An exercise-inducible metabolite that suppresses feeding and obesity.

Exercise confers protection against obesity, type 2 diabetes and other cardiometabolic diseases1-5. However, the molecular and cellular mechanisms that mediate the metabolic benefits of physical activity remain unclear6. Here we show that exercise stimulates the production of N-lactoyl-phenylalanine (Lac-Phe), a blood-borne signalling metabolite that suppresses feeding and obesity. The biosynthesis of Lac-Phe from lactate and phenylalanine occurs in CNDP2+ cells, including macrophages, monocytes and other immune and epithelial cells localized to diverse organs. In diet-induced obese mice, pharmacological-mediated increases in Lac-Phe reduces food intake without affecting movement or energy expenditure. Chronic administration of Lac-Phe decreases adiposity and body weight and improves glucose homeostasis. Conversely, genetic ablation of Lac-Phe biosynthesis in mice increases food intake and obesity following exercise training. Last, large activity-inducible increases in circulating Lac-Phe are also observed in humans and racehorses, establishing this metabolite as a molecular effector associated with physical activity across multiple activity modalities and mammalian species. These data define a conserved exercise-inducible metabolite that controls food intake and influences systemic energy balance.

Cell type-selective secretome profiling in vivo.

Secreted polypeptides are a fundamental axis of intercellular and endocrine communication. However, a global understanding of the composition and dynamics of cellular secretomes in intact mammalian organisms has been lacking. Here, we introduce a proximity biotinylation strategy that enables labeling, detection and enrichment of secreted polypeptides in a cell type-selective manner in mice. We generate a proteomic atlas of hepatocyte, myocyte, pericyte and myeloid cell secretomes by direct purification of biotinylated secreted proteins from blood plasma. Our secretome dataset validates known cell type-protein pairs, reveals secreted polypeptides that distinguish between cell types and identifies new cellular sources for classical plasma proteins. Lastly, we uncover a dynamic and previously undescribed nutrient-dependent reprogramming of the hepatocyte secretome characterized by the increased unconventional secretion of the cytosolic enzyme betaine-homocysteine S-methyltransferase (BHMT). This secretome profiling strategy enables dynamic and cell type-specific dissection of the plasma proteome and the secreted polypeptides that mediate intercellular signaling.

Adipose Tissue Lipokines: Recent Progress and Future Directions.

Beyond classical metabolic functions in energy storage and energy expenditure, adipose tissue is also a dynamic endocrine organ that secretes bioactive factors into blood plasma. Historically, studies of the adipose secretome have predominantly focused on polypeptide adipokines. Recently, adipose-derived blood-borne lipids ("lipokines") have emerged as a distinct class of endocrine factors. Lipokines are intimately connected to intracellular pathways of fatty acid metabolism and therefore uniquely poised to communicate the intracellular energy status of adipocytes to other nonadipose tissues including liver, muscle, and pancreas. Here, we discuss recent progress on our understanding of adipose-secreted lipokines as endocrine regulators of glucose and lipid metabolism. We also provide our perspective on future directions for adipose-secreted lipids, including limitations of the currently available experimental data as well as potential strategies for addressing the remaining open questions.

A Plasma Protein Network Regulates PM20D1 and N-Acyl Amino Acid Bioactivity.

N-acyl amino acids are a family of cold-inducible circulating lipids that stimulate thermogenesis. Their biosynthesis is mediated by a secreted enzyme called PM20D1. The extracellular mechanisms that regulate PM20D1 or N-acyl amino acid activity in the complex environment of blood plasma remains unknown. Using quantitative proteomics, here we show that PM20D1 circulates in tight association with both low- and high-density lipoproteins. Lipoprotein particles are powerful co-activators of PM20D1 activity in vitro and N-acyl amino acid biosynthesis in vivo. We also identify serum albumin as a physiologic N-acyl amino acid carrier, which spatially segregates N-acyl amino acids away from their sites of production, confers resistance to hydrolytic degradation, and establishes an equilibrium between thermogenic "free" versus inactive "bound" fractions. These data establish lipoprotein particles as principal extracellular sites of N-acyl amino acid biosynthesis and identify a lipoprotein-albumin network that regulates the activity of a circulating thermogenic lipid family.

Cooperative enzymatic control of N-acyl amino acids by PM20D1 and FAAH.

The N-acyl amino acids are a family of bioactive lipids with pleiotropic physiologic functions, including in energy homeostasis. Their endogenous levels are regulated by an extracellular mammalian N-acyl amino acid synthase/hydrolase called PM20D1 (peptidase M20 domain containing 1). Using an activity-guided biochemical approach, we report the molecular identification of fatty acid amide hydrolase (FAAH) as a second intracellular N-acyl amino acid synthase/hydrolase. In vitro, FAAH exhibits a more restricted substrate scope compared to PM20D1. In mice, genetic ablation or selective pharmacological inhibition of FAAH bidirectionally dysregulates intracellular, but not circulating, N-acyl amino acids. Dual blockade of both PM20D1 and FAAH reveals a dramatic and non-additive biochemical engagement of these two enzymatic pathways. These data establish FAAH as a second intracellular pathway for N-acyl amino acid metabolism and underscore enzymatic division of labor as an enabling strategy for the regulation of a structurally diverse bioactive lipid family.

Family-wide Annotation of Enzymatic Pathways by Parallel In Vivo Metabolomics.

Enzymes catalyze fundamental biochemical reactions that control cellular and organismal homeostasis. Here we present an approach for de novo biochemical pathway discovery across entire mammalian enzyme families using parallel viral transduction in mice and untargeted liquid chromatography-mass spectrometry. Applying this method to the M20 peptidases uncovers both known pathways of amino acid metabolism as well as a previously unknown CNDP2-regulated pathway for threonyl dipeptide catabolism. Ablation of CNDP2 in mice elevates threonyl dipeptides across multiple tissues, establishing the physiologic relevance of our biochemical assignments. Taken together, these data underscore the utility of parallel in vivo metabolomics for the family-wide discovery of enzymatic pathways.