently, liver receptor homolog-1 has emerged as a key regulator of intestinal function, coordinating unanticipated actions, such as cell renewal and local immune function with important implications to common intestinal diseases, including colorectal cancer and inflammatory bowel disease. Unlike most of the other nuclear receptors, liver receptor homolog-1 acts as a constitutively active transcription factor to drive the transcription of its target genes. Liver receptor homolog-1 activity however is to a major extent regulated by different corepressors and posttranslational modifications, which may account for its tissue-specific functions. This review will provide an update on the molecular aspects of liver receptor homolog-1 action and focus on some emerging aspects of its function in normal and diseased gut. This article is part of a Special Issue entitled: Translating nuclear receptors from health to disease.
Heritable changes to the transcriptome that are independent to changes in the genome are defined as epigenetics. DNA methylation and posttranslational modifications of histones, such as acetylation/deacetylation and methylation/demethylation of lysine residues, underlie these epigenetic phenomena, which impact on many physiological processes. This perspective focuses on the emerging biology of histone methylation and demethylation, highlighting how these reactions depend on metabolic coenzymes like S-adenosylmethionine, flavin adenine dinucleotide, and ?-ketoglutarate. Furthermore, we illustrate that methyltranferases and demethylases affect many metabolic pathways. Despite the preliminary evidence that methyltranferases and demethylases could link metabolic signals to chromatin and alter transcription, further research is indispensable to consolidate these enticing observations.
Calo et al. (2010) recently reported that the absence of the transcriptional regulator pRb enhances differentiation of mesenchymal precursors toward the brown adipocyte lineage in detriment of osteoblast and white fat populations. These findings may have therapeutic implications for cancer and bone and metabolic disease.
Bile acids (BAs) are amphipathic molecules that facilitate the uptake of lipids, and their levels fluctuate in the intestine as well as in the blood circulation depending on food intake. Besides their role in dietary lipid absorption, bile acids function as signaling molecules capable to activate specific receptors. These BA receptors are not only important in the regulation of bile acid synthesis and their metabolism, but also regulate glucose homeostasis, lipid metabolism, and energy expenditure. These processes are important in diabetes and other facets of the metabolic syndrome, which represents a considerable increasing health burden. In addition to the function of the nuclear receptor FXR? in regulating local effects in the organs of the enterohepatic axis, increasing evidence points to a crucial role of the G-protein coupled receptor (GPCR) TGR5 in mediating systemic actions of BAs. Here we discuss the current knowledge on BA receptors, with a strong focus on the cell membrane receptor TGR5, which emerges as a valuable target for intervention in metabolic diseases.
Molecular and cellular networks implicated in aging depend on a multitude of proteins that collectively mount adaptive and contingent metabolic responses to environmental challenges. Here, we discuss the intimate links between metabolic regulation and longevity and outline new approaches for analyzing and manipulating such links to promote human health span.
The AMP-activated protein kinase (AMPK) is a key regulator of catabolic versus anabolic processes. Its properties as an energy sensor allow it to couple the energy status of the cell to the metabolic environment. These adaptations not only take place through the acute modulation of key metabolic enzymes via direct phosphorylation, but also through a slower transcriptional adaptative response. The question of how AMPK regulates the expression of a number of gene sets, such as those related to mitochondrial biogenesis, energy production and oxidative protection, is only beginning to be elucidated, and still many questions remain to be answered. In this review we will try to integrate our current knowledge on how AMPK regulates transcription in muscle and liver, which will serve as examples to illustrate the major advances in the field and the key challenges ahead.
Related JoVE Video
Journal of Visualized Experiments
What is Visualize?
JoVE Visualize is a tool created to match the last 5 years of PubMed publications to methods in JoVE's video library.
How does it work?
We use abstracts found on PubMed and match them to JoVE videos to create a list of 10 to 30 related methods videos.
Video X seems to be unrelated to Abstract Y...
In developing our video relationships, we compare around 5 million PubMed articles to our library of over 4,500 methods videos. In some cases the language used in the PubMed abstracts makes matching that content to a JoVE video difficult. In other cases, there happens not to be any content in our video library that is relevant to the topic of a given abstract. In these cases, our algorithms are trying their best to display videos with relevant content, which can sometimes result in matched videos with only a slight relation.