Lipid Droplets in Cancer.

Lipid droplets have a unique structure among organelles consisting of a dense hydrophobic core of neutral lipids surrounded by a single layer of phospholipids decorated with various proteins. Often labeled merely as passive fat storage repositories, they in fact have a remarkably dynamic life cycle. Being formed within the endoplasmic reticulum membrane, lipid droplets rapidly grow, shrink, traverse the cytosol, and engage in contacts with other organelles to exchange proteins and lipids. Their lipid and protein composition changes dynamically in response to cellular states and nutrient availability. Remarkably, their biogenesis is induced when cells experience various forms of nutrient, energy, and redox imbalances, including lipid excess and complete nutrient deprivation. Cancer cells are continuously exposed to nutrient and oxygen fluctuations and have the capacity to switch between alternative nutrient acquisition and metabolic pathways in order to strive even during severe stress. Their supply of lipids is ensured by a series of nutrient uptake and scavenging mechanisms, upregulation of de novo lipid synthesis, repurposing of their structural lipids via enzymatic remodeling, or lipid recycling through autophagy. Importantly, most of these pathways of lipid acquisition converge at lipid droplets, which combine different lipid fluxes and control their usage based on specific cellular needs. It is thus not surprising that lipid droplet breakdown is an elaborately regulated process that occurs via a complex interplay of neutral lipases and autophagic degradation. Cancer cells employ lipid droplets to ensure energy production and redox balance, modulate autophagy, drive membrane synthesis, and control its composition, thereby minimizing stress and fostering tumor progression. As regulators of (poly)unsaturated fatty acid trafficking, lipid droplets are also emerging as modulators of lipid peroxidation and sensitivity to ferroptosis. Clearly, dysregulated lipid droplet turnover may also be detrimental to cancer cells, which should provide potential therapeutic opportunities in the future. In this review, we explore how lipid droplets consolidate lipid acquisition and trafficking pathways in order to match lipid supply with the requirements for cancer cell survival, growth, and metastasis.

The expanding organelle lipidomes: current knowledge and challenges.

Lipids in cell membranes and subcellular compartments play essential roles in numerous cellular processes, such as energy production, cell signaling and inflammation. A specific organelle lipidome is characterized by lipid synthesis and metabolism, intracellular trafficking, and lipid homeostasis in the organelle. Over the years, considerable effort has been directed to the identification of the lipid fingerprints of cellular organelles. However, these fingerprints are not fully characterized due to the large variety and structural complexity of lipids and the great variability in the abundance of different lipid species. The process becomes even more challenging when considering that the lipidome differs in health and disease contexts. This review summarizes the information available on the lipid composition of mammalian cell organelles, particularly the lipidome of the nucleus, mitochondrion, endoplasmic reticulum, Golgi apparatus, plasma membrane and organelles in the endocytic pathway. The lipid compositions of extracellular vesicles and lamellar bodies are also described. In addition, several examples of subcellular lipidome dynamics under physiological and pathological conditions are presented. Finally, challenges in mapping organelle lipidomes are discussed.

Synergy between 15-lipoxygenase and secreted PLA2 promotes inflammation by formation of TLR4 agonists from extracellular vesicles.

Damage-associated endogenous molecules induce innate immune response, thus making sterile inflammation medically relevant. Stress-derived extracellular vesicles (stressEVs) released during oxidative stress conditions were previously found to activate Toll-like receptor 4 (TLR4), resulting in expression of a different pattern of immune response proteins in comparison to lipopolysaccharide (LPS), underlying the differences between pathogen-induced and sterile inflammation. Here we report that synergistic activities of 15-lipoxygenase (15-LO) and secreted phospholipase A2 (sPLA2) are needed for the formation of TLR4 agonists, which were identified as lysophospholipids (lysoPLs) with oxidized unsaturated acyl chain. Hydroxy, hydroperoxy, and keto products of 2-arachidonoyl-lysoPI oxidation by 15-LO were identified by mass spectrometry (MS), and they activated the same gene pattern as stressEVs. Extracellular PLA2 activity was detected in the synovial fluid from rheumatoid arthritis and gout patients. Furthermore, injection of sPLA2 promoted K/BxN serum-induced arthritis in mice, whereby ankle swelling was partially TLR4 dependent. Results confirm the role of oxidized lysoPL of stressEVs in sterile inflammation that promotes chronic diseases. Both 15-LO and sPLA2 enzymes are induced during inflammation, which opens the opportunity for therapy without compromising innate immunity against pathogens.

Oleic Acid Protects Endothelial Cells from Silica-Coated Superparamagnetic Iron Oxide Nanoparticles (SPIONs)-Induced Oxidative Stress and Cell Death.

Superparamagnetic iron oxide nanoparticles (SPIONs) have great potential for use in medicine, but they may cause side effects due to oxidative stress. In our study, we investigated the effects of silica-coated SPIONs on endothelial cells and whether oleic acid (OA) can protect the cells from their harmful effects. We used viability assays, flow cytometry, infrared spectroscopy, fluorescence microscopy, and transmission electron microscopy. Our results show that silica-coated SPIONs are internalized by endothelial cells, where they increase the amount of reactive oxygen species (ROS) and cause cell death. Exposure to silica-coated SPIONs induced accumulation of lipid droplets (LD) that was not dependent on diacylglycerol acyltransferase (DGAT)-mediated LD biogenesis, suggesting that silica-coated SPIONs suppress LD degradation. Addition of exogenous OA promoted LD biogenesis and reduced SPION-dependent increases in oxidative stress and cell death. However, exogenous OA protected cells from SPION-induced cell damage even in the presence of DGAT inhibitors, implying that LDs are not required for the protective effect of exogenous OA. The molecular phenotype of the cells determined by Fourier transform infrared spectroscopy confirmed the destructive effect of silica-coated SPIONs and the ameliorative role of OA in the case of oxidative stress. Thus, exogenous OA protects endothelial cells from SPION-induced oxidative stress and cell death independent of its incorporation into triglycerides.

Astrocytes in stress accumulate lipid droplets.

When the brain is in a pathological state, the content of lipid droplets (LDs), the lipid storage organelles, is increased, particularly in glial cells, but rarely in neurons. The biology and mechanisms leading to LD accumulation in astrocytes, glial cells with key homeostatic functions, are poorly understood. We imaged fluorescently labeled LDs by microscopy in isolated and brain tissue rat astrocytes and in glia-like cells in Drosophila brain to determine the (sub)cellular localization, mobility, and content of LDs under various stress conditions characteristic for brain pathologies. LDs exhibited confined mobility proximal to mitochondria and endoplasmic reticulum that was attenuated by metabolic stress and by increased intracellular Ca2+ , likely to enhance the LD-organelle interaction imaged by electron microscopy. When de novo biogenesis of LDs was attenuated by inhibition of DGAT1 and DGAT2 enzymes, the astrocyte cell number was reduced by ~40%, suggesting that in astrocytes LD turnover is important for cell survival and/or proliferative cycle. Exposure to noradrenaline, a brain stress response system neuromodulator, and metabolic and hypoxic stress strongly facilitated LD accumulation in astrocytes. The observed response of stressed astrocytes may be viewed as a support for energy provision, but also to be neuroprotective against the stress-induced lipotoxicity.

Lipid Droplets and the Management of Cellular Stress.

Lipid droplets are cytosolic fat storage organelles present in most eukaryotic cells. Long regarded merely as inert fat reservoirs, they are now emerging as major regulators of cellular metabolism. They act as hubs that coordinate the pathways of lipid uptake, distribution, storage, and use in the cell. Recent studies have revealed that they are also essential components of the cellular stress response. One of the hallmark characteristics of lipid droplets is their capacity to buffer excess lipids and to finely tune their subsequent release based on specific cellular requirements. This simple feature of lipid droplet biology, buffering and delayed release of lipids, forms the basis for their pleiotropic roles in the cellular stress response. In stressed cells, lipid droplets maintain energy and redox homeostasis and protect against lipotoxicity by sequestering toxic lipids into their neutral lipid core. Their mobility and dynamic interactions with mitochondria enable an efficient delivery of fatty acids for optimal energy production. Lipid droplets are also involved in the maintenance of membrane and organelle homeostasis by regulating membrane composition, preventing lipid peroxidation and removing damaged proteins and lipids. Finally, they also engage in a symbiotic relationship with autophagy and act as reservoirs of bioactive lipids that regulate inflammation and immunity. Thus, lipid droplets are central managers of lipid metabolism that function as safeguards against various types of cellular stress.

Lipid droplets induced by secreted phospholipase A2 and unsaturated fatty acids protect breast cancer cells from nutrient and lipotoxic stress.

Cancer cells driven by the Ras oncogene scavenge unsaturated fatty acids (FAs) from their environment to counter nutrient stress. The human group X secreted phospholipase A2 (hGX sPLA2) releases FAs from membrane phospholipids, stimulates lipid droplet (LD) biogenesis in Ras-driven triple-negative breast cancer (TNBC) cells and enables their survival during starvation. Here we examined the role of LDs, induced by hGX sPLA2 and unsaturated FAs, in protection of TNBC cells against nutrient stress. We found that hGX sPLA2 releases a mixture of unsaturated FAs, including ω-3 and ω-6 polyunsaturated FAs (PUFAs), from TNBC cells. Starvation-induced breakdown of LDs induced by low micromolar concentrations of unsaturated FAs, including PUFAs, was associated with protection from cell death. Interestingly, adipose triglyceride lipase (ATGL) contributed to LD breakdown during starvation, but it was not required for the pro-survival effects of hGX sPLA2 and unsaturated FAs. High micromolar concentrations of PUFAs, but not OA, induced oxidative stress-dependent cell death in TNBC cells. Inhibition of triacylglycerol (TAG) synthesis suppressed LD biogenesis and potentiated PUFA-induced cell damage. On the contrary, stimulation of LD biogenesis by hGX sPLA2 and suppression of LD breakdown by ATGL depletion reduced PUFA-induced oxidative stress and cell death. Finally, lipidomic analyses revealed that sequestration of PUFAs in LDs by sPLA2-induced TAG remodelling and retention of PUFAs in LDs by inhibition of ATGL-mediated TAG lipolysis protect from PUFA lipotoxicity. LDs are thus antioxidant and pro-survival organelles that guard TNBC cells against nutrient and lipotoxic stress and emerge as attractive targets for novel therapeutic interventions.

Engineering recombinant Lactococcus lactis as a delivery vehicle for BPC-157 peptide with antioxidant activities.

Lactic acid bacteria (LAB) are attractive hosts for the expression of heterologous proteins and can be engineered to deliver therapeutic proteins or peptides to mucosal surfaces. The gastric stable pentadecapeptide BPC-157 is able to prevent and treat gastrointestinal inflammation by reducing the production of reactive oxygen species (ROS). In this study, we used LAB Lactococcus lactis as a vector to deliver BPC-157 by surface display and trypsin shedding or by secretion to the growth medium. Surface display of BPC-157 was achieved by fusing it with basic membrane protein A (BmpA) or with the peptidoglycan binding domain of AcmA and Usp45 secretion signal. While the expression of BmpA-fusion proteins was higher than that of AcmA/Usp45-fusion protein, the surface display ability of BPC-157 was approximately 14-fold higher with AcmA/Usp45-fusion protein. Release of BPC-157 from the bacterial surface or from isolated fusion proteins by trypsinization was demonstrated with anti-BPC-157 antibodies or by mass spectrometry. The concentration of BPC-157 delivered by surface display via AcmA/Usp45-fusion was 30 ng/ml. This increased to 117 ng/ml by Usp45 signal-mediated secretion, making the latter the most effective lactococcal delivery approach for BPC-157. Secreted BPC-157 significantly decreased ROS production in 149BR fibroblast cell model, suggesting its potential benefit in the treatment of intestinal inflammations. Additionally, a comparison of different modes of small peptide delivery by L. lactis, performed in the present study, will facilitate the future use of L. lactis as peptide delivery vehicle.

Lipid Droplets in Cancer: Guardians of Fat in a Stressful World.

Cancer cells possess remarkable abilities to adapt to adverse environmental conditions. Their survival during severe nutrient and oxidative stress depends on their capacity to acquire extracellular lipids and the plasticity of their mechanisms for intracellular lipid synthesis, mobilisation, and recycling. Lipid droplets, cytosolic fat storage organelles present in most cells from yeast to men, are emerging as major regulators of lipid metabolism, trafficking, and signalling in various cells and tissues exposed to stress. Their biogenesis is induced by nutrient and oxidative stress and they accumulate in various cancers. Lipid droplets act as switches that coordinate lipid trafficking and consumption for different purposes in the cell, such as energy production, protection against oxidative stress or membrane biogenesis during rapid cell growth. They sequester toxic lipids, such as fatty acids, cholesterol and ceramides, thereby preventing lipotoxic cell damage and engage in a complex relationship with autophagy. Here, we focus on the emerging mechanisms of stress-induced lipid droplet biogenesis; their roles during nutrient, lipotoxic, and oxidative stress; and the relationship between lipid droplets and autophagy. The recently discovered principles of lipid droplet biology can improve our understanding of the mechanisms that govern cancer cell adaptability and resilience to stress.

Lipid Droplet Formation in HeLa Cervical Cancer Cells Depends on Cell Density and the Concentration of Exogenous Unsaturated Fatty Acids.

Cytosolic lipid droplets (LDs) store excess fatty acids (FAs) in the form of neutral lipids and prevent starvation-induced cancer cell death. Here we studied the ability of mono- and polyunsaturated FAs to affect LD formation and survival in HeLa cervical cancer cells. We found that the LD content in HeLa cells increases with cell density, but it decreases in MDA-MB-231 breast cancer cells. Exogenously-added unsaturated FAs, including oleic (OA), linoleic (LA), arachidonic (AA), eicosapentaenoic (EPA) and docosahexaenoic acid (DHA) displayed a similar ability to alter LD formation in HeLa cells. There was a dual, concentration-dependent effect on neutral lipid accumulation: low micromolar concentrations of LA, AA and DHA reduced, while all FAs induced LD formation at higher concentrations. In serum starved He-La cells, OA stimulated LD formation, but, contrary to expectations, it promoted cell death. Our results reveal a link between cell population density and LD formation in HeLa cells and show that unsaturated FAs may both suppress or stimulate LD formation. This dynamic regulation of LD content must be accounted for when studying the effects of lipids and lipid metabolism-targeting drugs on LD metabolism in HeLa cells.

Disintegrins from the Venom of Vipera ammodytes ammodytes Efficiently Inhibit Migration of Breast Cancer Cells.

Integrins are plasma membrane proteins, whose dysfunction frequently results in cancer pathology, and therefore they represent important targets of anti-tumour therapy. Snake venoms are a rich source of disintegrins (Dis), proteins that specifically bind integrins and thus interfere with their functions. In an attempt to discover new molecules for treatment of breast cancer, the major type of cancer in women, we isolated a dimeric Dis (Vaa-Dis) from the venom of the nosehorned viper. By cell viability testing we demonstrated that 50 nM and higher concentrations of Vaa-Dis were toxic to highly invasive human breast adenocarcinoma cell line MDA-MB-231. Wound-healing assay revealed that already at one order of magnitude lower concentrations Vaa-Dis efficiently inhibited MDA-MB-231 cell migration. This exposed a promising anti-metastatic potential of Vaa-Dis and a good perspective of these natural snake venom proteins for further research and development towards the application in breast cancer treatment.

Phospholipase A2 group IIA is elevated in endometriomas but not in peritoneal fluid and serum of ovarian endometriosis patients.

Our previous gene expression analysis identified phospholipase A2 group IIA (PLA2G2A) as a potential biomarker of ovarian endometriosis. The aim of this study was to evaluate PLA2G2A mRNA and protein levels in tissue samples (endometriomas and normal endometrium) and in serum and peritoneal fluid of ovarian endometriosis patients and control women. One-hundred and sixteen women were included in this study: the case group included 70 ovarian endometriosis patients, and the control group included 38 healthy women and 8 patients with benign ovarian cysts. We observed 41.6-fold greater PLA2G2A mRNA levels in endometrioma tissue, compared to normal endometrium tissue. Using Western blotting, PLA2G2A was detected in all samples of endometriomas, but not in normal endometrium, and immunohistochemistry showed PLA2G2A-specific staining in epithelial cells of endometrioma paraffin sections. However, there were no significant differences in PLA2G2A levels between cases and controls according to ELISA of peritoneal fluid (6.0 ± 4.4 ng/ml, 6.6 ± 4.3 ng/ml; p = 0.5240) and serum (2.9 ± 2.1 ng/ml, 3.1 ± 2.2 ng/ml; p = 0.7989). Our data indicate that PLA2G2A is implicated in the pathophysiology of ovarian endometriosis, but that it cannot be used as a diagnostic biomarker.

Secreted phospholipases A2are differentially expressed and epigenetically silenced in human breast cancer cells.

Secreted phospholipases A2 (sPLA2s) have recently been associated with several cancers, but their role in breast cancer is unknown. Here we demonstrate that mRNA expression of group IIA, III and X sPLA2s differs both in vivo in tumour biopsies and in breast cancer cells in vitro. Their expression is differentially regulated by DNA methylation and histone acetylation and, significantly, all three genes are silenced in aggressive triple negative cells due to both mechanisms. The transcription start site promoter region and the upstream CpG islands, exclusive to the group X sPLA2 gene, have variable roles in the regulation of sPLA2 expression. Our results suggest that the differential expression of hGIIA, hGIII and hGX sPLA2s in breast cancer cells is a consequence of various degrees of epigenetic silencing due to DNA hypermethylation and histone deacetylation.

The efficient magneto-mechanical actuation of cancer cells using a very low concentration of non-interacting ferrimagnetic hexaferrite nanoplatelets.

Magneto-mechanical actuation (MMA) using the low-frequency alternating magnetic fields (AMFs) of magnetic nanoparticles internalized into cancer cells can be used to irreparably damage these cells. However, nanoparticles in cells usually agglomerate, thus greatly augmenting the delivered force compared to single nanoparticles. Here, we demonstrate that MMA also decreases the cell viability, with the MMA mediated by individual, non-interacting nanoparticles. The effect was demonstrated with ferrimagnetic (i.e., permanently magnetic) barium-hexaferrite nanoplatelets (NPLs, ∼50 nm wide and 3 nm thick) with a unique, perpendicular orientation of the magnetization. Two cancer-cell lines (MDA-MB-231 and HeLa) are exposed to the NPLs in-vitro under different cell-culture conditions and actuated with a uniaxial AMF. TEM analyses show that only a small number of NPLs internalize in the cells, always situated in membrane-enclosed compartments of the endosomal-lysosomal system. Most compartments contain 1-2 NPLs and only seldom are the NPLs found in small groups, but never in close contact or mutually oriented. Even at low concentrations, the single NPLs reduce the cell viability when actuated with AMFs, which is further increased when the cells are in starvation conditions. These results pave the way for more efficient in-vivo MMA at very low particle concentrations.

Oxylipin profiling for clinical research: Current status and future perspectives.

Oxylipins are potent lipid mediators with increasing interest in clinical research. They are usually measured in systemic circulation and can provide a wealth of information regarding key biological processes such as inflammation, vascular tone, or blood coagulation. Although procedures still require harmonization to generate comparable oxylipin datasets, performing comprehensive profiling of circulating oxylipins in large studies is feasible and no longer restricted by technical barriers. However, it is essential to improve and facilitate the biological interpretation of complex oxylipin profiles to truly leverage their potential in clinical research. This requires regular updating of our knowledge about the metabolism and the mode of action of oxylipins, and consideration of all factors that may influence circulating oxylipin profiles independently of the studied disease or condition. This review aims to provide the readers with updated and necessary information regarding oxylipin metabolism, their different forms in systemic circulation, the current limitations in deducing oxylipin cellular effects from in vitro bioactivity studies, the biological and technical confounding factors needed to consider for a proper interpretation of oxylipin profiles.

Group X secreted phospholipase A(2) induces lipid droplet formation and prolongs breast cancer cell survival.

BACKGROUND: Alterations in lipid metabolism are inherent to the metabolic transformations that support tumorigenesis. The relationship between the synthesis, storage and use of lipids and their importance in cancer is poorly understood. The human group X secreted phospholipase A2 (hGX sPLA2) releases fatty acids (FAs) from cell membranes and lipoproteins, but its involvement in the regulation of cellular FA metabolism and cancer is not known. RESULTS: Here we demonstrate that hGX sPLA2 induces lipid droplet (LD) formation in invasive breast cancer cells, stimulates their proliferation and prevents their death on serum deprivation. The effects of hGX sPLA2 are shown to be dependent on its enzymatic activity, are mimicked by oleic acid and include activation of protein kinase B/Akt, a cell survival signaling kinase. The hGX sPLA2-stimulated LD biogenesis is accompanied by AMP-activated protein kinase (AMPK) activation, up-regulation of FA oxidation enzymes and the LD-coating protein perilipin 2, and suppression of lipogenic gene expression. Prolonged activation of AMPK inhibited hGX sPLA2-induced LD formation, while etomoxir, an inhibitor of FA oxidation, abrogated both LD formation and cell survival. The hGX sPLA2-induced changes in lipid metabolism provide a minimal immediate proliferative advantage during growth under optimal conditions, but they confer to the breast cancer cells a sustained ability to resist apoptosis during nutrient and growth factor limitation. CONCLUSION: Our results identify hGX sPLA2 as a novel modulator of lipid metabolism that promotes breast cancer cell growth and survival by stimulating LD formation and FA oxidation.

Differential inhibition of LINE1 and LINE2 retrotransposition by vertebrate AID/APOBEC proteins.

BACKGROUND: The role of AID/APOBEC proteins in the mammalian immune response against retroviruses and retrotransposons is well established. G to A hypermutations, the hallmark of their cytidine deaminase activity, are present in several mammalian retrotransposons. However, the role of AID/APOBEC proteins in non-mammalian retroelement restriction is not completely understood. RESULTS: Here we provide the first evidence of anti-retroelement activity of a reptilian APOBEC protein. The green anole lizard A1 protein displayed potent DNA mutator activity and inhibited ex vivo retrotransposition of LINE1 and LINE2 ORF1 protein encoding elements, displaying a mechanism of action similar to that of the human A1 protein. In contrast, the human A3 proteins did not require ORF1 protein to inhibit LINE retrotransposition, suggesting a differential mechanism of anti-LINE action of A1 proteins, which emerged in amniotes, and A3 proteins, exclusive to placental mammals. In accordance, genomic analyses demonstrate differential G to A DNA editing of LINE retrotransposons in the lizard genome, which is also the first evidence for G to A DNA editing in non-mammalian genomes. CONCLUSION: Our data suggest that vertebrate APOBEC proteins differentially inhibit the retrotransposition of LINE elements and that the anti-retroelement activity of APOBEC proteins predates mammals.

Recombinant human erythropoietin alters gene expression and stimulates proliferation of MCF-7 breast cancer cells.

BACKGROUND: Functional erythropoietin (EPO) signaling is not specific only to erythroid lineages and has been confirmed in several solid tumors, including breast. Three different isoforms of erythropoietin receptor (EPOR) have been reported, the soluble (EPOR-S) and truncated (EPOR-T) forms acting antagonistically to the functional EPOR. In this study, we investigated the effect of human recombinant erythropoietin (rHuEPO) on cell proliferation, early gene response and the expression of EPOR isoforms in the MCF-7 breast cancer cell line. MATERIALS AND METHODS: The MCF-7 cells were cultured with or without rHuEPO for 72 h or 10 weeks and assessed for their growth characteristics, expression of early response genes and different EPOR isoforms. The expression profile of EPOR and EPOR-T was determined in a range of breast cancer cell lines and compared with their invasive properties. RESULTS: MCF-7 cell proliferation after rHuEPO treatment was dependent on the time of treatment and the concentration used. High rHuEPO concentrations (40 U/ml) stimulated cell proliferation independently of a preceding long-term exposure of MCF-7 cells to rHuEPO, while lower concentrations increased MCF-7 proliferation only after 10 weeks of treatment. Gene expression analysis showed activation of EGR1 and FOS, confirming the functionality of EPOR. rHuEPO treatment also slightly increased the expression of the functional EPOR isoform, which, however, persisted throughout the 10 weeks of treatment. The expression levels of EPOR-T were not influenced. There were no correlations between EPOR expression and the invasiveness of MCF-7, MDA-MB-231, Hs578T, Hs578Bst, SKBR3, T-47D and MCF-10A cell lines. CONCLUSIONS: rHuEPO modulates MCF-7 cell proliferation in time- and concentration-dependent manner. We confirmed EGR1, FOS and EPOR as transcription targets of the EPO-EPOR signaling loop, but could not correlate the expression of different EPOR isoforms with the invasiveness of breast cancer cell lines.

Lipid droplets and polyunsaturated fatty acid trafficking: Balancing life and death.

Lipid droplets are fat storage organelles ubiquitously distributed across the eukaryotic kingdom. They have a central role in regulating lipid metabolism and undergo a dynamic turnover of biogenesis and breakdown to meet cellular requirements for fatty acids, including polyunsaturated fatty acids. Polyunsaturated fatty acids esterified in membrane phospholipids define membrane fluidity and can be released by the activity of phospholipases A2 to act as ligands for nuclear receptors or to be metabolized into a wide spectrum of lipid signaling mediators. Polyunsaturated fatty acids in membrane phospholipids are also highly susceptible to lipid peroxidation, which if left uncontrolled leads to ferroptotic cell death. On the one hand, lipid droplets act as antioxidant organelles that control polyunsaturated fatty acid storage in triglycerides in order to reduce membrane lipid peroxidation, preserve organelle function and prevent cell death, including ferroptosis. On the other hand, lipid droplet breakdown fine-tunes the delivery of polyunsaturated fatty acids into metabolic and signaling pathways, but unrestricted lipid droplet breakdown may also lead to the release of lethal levels of polyunsaturated fatty acids. Precise regulation of lipid droplet turnover is thus essential for polyunsaturated fatty acid distribution and cellular homeostasis. In this review, we focus on emerging aspects of lipid droplet-mediated regulation of polyunsaturated fatty acid trafficking, including the management of membrane lipid peroxidation, ferroptosis and lipid mediator signaling.

The Combined Inhibition of Autophagy and Diacylglycerol Acyltransferase-Mediated Lipid Droplet Biogenesis Induces Cancer Cell Death during Acute Amino Acid Starvation.

Lipid droplets (LDs) are dynamic organelles involved in the management of fatty acid trafficking and metabolism. Recent studies suggest that autophagy and LDs serve complementary roles in the protection against nutrient stress, but the autophagy-LD interplay in cancer cells is not well understood. Here, we examined the relationship between autophagy and LDs in starving HeLa cervical cancer- and MDA-MB-231 breast cancer cells. We found that acute amino acid depletion induces autophagy and promotes diacylglycerol acyltransferase 1 (DGAT1)-mediated LD accumulation in HeLa cells. Inhibition of autophagy via late-stage autophagy inhibitors, or by knocking down autophagy-related 5 (ATG5), reduced LD accumulation in amino acid-starved cancer cells, suggesting that autophagy contributes to LD biogenesis. On the contrary, knockdown of adipose triglyceride lipase (ATGL) increased LD accumulation, suggesting that LD breakdown is mediated by lipolysis under these conditions. Concurrent inhibition of autophagy by silencing ATG5 and of LD biogenesis using DGAT inhibitors was effective in killing starving HeLa cells, whereas cell survival was not compromised by suppression of ATGL-mediated lipolysis. Autophagy-dependent LD biogenesis was also observed in the aggressive triple-negative MDA-MB-231 breast cancer cells deprived of amino acids, but these cells were not sensitized to starvation by the combined inhibition of LD biogenesis and autophagy. These findings reveal that while targeting autophagy-driven and DGAT-mediated LD biogenesis reduces the resilience of HeLa cervical cancer cells to amino acid deprivation, this strategy may not be successful in other cancer cell types.