Seipin is required for converting nascent to mature lipid droplets

How proteins control the biogenesis of cellular lipid droplets (LDs) is poorly understood. Using Drosophila and human cells, we show here that seipin, an ER protein implicated in LD biology, mediates a discrete step in LD formation—the conversion of small, nascent LDs to larger, mature LDs. Seipin forms discrete and dynamic foci in the ER that interact with nascent LDs to enable their growth. In the absence of seipin, numerous small, nascent LDs accumulate near the ER and most often fail to grow. Those that do grow prematurely acquire lipid synthesis enzymes and undergo expansion, eventually leading to the giant LDs characteristic of seipin deficiency. Our studies identify a discrete step of LD formation, namely the conversion of nascent LDs to mature LDs, and define a molecular role for seipin in this process, most likely by acting at ER-LD contact sites to enable lipid transfer to nascent LDs. DOI: http://dx.doi.org/10.7554/eLife.16582.00

Lipid Droplets in Brown Adipose Tissue Are Dispensable for Cold-Induced Thermogenesis

SUMMARYBrown adipocytes store metabolic energy as triglycerides (TG) in multilocular lipid droplets (LDs). Fatty acids released from brown adipocyte LDs by lipolysis are thought to activate and fuel UCP1-mediated thermogenesis. Here we test this hypothesis by preventing fatty acid storage in murine brown adipocytes through brown adipose tissue (BAT)-specific deletions of the TG synthesis enzymes, DGAT1 and DGAT2 (BA-DGAT KO). Despite the absence of LDs, BA-DGAT KO mice had functional BAT and maintained euthermia during acute or chronic cold exposure. As apparent adaptations to the lack of TG, brown adipocytes of BA-DGAT KO mice appear to utilize circulating glucose and fatty acids, as well as stored glycogen to fuel thermogenesis. Moreover, BA-DGAT KO mice were resistant to diet-induced glucose intolerance, likely due to increased glucose disposal by BAT. Thus, surprisingly, TGs in BAT are dispensable for its function, in part through adaptations to utilize other fuel sources.</jats:p

Assessment of lipidomic species in hepatocyte lipid droplets from stressed mouse models

Lipid droplets are considered to be the hub for storage and metabolism of cellular lipids. In previous work we have phenotyped the lipidome of murine hepatocyte lipid droplets using liquid chromatography-mass spectrometry (UHPLC-MS) plus integrated MS/MS, followed by automatic analysis of the MS data. The organelles were isolated after intervention studies involving nutritional stress (extended feeding of a high fat diet or short term fasting), genetic stress due to knock-out of adipocyte triglyceride lipase, or by combined application of nutritional and genetic stress together (‘super stress’). Lipidomics at the level of lipid species (profiling of lipid classes) and lipid molecular species (structural analysis in parallel) has unraveled clear lipid droplet phenotypes as judged by patterns seen best in triacylglycerol (TG) lipidomes, but also in diacylglycerol and phosphatidylcholine lipidomes. The combined view of these data presented here validates the methods used and provides high quality lipidomic data for further bioinformatic inspections. Examples are given for identification of TG species subsets considered surrogates for whole TG lipidomes

Conditional targeting of phosphatidylserine decarboxylase to lipid droplets

ABSTRACT Phosphatidylethanolamine is an abundant component of most cellular membranes whose physical and chemical properties modulate multiple aspects of organelle membrane dynamics. An evolutionarily ancient mechanism for producing phosphatidylethanolamine is to decarboxylate phosphatidylserine and the enzyme catalyzing this reaction, phosphatidylserine decarboxylase, localizes to the inner membrane of the mitochondrion. We characterize a second form of phosphatidylserine decarboxylase, termed PISD-LD, that is generated by alternative splicing of PISD pre-mRNA and localizes to lipid droplets and to mitochondria. Sub-cellular targeting is controlled by a common segment of PISD-LD that is distinct from the catalytic domain and is regulated by nutritional state. Growth conditions that promote neutral lipid storage in lipid droplets favors targeting to lipid droplets, while targeting to mitochondria is favored by conditions that promote consumption of lipid droplets. Depletion of both forms of phosphatidylserine decarboxylase impairs triacylglycerol synthesis when cells are challenged with free fatty acid, indicating a crucial role phosphatidylserine decarboxylase in neutral lipid storage. The results reveal a previously unappreciated role for phosphatidylserine decarboxylase in lipid droplet biogenesis.</jats:p

Lipid Data Analyzer: unattended identification and quantitation of lipids in LC-MS data

Abstract Motivation: The accurate measurement of the lipidome permits insights into physiological and pathological processes. Of the present high-throughput technologies, LC-MS especially bears potential of monitoring quantitative changes in hundreds of lipids simultaneously. In order to extract valuable information from huge amount of mass spectrometry data, the aid of automated, reliable, highly sensitive and specific analysis algorithms is indispensable. Results: We present here a novel approach for the quantitation of lipids in LC-MS data. The new algorithm obtains its analytical power by two major innovations: (i) a 3D algorithm that confines the peak borders in m/z and time direction and (ii) the use of the theoretical isotopic distribution of an analyte as selection/exclusion criterion. The algorithm is integrated in the Lipid Data Analyzer (LDA) application which additionally provides standardization, a statistics module for results analysis, a batch mode for unattended analysis of several runs and a 3D viewer for the manual verification. The statistics module offers sample grouping, tests between sample groups and export functionalities, where the results are visualized by heat maps and bar charts. The presented algorithm has been applied to data from a controlled experiment and to biological data, containing analytes distributed over an intensity range of 106. Our approach shows improved sensitivity and an extremely high positive predictive value compared with existing methods. Consequently, the novel algorithm, integrated in a user-friendly application, is a valuable improvement in the high-throughput analysis of the lipidome. Implementation and availability: The Java application is freely available for non-commercial users at http://genome.tugraz.at/lda. Raw data associated with this manuscript may be downloaded from ProteomeCommons.org Tranche using the following hash: ZBh3nS5bXk6I/Vn32tB5Vh0qnMpVIW71HByFFQqM0RmdF4/4Hcn H3Wggh9kU2teYVOtM1JWwHIeMHqSS/bc2yYNFmyUAAAAAAACl DQ == Contact:  [email protected] Supplementary information:  Supplementary data are available from Bioinformatics online.</jats:p

Lipodystrophy can be uncoupled from detrimental metabolic consequences

SUMMARYAdipose tissue has at least two major functions: storing metabolic energy as triacylglycerols (TG) and coordinating metabolism by secreting hormones, such as leptin. In lipodystrophies, defects of storing TG are typically accompanied by metabolic abnormalities, such as hepatic steatosis, and endocrine perturbations. Thus, the concept emerged that the endocrine function of adipose tissue is coordinated with, and requires, TG stores. To test this notion, we selectively depleted adipose TG stores by deleting the TG synthesis enzymes, DGAT1 and DGAT2, in murine adipose tissue (ADGAT DKO mice). Despite markedly reduced TG storage, ADGAT DKO mice maintained ample adipose tissue endocrine function and surprisingly did not develop metabolic perturbations, even when fed a high-fat diet, owing to increased energy expenditure and beiging of white adipose tissue. These findings, thus, reveal that adipose tissue performs TG storage and endocrine functions largely independently from each other.</jats:sec

Mice lacking triglyceride synthesis enzymes in adipose tissue are resistant to diet-induced obesity

Triglycerides (TGs) in adipocytes provide the major stores of metabolic energy in the body. Optimal amounts of TG stores are desirable as insufficient capacity to store TG, as in lipodystrophy, or exceeding the capacity for storage, as in obesity, results in metabolic disease. We hypothesized that mice lacking TG storage in adipocytes would result in excess TG storage in cell types other than adipocytes and severe lipotoxicity accompanied by metabolic disease. To test this hypothesis, we selectively deleted both TG synthesis enzymes, DGAT1 and DGAT2, in adipocytes (ADGAT DKO mice). As expected with depleted energy stores, ADGAT DKO mice did not tolerate fasting well and, with prolonged fasting, entered torpor. However, ADGAT DKO mice were unexpectedly otherwise metabolically healthy and did not accumulate TGs ectopically or develop associated metabolic perturbations, even when fed a high-fat diet. The favorable metabolic phenotype resulted from activation of energy expenditure, in part via BAT (brown adipose tissue) activation and beiging of white adipose tissue. Thus, the ADGAT DKO mice provide a fascinating new model to study the coupling of metabolic energy storage to energy expenditure