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Lipid droplets are dynamic organelles that store neutral lipids, serve the metabolic needs of cells, and sequester lipids to prevent lipotoxicity and membrane damage. Here we review the current understanding of the mechanisms of lipid droplet biogenesis and turnover, the transfer of lipids and metabolites at membrane contact sites, and the role of lipid droplets in regulating fatty acid flux in lipotoxicity and cell death., (© 2024. Springer Nature Limited.)

Despite the key roles of perilipin-2 (PLIN2) in governing lipid droplet (LD) metabolism, the mechanisms that regulate PLIN2 levels remain incompletely understood. Here, we leverage a set of genome-edited human PLIN2 reporter cell lines in a series of CRISPR-Cas9 loss-of-function screens, identifying genetic modifiers that influence PLIN2 expression and post-translational stability under different metabolic conditions and in different cell types. These regulators include canonical genes that control lipid metabolism as well as genes involved in ubiquitination, transcription, and mitochondrial function. We further demonstrate a role for the E3 ligase MARCH6 in regulating triacylglycerol biosynthesis, thereby influencing LD abundance and PLIN2 stability. Finally, our CRISPR screens and several published screens provide the foundation for CRISPRlipid (http://crisprlipid.org), an online data commons for lipid-related functional genomics data. Our study identifies mechanisms of PLIN2 and LD regulation and provides an extensive resource for the exploration of LD biology and lipid metabolism., Competing Interests: Declaration of interests J.A.O. is a member of the scientific advisory board for Vicinitas Therapeutics and has patent applications related to ferroptosis. M.K. is an inventor on a US patent related to CRISPRi and CRISPRa screening; serves on the scientific advisory boards of Engine Biosciences, Casma Therapeutics, Cajal Neuroscience and Alector; and is a consultant to Modulo Bio and Recursion Therapeutics. F.F. and M.A.N.’s participation in this project was part of a competitive contract awarded to Data Tecnica International LLC by the National Institutes of Health to support open science research. M.A.N. also currently serves on the scientific advisory board for Clover Therapeutics and is an advisor to Neuron23 Inc. M.C.B. has outside interest in DEM Biopharma., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

Lipid droplets (LDs) are organelles of cellular lipid storage with fundamental roles in energy metabolism and cell membrane homeostasis. There has been an explosion of research into the biology of LDs, in part due to their relevance in diseases of lipid storage, such as atherosclerosis, obesity, type 2 diabetes, and hepatic steatosis. Consequently, there is an increasing need for a resource that combines datasets from systematic analyses of LD biology. Here, we integrate high-confidence, systematically generated human, mouse, and fly data from studies on LDs in the framework of an online platform named the "Lipid Droplet Knowledge Portal" (https://lipiddroplet.org/). This scalable and interactive portal includes comprehensive datasets, across a variety of cell types, for LD biology, including transcriptional profiles of induced lipid storage, organellar proteomics, genome-wide screen phenotypes, and ties to human genetics. This resource is a powerful platform that can be utilized to identify determinants of lipid storage., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2022 The Authors. Published by Elsevier Inc. All rights reserved.)

Lipid droplets are endoplasmic reticulum-derived neutral lipid storage organelles that play critical roles in cellular lipid and energy homeostasis. Here, we present a protocol for the identification of high-confidence lipid droplet proteomes in a cell culture model. This approach overcomes limitations associated with standard biochemical fractionation techniques, employing an engineered ascorbate peroxidase (APEX2) to biotinylate endogenous lipid droplet proteins in living cells for subsequent purification and identification by proteomics. For complete details on the use and execution of this protocol, please refer to Bersuker et al. (2018)., Competing Interests: The authors declare no competing interests., (© 2021 The Authors.)

Lipid droplets (LDs) are endoplasmic reticulum-derived organelles that consist of a core of neutral lipids encircled by a phospholipid monolayer decorated with proteins. As hubs of cellular lipid and energy metabolism, LDs are inherently involved in the etiology of prevalent metabolic diseases such as obesity and nonalcoholic fatty liver disease. The functions of LDs are regulated by a unique set of associated proteins, the LD proteome, which includes integral membrane and peripheral proteins. These proteins control key activities of LDs such as triacylglycerol synthesis and breakdown, nutrient sensing and signal integration, and interactions with other organelles. Here we review the mechanisms that regulate the composition of the LD proteome, such as pathways that mediate selective and bulk LD protein degradation and potential connections between LDs and cellular protein quality control.

Lipid droplets (LDs) are neutral lipid storage organelles assembled at the endoplasmic reticulum (ER). A new study reveals that the high membrane curvature of ER tubules catalyzes the nucleation of a neutral lipid lens, an early step in LD biogenesis., (Copyright © 2020 Elsevier Inc. All rights reserved.)

In this issue of Developmental Cell, Chorlay et al. (2019) provide evidence that asymmetric membrane surface tension determines the directionality of lipid droplet (LD) emergence. Furthermore, phospholipid synthesis "refills" the outer leaflet of the endoplasmic reticulum (ER) membrane to maintain cytosolic LD emergence and prevent disruptions to ER homeostasis., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Lipid droplets (LDs) are hubs for lipid metabolism that form membrane contact sites with multiple organelles. In this issue, Hariri et al. (2019. J. Cell Biol. https://doi.org/10.1083/jcb.201808119) reveal the functions of Mdm1-mediated endoplasmic reticulum (ER)-LD tethering in yeast and Datta et al. (2019. J. Cell Biol. https://doi.org/10.1083/jcb.201808133) identify a role for the Mdm1 orthologue, Snx14, as an ER-LD tether that regulates lipid metabolism in human cells., (© 2019 Nguyen and Olzmann.)

Lipid droplets are storage organelles at the centre of lipid and energy homeostasis. They have a unique architecture consisting of a hydrophobic core of neutral lipids, which is enclosed by a phospholipid monolayer that is decorated by a specific set of proteins. Originating from the endoplasmic reticulum, lipid droplets can associate with most other cellular organelles through membrane contact sites. It is becoming apparent that these contacts between lipid droplets and other organelles are highly dynamic and coupled to the cycles of lipid droplet expansion and shrinkage. Importantly, lipid droplet biogenesis and degradation, as well as their interactions with other organelles, are tightly coupled to cellular metabolism and are critical to buffer the levels of toxic lipid species. Thus, lipid droplets facilitate the coordination and communication between different organelles and act as vital hubs of cellular metabolism.

Lipid droplets (LDs) are ubiquitous lipid storage organelles composed of a neutral lipid core surrounded by a phospholipid monolayer that is decorated with integral and peripheral proteins. Accurate identification of LD proteins using biochemical fractionation methods has been challenging due to the presence of contaminant proteins from co-fractionating organelles. Here, we describe a method to identify high-confidence LD proteomes that employs an engineered ascorbate peroxidase (APEX2) to induce spatially and temporally restricted biotinylation of LD proteins. This proximity labeling method can be broadly applied to define the composition of the LD proteome in any cultured cell line and can be utilized to examine LD proteome dynamics.

Lipid droplet (LD) functions are regulated by a complement of integral and peripheral proteins that associate with the bounding LD phospholipid monolayer. Defining the composition of the LD proteome has remained a challenge due to the presence of contaminating proteins in LD-enriched buoyant fractions. To overcome this limitation, we developed a proximity labeling strategy that exploits LD-targeted APEX2 to biotinylate LD proteins in living cells. Application of this approach to two different cell types identified the vast majority of previously validated LD proteins, excluded common contaminating proteins, and revealed new LD proteins. Moreover, quantitative analysis of LD proteome dynamics uncovered a role for endoplasmic reticulum-associated degradation in controlling the composition of the LD proteome. These data provide an important resource for future LD studies and demonstrate the utility of proximity labeling to study the regulation of LD proteomes., (Copyright © 2017 Elsevier Inc. All rights reserved.)

Lipid droplets (LDs) are ubiquitous, endoplasmic reticulum (ER)-derived organelles that mediate the sequestration of neutral lipids (e.g. triacylglycerol and sterol esters), providing a dynamic cellular storage depot for rapid lipid mobilization in response to increased cellular demands. LDs have a unique ultrastructure, consisting of a core of neutral lipids encircled by a phospholipid monolayer that is decorated with integral and peripheral proteins. The LD proteome contains numerous lipid metabolic enzymes, regulatory scaffold proteins, proteins involved in LD clustering and fusion, and other proteins of unknown functions. The cellular role of LDs is inherently determined by the composition of its proteome and alteration of the LD protein coat provides a powerful mechanism to adapt LDs to fluctuating metabolic states. Here, we review the current understanding of the molecular mechanisms that govern LD protein targeting and degradation. This article is part of a Special Issue entitled: Recent Advances in Lipid Droplet Biology edited by Rosalind Coleman and Matthijs Hesselink., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Lipid droplets (LDs) provide an "on-demand" source of fatty acids (FAs) that can be mobilized in response to fluctuations in nutrient abundance. Surprisingly, the amount of LDs increases during prolonged periods of nutrient deprivation. Why cells store FAs in LDs during an energy crisis is unknown. Our data demonstrate that mTORC1-regulated autophagy is necessary and sufficient for starvation-induced LD biogenesis. The ER-resident diacylglycerol acyltransferase 1 (DGAT1) selectively channels autophagy-liberated FAs into new, clustered LDs that are in close proximity to mitochondria and are lipolytically degraded. However, LDs are not required for FA delivery to mitochondria but instead function to prevent acylcarnitine accumulation and lipotoxic dysregulation of mitochondria. Our data support a model in which LDs provide a lipid buffering system that sequesters FAs released during the autophagic degradation of membranous organelles, reducing lipotoxicity. These findings reveal an unrecognized aspect of the cellular adaptive response to starvation, mediated by LDs., (Copyright © 2017 Elsevier Inc. All rights reserved.)

The endoplasmic reticulum (ER) mediates the folding, maturation, and deployment of the secretory proteome. Proteins that fail to achieve their native conformation are retained in the ER and targeted for clearance by ER-associated degradation (ERAD), a sophisticated process that mediates the ubiquitin-dependent delivery of substrates to the 26S proteasome for proteolysis. Recent findings indicate that inhibition of long-chain acyl-CoA synthetases with triacsin C, a fatty acid analogue, impairs lipid droplet (LD) biogenesis and ERAD, suggesting a role for LDs in ERAD. However, whether LDs are involved in the ERAD process remains an outstanding question. Using chemical and genetic approaches to disrupt diacylglycerol acyltransferase (DGAT)-dependent LD biogenesis, we provide evidence that LDs are dispensable for ERAD in mammalian cells. Instead, our results suggest that triacsin C causes global alterations in the cellular lipid landscape that disrupt ER proteostasis by interfering with the glycan trimming and dislocation steps of ERAD. Prolonged triacsin C treatment activates both the IRE1 and PERK branches of the unfolded protein response and ultimately leads to IRE1-dependent cell death. These findings identify an intimate relationship between fatty acid metabolism and ER proteostasis that influences cell viability., (© 2017 To, Peterson, et al. This article is distributed by The American Society for Cell Biology under license from the author(s). Two months after publication it is available to the public under an Attribution–Noncommercial–Share Alike 3.0 Unported Creative Commons License (http://creativecommons.org/licenses/by-nc-sa/3.0).)

Lipid droplets (LDs) are neutral lipid storage organelles that provide a rapidly accessible source of fatty acids (FAs) for energy during periods of nutrient deprivation. Surprisingly, lipids released by the macroautophagic/autophagic breakdown of membranous organelles are packaged and stored in new LDs during periods of prolonged starvation. Why cells would store FAs during an energy crisis was unknown. In our recent study, we demonstrated that FAs released during MTORC1-regulated autophagy are selectively channeled by DGAT1 (diacylglycerol O-acyltransferase 1) into triacylglycerol (TAG)-rich LDs. These DGAT1-dependent LDs sequester FAs and prevent the accumulation of acylcarnitines, which otherwise directly disrupt mitochondrial integrity. Our findings establish LD biogenesis as a general cellular response to periods of high autophagic flux that provide a lipid buffering system to mitigate lipotoxic cellular damage.