Facultative protein selenation regulates redox sensitivity, adipose tissue thermogenesis, and obesity.

Oxidation of cysteine thiols by physiological reactive oxygen species (ROS) initiates thermogenesis in brown and beige adipose tissues. Cellular selenocysteines, where sulfur is replaced with selenium, exhibit enhanced reactivity with ROS. Despite their critical roles in physiology, methods for broad and direct detection of proteogenic selenocysteines are limited. Here we developed a mass spectrometric method to interrogate incorporation of selenium into proteins. Unexpectedly, this approach revealed facultative incorporation of selenium as selenocysteine or selenomethionine into proteins that lack canonical encoding for selenocysteine. Selenium was selectively incorporated into regulatory sites on key metabolic proteins, including as selenocysteine-replacing cysteine at position 253 in uncoupling protein 1 (UCP1). This facultative utilization of selenium was initiated by increasing cellular levels of organic, but not inorganic, forms of selenium. Remarkably, dietary selenium supplementation elevated facultative incorporation into UCP1, elevated energy expenditure through thermogenic adipose tissue, and protected against obesity. Together, these findings reveal the existence of facultative protein selenation, which correlates with impacts on thermogenic adipocyte function and presumably other biological processes as well.

Accumulation of succinate controls activation of adipose tissue thermogenesis.

Thermogenesis by brown and beige adipose tissue, which requires activation by external stimuli, can counter metabolic disease1. Thermogenic respiration is initiated by adipocyte lipolysis through cyclic AMP-protein kinase A signalling; this pathway has been subject to longstanding clinical investigation2-4. Here we apply a comparative metabolomics approach and identify an independent metabolic pathway that controls acute activation of adipose tissue thermogenesis in vivo. We show that substantial and selective accumulation of the tricarboxylic acid cycle intermediate succinate is a metabolic signature of adipose tissue thermogenesis upon activation by exposure to cold. Succinate accumulation occurs independently of adrenergic signalling, and is sufficient to elevate thermogenic respiration in brown adipocytes. Selective accumulation of succinate may be driven by a capacity of brown adipocytes to sequester elevated circulating succinate. Furthermore, brown adipose tissue thermogenesis can be initiated by systemic administration of succinate in mice. Succinate from the extracellular milieu is rapidly metabolized by brown adipocytes, and its oxidation by succinate dehydrogenase is required for activation of thermogenesis. We identify a mechanism whereby succinate dehydrogenase-mediated oxidation of succinate initiates production of reactive oxygen species, and drives thermogenic respiration, whereas inhibition of succinate dehydrogenase supresses thermogenesis. Finally, we show that pharmacological elevation of circulating succinate drives UCP1-dependent thermogenesis by brown adipose tissue in vivo, which stimulates robust protection against diet-induced obesity and improves glucose tolerance. These findings reveal an unexpected mechanism for control of thermogenesis, using succinate as a systemically-derived thermogenic molecule.

Genetic Depletion of Adipocyte Creatine Metabolism Inhibits Diet-Induced Thermogenesis and Drives Obesity.

Diet-induced thermogenesis is an important homeostatic mechanism that limits weight gain in response to caloric excess and contributes to the relative stability of body weight in most individuals. We previously demonstrated that creatine enhances energy expenditure through stimulation of mitochondrial ATP turnover, but the physiological role and importance of creatine energetics in adipose tissue have not been explored. Here, we have inactivated the first and rate-limiting enzyme of creatine biosynthesis, glycine amidinotransferase (GATM), selectively in fat (Adipo-Gatm KO). Adipo-Gatm KO mice are prone to diet-induced obesity due to the suppression of elevated energy expenditure that occurs in response to high-calorie feeding. This is paralleled by a blunted capacity for ß3-adrenergic activation of metabolic rate, which is rescued by dietary creatine supplementation. These results provide strong in vivo genetic support for a role of GATM and creatine metabolism in energy expenditure, diet-induced thermogenesis, and defense against diet-induced obesity.

UCP1 deficiency causes brown fat respiratory chain depletion and sensitizes mitochondria to calcium overload-induced dysfunction.

Brown adipose tissue (BAT) mitochondria exhibit high oxidative capacity and abundant expression of both electron transport chain components and uncoupling protein 1 (UCP1). UCP1 dissipates the mitochondrial proton motive force (Δp) generated by the respiratory chain and increases thermogenesis. Here we find that in mice genetically lacking UCP1, cold-induced activation of metabolism triggers innate immune signaling and markers of cell death in BAT. Moreover, global proteomic analysis reveals that this cascade induced by UCP1 deletion is associated with a dramatic reduction in electron transport chain abundance. UCP1-deficient BAT mitochondria exhibit reduced mitochondrial calcium buffering capacity and are highly sensitive to mitochondrial permeability transition induced by reactive oxygen species (ROS) and calcium overload. This dysfunction depends on ROS production by reverse electron transport through mitochondrial complex I, and can be rescued by inhibition of electron transfer through complex I or pharmacologic depletion of ROS levels. Our findings indicate that the interscapular BAT of Ucp1 knockout mice exhibits mitochondrial disruptions that extend well beyond the deletion of UCP1 itself. This finding should be carefully considered when using this mouse model to examine the role of UCP1 in physiology.

Mitochondrial ROS regulate thermogenic energy expenditure and sulfenylation of UCP1.

Brown and beige adipose tissues can dissipate chemical energy as heat through thermogenic respiration, which requires uncoupling protein 1 (UCP1). Thermogenesis from these adipocytes can combat obesity and diabetes, encouraging investigation of factors that control UCP1-dependent respiration in vivo. Here we show that acutely activated thermogenesis in brown adipose tissue is defined by a substantial increase in levels of mitochondrial reactive oxygen species (ROS). Remarkably, this process supports in vivo thermogenesis, as pharmacological depletion of mitochondrial ROS results in hypothermia upon cold exposure, and inhibits UCP1-dependent increases in whole-body energy expenditure. We further establish that thermogenic ROS alter the redox status of cysteine thiols in brown adipose tissue to drive increased respiration, and that Cys253 of UCP1 is a key target. UCP1 Cys253 is sulfenylated during thermogenesis, while mutation of this site desensitizes the purine-nucleotide-inhibited state of the carrier to adrenergic activation and uncoupling. These studies identify mitochondrial ROS induction in brown adipose tissue as a mechanism that supports UCP1-dependent thermogenesis and whole-body energy expenditure, which opens the way to improved therapeutic strategies for combating metabolic disorders.

A creatine-driven substrate cycle enhances energy expenditure and thermogenesis in beige fat.

Thermogenic brown and beige adipose tissues dissipate chemical energy as heat, and their thermogenic activities can combat obesity and diabetes. Herein the functional adaptations to cold of brown and beige adipose depots are examined using quantitative mitochondrial proteomics. We identify arginine/creatine metabolism as a beige adipose signature and demonstrate that creatine enhances respiration in beige-fat mitochondria when ADP is limiting. In murine beige fat, cold exposure stimulates mitochondrial creatine kinase activity and induces coordinated expression of genes associated with creatine metabolism. Pharmacological reduction of creatine levels decreases whole-body energy expenditure after administration of a ß3-agonist and reduces beige and brown adipose metabolic rate. Genes of creatine metabolism are compensatorily induced when UCP1-dependent thermogenesis is ablated, and creatine reduction in Ucp1-deficient mice reduces core body temperature. These findings link a futile cycle of creatine metabolism to adipose tissue energy expenditure and thermal homeostasis. PAPERCLIP.