COMMENTARY

PHOSPHO1 puts the breaks on thermogenesis in brown adipocytes COMMENTARY Christy M. Gliniaka and Philipp E. Scherera,b,1

The nonshivering heat production in mitochondria-rich the enzyme in BAT cannot easily be predicted. A recent brown or beige adipocytes allows rodents and humans study identified that PHOSPHO1 regulates oxygen to adapt to cold stress (1). The best-characterized ther- consumption in BAT (2). Jiang et al. (5) extend this mogenic effector is uncoupling protein 1 (UCP1), which observation and performed several bioinformatic dissipates energy as heat by proton transport across the analyses that show that the molecular network around mitochondrial inner membrane. Of note, there are also PHOSPHO1 correlates with increased classical thermo- important UCP1-independent pathways mediating genic genes, such as Ucp1, and with proteins involved brown adipose tissue (BAT) thermogenesis (2). The dis- in mitochondrial electron transport and lipid catabolism. covery of significant brown and beige adipose depots A feature of activated BAT is tolerance to cold in humans initiated efforts to leverage the thermogenic stress. Sympathetic nerve activity stimulates BAT via processasameanstoincreasethemetabolicrateto norepinephrine activation of adrenergic receptors. prevent metabolic disease, such as obesity and diabe- This not only promotes thermogenic gene expression tes (3). Currently, there are limited browning regimens but also stimulates lipolysis of triacylglycerol, which for increased energy expenditure in humans short of increases intracellular free fatty acid (FFA) concentra- cold exposure (4). In PNAS, Jiang et al. (5) investi- tions. The FFAs bind to and activate UCP1, thus gated how the enzyme phosphoethanolamine and switching on UCP1-mediated heat production. The phosphocholine phosphatase 1 (PHOSPHO1) regu- authors describe that PHOSPHO1 knockout (KO) mice lates thermogenesis in BAT, triggering changes in sys- show an improved ability to maintain body tempera- temic insulin sensitivity and energy balance. PHOSPHO1 ture during acute cold exposure compared with wild- catalyzes the hydrolysis of phosphocholine (PC) to cho- type (WT) mice. Moreover, upon cold exposure, line as well as of phosphoethanolamine (PEA) to eth- PHOSPHO1 null mice have increased thermogenic anolamine and in the process, releases an inorganic gene expression in BAT and reduced lipid stores. This phosphate group. is consistent with the events during short-term cold ex- PHOSPHO1 is not widely expressed, although it has posure when BAT initially oxidizes intracellular lipid been well characterized as the PC and PEA phosphatase stores to produce heat. Differentiated brown adipo- in bone, where free inorganic phosphate is essential for cytes from PHOSPHO1 null mice displayed increased bone mineralization (6). Additionally, PHOSPHO1 is a maximal oxygen consumption. Therefore, the gene ex- key regulator that coordinates the change in PC and pression and functional data combined suggest that phospholipid composition during terminal erythropoiesis PHOSPHO1 enzymatic activity inhibits a full thermo- (7). It is remarkable that bone and red blood cell progen- genic response in BAT, and when PHOSPHO1 is de- itors rely on different aspects of PHOSPHO1 enzymatic pleted, energy expenditure and BAT thermogenesis activity. The authors discovered that PHOSPHO1 is intensify. highly enriched in murine BAT and accumulates during Given the fact that the induction of thermogenesis in vitro differentiation of primary brown adipocytes. can prevent obesity and other metabolic disease PHOSPHO1 expression also increased during cold ex- caused by overnutrition in mice (8), Jiang et al. (5) posure in mice. Furthermore, they determined that met- examined energy balance in PHOSPHO1 KO mice abolically active BAT from humans at the fetal stage has that were fed a high-fat diet (HFD). The KO mice high levels of PHOSPHO1 vs. adult fat tissues. gained less weight than controls, primarily due to de- There is little known about the role of PHOSPHO1 in creased fat mass. KO mice displayed increased oxy- adipose tissue. Based on the literature, the function of gen consumption with no change in food intake.

aTouchstone Diabetes Center, The University of Texas Southwestern Medical Center, Dallas, TX 75390-8549; and bDepartment of Cell Biology, The University of Texas Southwestern Medical Center, Dallas, TX 75390-8549 Author contributions: C.M.G. and P.E.S. wrote the paper. The authors declare no competing interest. Published under the PNAS license. See companion article, “Phosphocholine accumulation and PHOSPHO1 depletion promote adipose tissue thermogenesis,” 10.1073/pnas. 1916550117. 1To whom correspondence may be addressed. Email: [email protected].

www.pnas.org/cgi/doi/10.1073/pnas.2011052117 PNAS Latest Articles | 1of3 Downloaded by guest on October 2, 2021 Fig. 1. Pleiotropic effects of PHOSPHO1 elimination on thermogenesis in brown and beige adipocytes. The biosynthesis of PtdChol and PE occurs primarily in the ER. In the cytosol, PHOSPHO1 catalyzes the hydrolysis of PC to choline or PEA to ethanolamine (Etn). The lack of PHOSPHO1 (highlighted in yellow as PHOSPHO1 KO) stimulates adrenergic receptor-mediated lipolysis, enhances UCP1 activity, stimulates glucose transport, and leads to enhanced transcription of thermogenic genes. Lack of PHOSPHO1 likely affects the ratio of PtdChol to PE, a change that profoundly affects the biophysical properties of membranes. Other enzymes in the pathway include choline kinase (CK), phosphocholine cytidylyltransferase 1a (CCTα), choline phosphotransferase (CPT), ethanolamine kinase (EK), phosphoethanolamine cytidylyltransferase (ET), choline-ethanolamine phosphotransferase (CEPT), and phosphatidylethanolamine N-methyltransferase (PEMT). Acyl- CoA, acetyl coenzyme A; cAMP, cyclic adenosine monophosphate; CPT1, carnitine palmitoyltransferase 1; CPT2, carnitine palmitoyltransferase 2; ETC, electron transport chain; Glut4, glucose transporter 4; TG, triglyceride.

Therefore, the weight loss could be explained by increased en- phospholipid, comprising 40 to 50% of phospholipids in mamma- ergy expenditure due to thermogenesis in the KOs compared lian cells, with PE comprising 15 to 25% of all phospholipids (10). with controls. PHOSPHO1 KO mice fed HFD were also more glu- In Jiang et al. (5), PC was one of the top metabolites induced cose tolerant and insulin sensitive compared with controls. Acute during BAT thermogenesis, which is suggestive that PC may be cold exposure increases glucose uptake in BAT, which may ex- driving effective thermogenesis in PHOSPHO1 KO mice. It would plain the glucose-lowering response (9). Overall, these findings be interesting to test whether PEA also rescued any of the effects highlight the glucose-lowering potential of BAT in rodents. of PHOSPO1 KO in BAT. Whether a similar degree of improvement can be reached clini- PHOSPHO1 appears to have UCP1-dependent and -independent cally through the manipulation of PC remains to be seen. effects on lipid metabolism and fatty acid oxidation. For What is the mechanism underlying how the enzyme PHOSPHO1 example, in mice, the ablation of PHOSPHO1 or PC treatment regulates BAT thermogenesis? Some consideration was given to increased UCP1 expression. On the other hand, PC is a precursor the idea that PHOSPHO1 may be involved directly in the futile cycle to PtdChol (10). PC and PE can give rise to a diversity of lipid of creatine metabolism, a mechanism that increases energy expen- species that contribute to membrane structure and permeability, diture in BAT. However, PHOSPHO1 has limited phosphatase essential for effective thermogenesis. The acyl chains can be activity toward phosphocreatine (2). Similar to the role of remodeled by phospholipases (A, C, and D) and by lysophospholipid PHOSPHO1 in bone, it is possible that the liberated inorganic transferases to generate bioactive lipid mediators. Also, inhibition of phosphate contributes to thermogenesis; however, there is the synthesis of PtdChol or PE results in elevated diacylglycerol levels little evidence to support this. Jiang et al. (5) discovered that and/or reduced sphingomyelin levels and can have profound effects PHOSPHO1 KO mice had a higher PC to choline ratio in the serum on endoplasmic reticulum (ER) and Golgi-mediated functions and compared with WT. Therefore, the effects of PHOSPHO1 KO in triglyceride accumulation (11, 12). BAT may be mediated through increased intracellular PC. PC treat- Changes in the absolute concentrations of phospholipids are ment of brown adipocytes phenocopied the thermogenic as- important, but a delicate balance of cellular PtdChol/PE ratios is pects of the PHOSPHO1 KO mouse. This supports the idea required to maintain energy balance and prevention of disease. In that the thermogenic program induced by PHOSPHO1 is cell mitochondria, changes in the PtdChol/PE molar ratio affect autonomous and indicates that the effects of PHOSPHO1 ablation energy production (13). Kennedy and Weiss discovered unex- in BAT may be due to an accumulation of PC in critical intracellular pectedly (14) that the high-energy compound cytidine triphos- membranes. phate (CTP) is the substrate for the biosynthesis of PtdChol and Since PC serves as a precursor to many lipids, the mechanisms PE, rather than adenosine triphosphate. Thus, PC cytidylyl- by which the lack of PHOSPHO1 regulates cellular metabolism in transferase 1a catalyzes the conversion of PC and CTP to cyti- BAT are not clear. While it is very compelling that PC-treated mice dine diphosphocholine (CDP-choline) plus pyrophosphate. phenocopy the effects seen in PHOSPHO1 KO mice, an important This is the rate-limiting step that determines the flux of choline consideration for future studies is whether PHOSPHO1 also alters to PtdChol. The reactions of the CDP-ethanolamine pathway the metabolism of its other substrate, PEA. Cells maintain a large for PE synthesis parallel those of the CDP-choline pathway for pool of PC and PEA as precursors for the rate-limiting steps for PC synthesis. PEA cytidylyltransferase converts CTP and PEA to phosphatidylcholine (PtdChol) and phosphatidylethanolamine CDP-ethanolamine and pyrophosphate and is normally the (PE) synthesis, respectively. PtdChol is the most abundant rate-limiting enzyme of the pathway.

2of3 | www.pnas.org/cgi/doi/10.1073/pnas.2011052117 Gliniak and Scherer Downloaded by guest on October 2, 2021 The clinical importance of maintaining the hepatic PtdChol/PE increase in the relative amount of PE on the surface of lipid drop- ratio in an appropriate range is well appreciated (15). A small lets can promote fusion of smaller droplets into larger ones (17), reduction in the PtdChol/PE molar ratio predicts nonalcoholic an essential process regulating the constant flux of lipids in adi- liver disease (NAFLD), whereas an extreme reduction in the pocytes (Fig. 1). PtdChol/PE molar ratio predicts liver failure (10). Patients with Future studies using inducible, tissue-specific mouse models NAFLD have low PtdChol levels and high PE levels (i.e., the modulating PHOSPHO1 and other enzymes that alter the synthe- PtdChol/PE molar ratio is low) in the liver compared with healthy sis of PtdChol and PE will help elucidate the effects of these patients. PtdChol/PE molar ratios also regulate hepatic triglycer- phospholipids. Overall, it may be said that an enhanced un- ide metabolism and the formation, stability, and clearance of derstanding of the mechanisms that regulate phospholipid me- cholesterol lipoproteins. Of note, hepatic PtdChol can be synthe- tabolism will support the development of therapies that normalize sized via the choline pathway or by methylation of PE via PE PtdChol/PE molar ratios. This could prove immensely valuable in N-methyltransferase (15). slowing the progression of obesity, diabetes, and NAFLD, beyond Phospholipid ratios are also important in adipocytes. One role the effects in BAT. Jiang et al. (5) have taken a bold step in high- of adipocytes is to prevent lipotoxicity by incorporating excess lighting what powerful impact the manipulation of this pathway FFAs to triglycerides, which is stored in cytosolic lipid droplets. can have on systemic metabolism. The relative ratio of PtdChol and PE regulates the size and dynamics of lipid droplets. It has been shown that inhibition of PtdChol biosynthesis during conditions that promote triglyceride Acknowledgments We are supported by NIH Grants R01-DK55758, RC2-DK118620, P01- storage increases the size of the lipid droplets, possibly in part DK088761, R01-DK099110, and P01-AG051459 (to P.E.S.). C.M.G. is supported because there is limited PtdChol for lipid droplet surfaces, thus by NIH Grants T32-DK007307 and F32-DK122623. P.E.S. was supported by an promoting the fusion of lipid droplets (16, 17). Additionally, an unrestricted research grant from the Novo Nordisk Foundation.

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