No abstract

Summary Mitochondrial function is important for aspartate biosynthesis in proliferating cells. Here, we show that mitochondrial aspartate export via the aspartate-glutamate carrier 1 (AGC1) supports cell proliferation and cellular redox homeostasis. Insufficient cytosolic aspartate delivery leads to cell death when TCA cycle carbon is reduced following glutamine withdrawal and/or glutaminase inhibition. Moreover, loss of AGC1 reduces allograft tumor growth that is further compromised by treatment with the glutaminase inhibitor CB-839. Together, these findings argue that mitochondrial aspartate export sustains cell survival in low-glutamine environments and AGC1 inhibition can synergize with glutaminase inhibition to limit tumor growth.

The current dogma is that obesity-associated hepatic inflammation is due to increased Kupffer cell (KC) activation. However, recruited hepatic macrophages (RHMs) were recently shown to represent a sizable liver macrophage population in the context of obesity. Therefore, we assessed whether KCs and RHMs, or both, represent the major liver inflammatory cell type in obesity. We used a combination of in vivo macrophage tracking methodologies and adoptive transfer techniques in which KCs and RHMs are differentially labeled with fluorescent markers. With these approaches, the inflammatory phenotype of these distinct macrophage populations was determined under lean and obese conditions. In vivo macrophage tracking revealed an approximately sixfold higher number of RHMs in obese mice than in lean mice, whereas the number of KCs was comparable. In addition, RHMs comprised smaller size and immature, monocyte-derived cells compared with KCs. Furthermore, RHMs from obese mice were more inflamed and expressed higher levels of tumor necrosis factor-α and interleukin-6 than RHMs from lean mice. A comparison of the MCP-1/C-C chemokine receptor type 2 (CCR2) chemokine system between the two cell types showed that the ligand (MCP-1) is more highly expressed in KCs than in RHMs, whereas CCR2 expression is approximately fivefold greater in RHMs. We conclude that KCs can participate in obesity-induced inflammation by causing the recruitment of RHMs, which are distinct from KCs and are not precursors to KCs. These RHMs then enhance the severity of obesity-induced inflammation and hepatic insulin resistance.

Triacylglycerols are stored in eukaryotic cells within lipid droplets (LD).The LD core is enwrapped by a phospholipid monolayer with phosphatidylcholine (PC), the major phospholipid, and phosphatidylethanolamine (PE), a minor component. We demonstrate that the onset of LD formation is characterized by a change in cellular PC, PE, and phosphatidylserine (PS). With induction of differentiation of 3T3-L1 fibroblasts into adipocytes, the cellular PC/PE ratio decreased concomitant with LD formation, with the most pronounced decline between confluency and day 5. The mRNA for PS synthase-1 (forms PS from PC) and PS decarboxylase (forms PE from PS) increased after day 5. Activity and protein of PE N-methyltransferase (PEMT), which produces PC by methylation of PE, are absent in 3T3-L1 fibroblasts but were induced at day 5. High fat challenge induced PEMT expression in mouse adipose tissue. PE, produced via PS decarboxylase, was the preferred substrate for methylation to PC. A PEMT-GFP fusion protein decorated the periphery of LD. PEMT knockdown in 3T3-L1 adipocytes correlated with increased basal triacylglycerol hydrolysis. Pemt ؊/؊ mice developed desensitization against adenosine-mediated inhibition of basal hydrolysis in adipose tissue, and adipocyte hypotrophy was observed in Pemt ؊/؊ animals on a high fat diet. Knock-out of PEMT in adipose tissue down-regulated PS synthase-1 mRNA, suggesting coordination between PE supply and converting pathways during LD biosynthesis. We conclude that two consecutive processes not previously related to LD biogenesis, (i) PE production via PS and (ii) PE conversion via PEMT, are implicated in LD formation and stability.Cytosolic neutral lipid droplets (LD) 7 within eukaryotic cells represent intracellular storage compartments for triacylglycerols (TG) and cholesteryl esters to bridge alimentary or metabolic gaps (1). Adipocytes are the body's primary depots for efficient TG storage. Upon defined stimulation of adipocytes (2), fatty acids are released from TG droplets to supply energy or to provide essential components for the synthesis of biological membranes. Given these important functions, LD of all cell types are considered as dynamic organelles that represent fundamental components of intracellular lipid homeostasis (3).It is generally accepted that LD originate from the cytosolic leaflet of the endoplasmic reticulum (ER), contain a core of neutral lipids, and are surrounded by a phospholipid (PL) monolayer (4, 5). The finding that cytosolic LD in adipocytes contain at their periphery minor amounts of ER proteins such as BiP (6) and calnexin might reflect their site of origin in the ER. The primary LD proteins are so-called cage proteins, which not only stabilize LD but also protect their neutral lipid cores from unregulated degradation (7). Several other proteins contributing to the regulation of intracellular vesicle trafficking or targeting (3), and components of the intermediate filament protein machinery, have also been shown to be associated with intracellular LD (8, 9).D...