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Drosophila melanogaster is a popular model organism to elucidate the molecular mechanisms that underlie the structure and function of the eye as well as the causes of retinopathies, aging, light-induced damage, or dietary deficiencies. Large-scale screens have isolated genes whose mutation causes morphological and functional ocular defects, which led to the discovery of key components of the phototransduction cascade. However, the proteome of the Drosophila eye is poorly characterized. Here, we used GeLC-MS/MS to quantify 3516 proteins, including the absolute (molar) quantities of 43 proteins in the eye of adult male Drosophila reared on standard laboratory food. This work provides a generic and expandable resource for further genetic, pharmacological, and dietary studies., (© 2023 The Authors. PROTEOMICS published by Wiley‐VCH GmbH.)

Organisms respond to dietary and environmental challenges by altering the molecular composition of their glycerolipids and glycerophospholipids (GPLs), which may favorably adjust the physicochemical properties of lipid membranes. However, how lipidome changes affect the membrane proteome and, eventually, the physiology of specific organs is an open question. We addressed this issue in Drosophila melanogaster , which is not able to synthesize sterols and polyunsaturated fatty acids but can acquire them from food. We developed a series of semisynthetic foods to manipulate the length and unsaturation of fatty acid moieties in GPLs and singled out proteins whose abundance is specifically affected by membrane lipid unsaturation in the Drosophila eye. Unexpectedly, we identified a group of proteins that have muscle-related functions and increased their abundances under unsaturated eye lipidome conditions. In contrast, the abundance of two stress response proteins, Turandot A and Smg5, is decreased by lipid unsaturation. Our findings could guide the genetic dissection of homeostatic mechanisms that maintain visual function when the eye is exposed to environmental and dietary challenges.

Lipidomic analysis in diverse biological settings has become a frequent tool to increase our understanding of the processes of life. Cellular lipids play important roles not only as being the main components of cellular membranes, but also in the regulation of cell homeostasis as lipid signaling molecules. Yeast has been harnessed for biomedical research based on its good conservation of genetics and fundamental cell organisation principles and molecular pathways. Further application in so-called humanised yeast models have been developed which take advantage of yeast as providing the basics of a living cell with full control over heterologous expression. Here we present evidence that high-performance thin-layer chromatography (HPTLC) represents an effective alternative to replace cost intensive mass spectrometry-based lipidomic analyses. We provide statistical comparison of identical samples by both methods, which support the use of HPTLC for quantitative analysis of the main yeast lipid classes., Competing Interests: Conflict of Interest: The authors declare no conflict of interest., (Copyright: © 2024 Meyer et al.)

The dynamic nature of the actin cytoskeleton is required to coordinate many cellular processes, and a loss of its plasticity has been linked to accelerated cell aging and attenuation of adaptive response mechanisms. Cofilin is an actin-binding protein that controls actin dynamics and has been linked to mitochondrial signaling pathways that control drug resistance and cell death. Here we show that cofilin-driven chronic depolarization of the actin cytoskeleton activates cell wall integrity mitogen-activated protein kinase (MAPK) signalling and disrupts lipid homeostasis in a voltage-dependent anion channel (VDAC)-dependent manner. Expression of the cof1-5 mutation, which reduces the dynamic nature of actin, triggers loss of cell wall integrity, vacuole fragmentation, disruption of lipid homeostasis, lipid droplet (LD) accumulation, and the promotion of cell death. The integrity of the actin cytoskeleton is therefore essential to maintain the fidelity of MAPK signaling, lipid homeostasis, and cell health in S. cerevisiae ., Competing Interests: The authors declare no competing interests., (© 2023 The Authors.)

Alcohol consumption and high caloric diet are leading causes of progressive fatty liver disease. Genetic variant rs738409 in patatin-like phospholipase domain-containing protein 3 (PNPLA3 rs738409 C>G) has been repeatedly described as one of the major risk loci for alcoholic liver cirrhosis (ALC) and hepatocellular carcinoma (HCC) in humans, however, the mechanism behind this association is incompletely understood. We generated mice carrying the rs738409 variant (PNPLA3 I148M) in order to detect genotype-phenotype relationships in mice upon chow and alcohol-high fat/high sugar diet (EtOH/WD). We could clearly demonstrate that the presence of rs738409 per se is sufficient to induce spontaneous development of steatosis after 1 year in mice on a chow diet, whereas in the setting of unhealthy diet feeding, PNPLA3 I148M did not affect hepatic inflammation or fibrosis, but induced a striking lipid remodeling, microvesicular steatosis and protected from HCC formation. Using shot gun lipidomics, we detected a striking restoration of reduced long chain-polyunsaturated fatty acids (LC-PUFA)-containing TGs, docosapentaenoic acid (C22:5 n3) and omega-3-derived eicosanoids (5-HEPE, 20-HEPE, 19,20-EDP, 21-HDHA) in PNPLA3 I148M mice upon EtOH/WD. At the molecular level, PNPLA3 I148M modulated enzymes for fatty acid and TG transport and metabolism. These findings suggest (dietary) lipids as an important and independent driver of hepatic tumorigenesis. Genetic variant in PNPLA3 exerted protective effects in mice, conflicting with findings in humans. Species-related differences in physiology and metabolism should be taken into account when modeling unhealthy human lifestyle, as genetic mouse models may not always allow for translation of insight gained in humans., Competing Interests: Declaration of Competing Interest The authors declare that they have no competing interests., (Copyright © 2022 Elsevier Inc. All rights reserved.)

The requirement of vitamin A for the synthesis of the visual chromophore and the light-sensing pigments has been studied in vertebrate and invertebrate model organisms. To identify the molecular mechanisms that orchestrate the ocular response to vitamin A deprivation, we took advantage of the fact that Drosophila melanogaster predominantly requires vitamin A for vision, but not for development or survival. We analyzed the impacts of vitamin A deficiency on the morphology, the lipidome, and the proteome of the Drosophila eye. We found that chronic vitamin A deprivation damaged the light-sensing compartments and caused a dramatic loss of visual pigments, but also decreased the molar abundance of most phototransduction proteins that amplify and transduce the visual signal. Unexpectedly, vitamin A deficiency also decreased the abundances of specific subunits of mitochondrial TCA cycle and respiratory chain components but increased the levels of cuticle- and lens-related proteins. In contrast, we found no apparent effects of vitamin A deficiency on the ocular lipidome. In summary, chronic vitamin A deficiency decreases the levels of most components of the visual signaling pathway, but also affects molecular pathways that are not vision-specific and whose mechanistic connection to vitamin A remains to be elucidated.

Parkinson's disease (PD) is a neurodegenerative disorder characterized by alpha-synuclein (αSyn) aggregation and associated with abnormalities in lipid metabolism. The accumulation of lipids in cytoplasmic organelles called lipid droplets (LDs) was observed in cellular models of PD. To investigate the pathophysiological consequences of interactions between αSyn and proteins that regulate the homeostasis of LDs, we used a transgenic Drosophila model of PD, in which human αSyn is specifically expressed in photoreceptor neurons. We first found that overexpression of the LD-coating proteins Perilipin 1 or 2 (dPlin1/2), which limit the access of lipases to LDs, markedly increased triacylglyclerol (TG) loaded LDs in neurons. However, dPlin-induced-LDs in neurons are independent of lipid anabolic (diacylglycerol acyltransferase 1/midway, fatty acid transport protein/dFatp) and catabolic (brummer TG lipase) enzymes, indicating that alternative mechanisms regulate neuronal LD homeostasis. Interestingly, the accumulation of LDs induced by various LD proteins (dPlin1, dPlin2, CG7900 or KlarsichtLD-BD) was synergistically amplified by the co-expression of αSyn, which localized to LDs in both Drosophila photoreceptor neurons and in human neuroblastoma cells. Finally, the accumulation of LDs increased the resistance of αSyn to proteolytic digestion, a characteristic of αSyn aggregation in human neurons. We propose that αSyn cooperates with LD proteins to inhibit lipolysis and that binding of αSyn to LDs contributes to the pathogenic misfolding and aggregation of αSyn in neurons., Competing Interests: The authors have declared that no competing interests exist.

Autophagy is a cellular recycling program which efficiently reduces the cellular burden of ageing. Autophagy is characterised by nucleation of isolation membranes, which grow in size and further expand to form autophagosomes, engulfing cellular material to be degraded by fusion with lysosomes (vacuole in yeast). Autophagosomal membranes do not bud from a single cell organelle, but are generated de novo. Several lipid sources for autophagosomal membranes have been identified, but the whole process of their generation is complex and not entirely understood. In this study, we investigated how the mitochondrial outer membrane protein porin 1 (Por1), the yeast orthologue of mammalian voltage-dependent anion channel (VDAC), affects autophagy in yeast. We show that POR1 deficiency reduces the autophagic capacity and leads to changes in vacuole and lipid homeostasis. We further investigated whether limited phosphatidylethanolamine (PE) availability in por1 ∆ was causative for reduced autophagy by overexpression of the PE-generating phosphatidylserine decarboxylase 1 (Psd1). Altogether, our results show that POR1 deficiency is associated with reduced autophagy, which can be circumvented by additional PSD1 overexpression. This suggests a role for Por1 in Psd1-mediated autophagy regulation.

There is little in vitro data available on long-term effects of TiO 2 exposure. Such data are important for improving the understanding of underlying mechanisms of adverse health effects of TiO 2 . Here, we exposed pulmonary epithelial cells to two doses (0.96 and 1.92 µg/cm 2 ) of TiO 2 for 13 weeks and effects on cell cycle and cell death mechanisms, i.e., apoptosis and autophagy were determined after 4, 8 and 13 weeks of exposure. Changes in telomere length, cellular protein levels and lipid classes were also analyzed at 13 weeks of exposure. We observed that the TiO 2 exposure increased the fraction of cells in G1-phase and reduced the fraction of cells in G2-phase, which was accompanied by an increase in the fraction of late apoptotic/necrotic cells. This corresponded with an induced expression of key apoptotic proteins i.e., BAD and BAX, and an accumulation of several lipid classes involved in cellular stress and apoptosis. These findings were further supported by quantitative proteome profiling data showing an increase in proteins involved in cell stress and genomic maintenance pathways following TiO 2 exposure. Altogether, we suggest that cell stress response and cell death pathways may be important molecular events in long-term health effects of TiO 2 .

Objective: The rs641738C>T variant located near the membrane-bound O-acyltransferase domain containing 7 (MBOAT7) locus is associated with fibrosis in liver diseases, including non-alcoholic fatty liver disease (NAFLD), alcohol-related liver disease, hepatitis B and C. We aim to understand the mechanism by which the rs641738C>T variant contributes to pathogenesis of NAFLD., Design: Mice with hepatocyte-specific deletion of MBOAT7 (Mboat7 Δhep ) were generated and livers were characterised by histology, flow cytometry, qPCR, RNA sequencing and lipidomics. We analysed the association of rs641738C>T genotype with liver inflammation and fibrosis in 846 NAFLD patients and obtained genotype-specific liver lipidomes from 280 human biopsies., Results: Allelic imbalance analysis of heterozygous human liver samples pointed to lower expression of the MBOAT7 transcript on the rs641738C>T haplotype. Mboat7 Δhep mice showed spontaneous steatosis characterised by increased hepatic cholesterol ester content after 10 weeks. After 6 weeks on a high fat, methionine-low, choline-deficient diet, mice developed increased hepatic fibrosis as measured by picrosirius staining (p < 0.05), hydroxyproline content (p < 0.05) and transcriptomics, while the inflammatory cell populations and inflammatory mediators were minimally affected. In a human biopsied NAFLD cohort, MBOAT7 rs641738C>T was associated with fibrosis (p = 0.004) independent of the presence of histological inflammation. Liver lipidomes of Mboat7 Δhep mice and human rs641738TT carriers with fibrosis showed increased total lysophosphatidylinositol levels. The altered lysophosphatidylinositol and phosphatidylinositol subspecies in MBOAT7 Δhep livers and human rs641738TT carriers were similar., Conclusion: Mboat7 deficiency in mice and human points to an inflammation-independent pathway of liver fibrosis that may be mediated by lipid signalling and a potentially targetable treatment option in NAFLD., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2021. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.)

Triacylglycerol (TG) and steryl ester (SE) lipid storage is a universal strategy to maintain organismal energy and membrane homeostasis. Cycles of building and mobilizing storage fat are fundamental in (re)distributing lipid substrates between tissues or to progress ontogenetic transitions. In this study, we show that Hormone-sensitive lipase (Hsl) specifically controls SE mobilization to initiate intergenerational sterol transfer in Drosophila melanogaster . Tissue-autonomous Hsl functions in the maternal fat body and germline coordinately prevent adult SE overstorage and maximize sterol allocation to embryos. While Hsl-deficiency is largely dispensable for normal development on sterol-rich diets, animals depend on adipocyte Hsl for optimal fecundity when dietary sterol becomes limiting. Notably, accumulation of SE but not of TG is a characteristic of Hsl-deficient cells across phyla including murine white adipocytes. In summary, we identified Hsl as an ancestral regulator of SE degradation, which improves intergenerational sterol transfer and reproductive success in flies., Competing Interests: CH, OK, HH, WM, IP, LS, GS, HX, SG, AS, RK No competing interests declared, (© 2021, Heier et al.)

Sentinel species are playing an indispensable role in monitoring environmental pollution in aquatic ecosystems. Many pollutants found in water prove to be endocrine disrupting chemicals that could cause disruptions in lipid homeostasis in aquatic species. A comprehensive profiling of the lipidome of these species is thus an essential step toward understanding the mechanism of toxicity induced by pollutants. Both the composition and spatial distribution of lipids in freshwater crustacean Gammarus fossarum were extensively examined herein. The baseline lipidome of gammarids of different sex and reproductive stages was established by high throughput shotgun lipidomics. Spatial lipid mapping by high resolution mass spectrometry imaging led to the discovery of sulfate-based lipids in hepatopancreas and their accumulation in mature oocytes. A diverse and dynamic lipid composition in G. fossarum was uncovered, which deepens our understanding of the biochemical changes during development and which could serve as a reference for future ecotoxicological studies., Competing Interests: The authors declare no competing interests., (© 2021 The Author(s).)

Nonalcoholic fatty liver disease (NAFLD) is a common metabolic dysfunction leading to hepatic steatosis. However, NAFLD's global impact on the liver lipidome is poorly understood. Using high-resolution shotgun mass spectrometry, we quantified the molar abundance of 316 species from 22 major lipid classes in liver biopsies of 365 patients, including nonsteatotic patients with normal or excessive weight, patients diagnosed with NAFL (nonalcoholic fatty liver) or NASH (nonalcoholic steatohepatitis), and patients bearing common mutations of NAFLD-related protein factors. We confirmed the progressive accumulation of di- and triacylglycerols and cholesteryl esters in the liver of NAFL and NASH patients, while the bulk composition of glycerophospho- and sphingolipids remained unchanged. Further stratification by biclustering analysis identified sphingomyelin species comprising n24:2 fatty acid moieties as membrane lipid markers of NAFLD. Normalized relative abundance of sphingomyelins SM 43:3;2 and SM 43:1;2 containing n24:2 and n24:0 fatty acid moieties, respectively, showed opposite trends during NAFLD progression and distinguished NAFL and NASH lipidomes from the lipidome of nonsteatotic livers. Together with several glycerophospholipids containing a C22:6 fatty acid moiety, these lipids serve as markers of early and advanced stages of NAFL., Competing Interests: Conflict of interest The authors declare that they have no conflicts of interest with the contents of this article., (Copyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.)

Aminotransaminases, including aspartate aminotransaminase (AST) and alanine aminotransaminase (ALT), are strongly involved in cancer cell metabolism and have been associated with prognosis in different types of cancer. The purpose of the present study was to evaluate the prognostic significance of the pre-treatment AST/ALT ratio in a large European cohort of patients with oral and oropharyngeal squamous cell cancer (OOSCC). Data from 515 patients treated for OOSCC at a tertiary academic center from 2000-2017 were retrospectively analyzed. Levels of AST and ALT were measured prior to the start of treatment. Uni- and multivariate Cox regression analyses were applied to evaluate the prognostic value of the AST/ALT ratio for cancer-specific survival (CSS) and overall survival (OS), survival rates were calculated. Univariate analyses showed a significant association of the AST/ALT ratio with CSS (hazard ratio (HR) 1.71, 95% confidence interval (CI) 1.38-2.12; p < 0.001) and OS (HR 1.69, 95% CI 1.41-2.02; p < 0.001). In multivariate analysis, the AST/ALT ratio remained an independent prognostic factor for CSS and OS (HR 1.45, 95% CI 1.12-1.88, p = 0.005 and HR 1.42, 95% CI 1.14-1.77, p = 0.002). Applying receiver operating characteristics (ROC) curve analysis, the optimal cut-off level for the AST/ALT ratio was 1.44, respectively. In multivariate analysis, an AST/ALT ratio > 1.44 was an independent prognostic factor for poor CSS and OS (HR 1.64, 95% CI 1.10-2.43, p = 0.014 and HR 1.55, 95% CI 1.12-2.15; p = 0.008). We conclude that the AST/ALT ratio is a prognostic marker for survival in OOSCC patients and could contribute to a better risk stratification and improved oncological therapy decisions.

Background: There is considerable evidence that platelets contribute to cancer growth and metastatic dissemination. In recent studies, altered mean platelet volume (MPV) has been associated with prognosis in different types of cancer. However, the prognostic role of the MPV in head and neck squamous cell cancer (HNSCC) is currently discussed controversially. The present study was performed to analyze and further elucidate the prognostic significance of the MPV in HNSCC., Methods: A total of 319 oropharyngeal squamous cell cancer (OPSCC) patients treated with radiotherapy at a tertiary academic center were enrolled in the present study. Kaplan-Meier method as well as uni- and multivariate Cox proportional hazards were used to evaluate the impact of MPV on cancer-specific survival (CSS), locoregional control (LC) and recurrence-free survival (RFS)., Results: The median MPV was 10.30 fL (mean 10.26 ± 1.17fL). Univariate analyses showed a significant association of the MPV with CSS (HR 0.85, 95% CI 0.74-0.98, p = 0.025), LC (HR 0.86, 95% CI 0.74-0.99, p = 0.034) and RFS (HR 0.87, 95% CI 0.76-0.996; p = 0.043). In multivariate analysis, the MPV remained an independent prognostic factor for CSS (HR 0.77, 95% CI 0.63-0.93, p = 0.008), LC (HR 0.80, 95% CI 0.65-0.98, p = 0.030), and RFS (HR 0.83, 95% CI 0.685-0.999, p = 0.049)., Conclusions: Our findings indicate that the MPV is a prognostic marker in OPSCC patients and may contribute to future individual risk assessment.

Metabolic research is a challenge because of the variety of data within experimental series and the difficulty of replicating results among scientific groups. The fruit fly, Drosophila melanogaster, is a cost-effective and reliable pioneer model to screen dietary variables for metabolic research. One of the main reasons for problems in this field are differences in food recipes, diet-associated microbial environments and the pharmacokinetic behavior of nutrients across the gut-blood barrier. To prevent such experimental shortcomings, a common strategy is to pool scores of subjects into one sample to create an average statement. However, this approach lacks information about the biological spread and may provoke misleading interpretations. We propose to use the developmental rate of individual Drosophila larvae as a metabolic sensor. To do so, we introduce here a 96-well plate-based assay, which allows screening for multiple variables including food quality, microbial load, and genetic differences. We demonstrate that on a diet that is rich in calories, pupation is sensitive to the variation of dietary lipid compounds and that genotypes considered as wild-types/controls produce different developmental profiles. Our platform is suited for later automation and represents a potent high-throughput screening tool for the pharmacology and food industry. If used systematically, our assay could become a powerful reference tool to compare the quality of used dietary configurations with published benchmark recipes., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

During cold acclimation fruit flies switch their feeding from yeast to plant food, however there are no robust molecular markers to monitor this in the wild. Drosophila melanogaster is a sterol auxotroph and relies on dietary sterols to produce lipid membranes, lipoproteins and molting hormones. We employed shotgun lipidomics to quantify eight major food sterols in total lipid extracts of heads and genital tracts of adult male and female flies. We found that their sterol composition is dynamic and reflective of fly diet in an organ-specific manner. Season-dependent changes observed in the organs of wild-living flies suggested that the molar ratio between yeast (ergosterol, zymosterol) and plant (sitosterol, stigmasterol) sterols is a quantifiable, generic and unequivocal marker of their feeding behavior suitable for ecological and environmental population-based studies. The enrichment of phytosterols over yeast sterols in wild-living flies at low temperatures is consistent with switching from yeast to plant diet and corroborates the concomitantly increased unsaturation of their membrane lipids., Competing Interests: Declaration of competing interest The authors declare that they have no competing interests., (Copyright © 2020 Elsevier B.V. All rights reserved.)

The purpose of the present study was to evaluate the prognostic significance of the pre- treatment C-reactive protein (CRP) level in a cohort of 503 patients with oral and oropharyngeal cancer treated at a tertiary academic center between 2000 and 2017. Cancer-specific survival (CSS), overall survival (OS) and loco-regional control (LC) were calculated using Kaplan-Meier analysis. To evaluate the prognostic value of the CRP level for the clinical endpoints, univariate and multivariate Cox regression models were applied. The median follow-up period was 61 months. Patients were divided into elevated CRP (≥5 mg/L) and normal CRP groups, according to pre-treatment plasma levels. An increased CRP level was significantly associated with shorter CSS ( p < 0.001, log-rank test), as well as with shorter OS ( p < 0.001, log-rank test) and loco-regional control ( p = 0.001, log-rank test). In addition, multivariate analysis identified CRP as an independent predictor for CSS (hazard ratio (HR) 1.59, 95% confidence interval (CI) 1.08-2.35; p = 0.020) as well as for OS (HR 1.62, 95%CI 1.17-2.24; p = 0.004) and LC (HR 1.50, 95%CI 1.06-2.14; p = 0.023). In subgroup analysis, Kaplan Meier curves revealed that an elevated pre-treatment CRP level was a consistent prognostic factor for poor CSS ( p = 0.003, log-rank test), OS ( p = 0.001, log-rank test), and LC ( p = 0.028, log-rank test) in patients treated with definitive (chemo-) radiotherapy, whereas a significant association in patients undergoing surgery and postoperative radiotherapy was not detected. The pre-treatment CRP level seems to represent a prognostic factor for CSS, OS, and LC in patients with oral and oropharyngeal cancer, particularly in those treated with definitive (chemo-) radiotherapy. Additional large-scale prospective studies are warranted to confirm and extend our findings.

Autophagy, a membrane-dependent catabolic process, ensures survival of aging cells and depends on the cellular energetic status. Acetyl-CoA carboxylase 1 (Acc1) connects central energy metabolism to lipid biosynthesis and is rate-limiting for the de novo synthesis of lipids. However, it is unclear how de novo lipogenesis and its metabolic consequences affect autophagic activity. Here, we show that in aging yeast, autophagy levels highly depend on the activity of Acc1. Constitutively active Acc1 ( acc1 S/A ) or a deletion of the Acc1 negative regulator, Snf1 (yeast AMPK), shows elevated autophagy levels, which can be reversed by the Acc1 inhibitor soraphen A. Vice versa, pharmacological inhibition of Acc1 drastically reduces cell survival and results in the accumulation of Atg8-positive structures at the vacuolar membrane, suggesting late defects in the autophagic cascade. As expected, acc1 S/A cells exhibit a reduction in acetate/acetyl-CoA availability along with elevated cellular lipid content. However, concomitant administration of acetate fails to fully revert the increase in autophagy exerted by acc1 S/A Instead, administration of oleate, while mimicking constitutively active Acc1 in WT cells, alleviates the vacuolar fusion defects induced by Acc1 inhibition. Our results argue for a largely lipid-dependent process of autophagy regulation downstream of Acc1. We present a versatile genetic model to investigate the complex relationship between acetate metabolism, lipid homeostasis, and autophagy and propose Acc1-dependent lipogenesis as a fundamental metabolic path downstream of Snf1 to maintain autophagy and survival during cellular aging., (© 2019 Gross et al.)

Kleiber's law, or the 3/4 -power law scaling of the metabolic rate with body mass, is considered one of the few quantitative laws in biology, yet its physiological basis remains unknown. Here, we report Kleiber's law scaling in the planarian Schmidtea mediterranea . Its reversible and life history-independent changes in adult body mass over 3 orders of magnitude reveal that Kleiber's law does not emerge from the size-dependent decrease in cellular metabolic rate, but from a size-dependent increase in mass per cell. Through a combination of experiment and theoretical analysis of the organismal energy balance, we further show that the mass allometry is caused by body size dependent energy storage. Our results reveal the physiological origins of Kleiber's law in planarians and have general implications for understanding a fundamental scaling law in biology., Competing Interests: AT, SW, OF, JP, OK, YQ, KF, AS, BF, JR No competing interests declared, FJ Reviewing editor, eLife, (© 2019, Thommen et al.)

Shotgun analysis provides a quantitative snapshot of the lipidome composition of cells, tissues, or model organisms; however, it does not elucidate the spatial distribution of lipids. Here we demonstrate that shotgun analysis could quantify low-picomole amounts of lipids isolated by laser capture microdissection (LCM) of hundred micrometer-sized histological zones visualized at the cryosections of tissues. We identified metabolically distinct periportal (pp) and pericentral (pc) zones by immunostaining of 20 μm thick cryosections of a healthy mouse liver. LCM was used to ablate, catapult, and collect the tissue material from 10 to 20 individual zones covering a total area of 0.3-0.5 mm 2 and containing ca. 500 cells. Top-down shotgun profiling relying upon computational stitching of 61 targeted selective ion monitoring ( t-SIM) spectra quantified more than 200 lipid species from 17 lipid classes including glycero- and glycerophospholipids, sphingolipids, cholesterol esters, and cholesterol. Shotgun LCM revealed the overall commonality of the full lipidome composition of pp and pc zones along with significant ( p < 0.001) difference in the relative abundance of 13 lipid species. Follow-up proteomics analyses of pellets recovered from an aqueous phase saved after the lipid extraction identified 13 known and 7 new protein markers exclusively present in pp or in pc zones and independently validated the specificity of their visualization, isolation, and histological assignment.

The effects of long-term chronic exposure of human lung cells to multi-walled carbon nanotubes (MWCNT) and their impact upon cellular proteins and lipids were investigated. Since the lung is the major target organ, an in vitro normal bronchial epithelial cell line model was used. Additionally, to better mimic exposure to manufactured nanomaterials at occupational settings, cells were continuously exposed to two non-toxic and low doses of a MWCNT for 13-weeks. MWCNT-treatment increased ROS levels in cells without increasing oxidative DNA damage and resulted in differential expression of multiple anti- and pro-apoptotic proteins. The proteomic analysis of the MWCNT-exposed cells showed that among more than 5000 identified proteins; more than 200 were differentially expressed in the treated cells. Functional analyses revealed association of these differentially regulated proteins to cellular processes such as cell death and survival, cellular assembly, and organization. Similarly, shotgun lipidomic profiling revealed accumulation of multiple lipid classes. Our results indicate that long-term MWCNT-exposure of human normal lung cells at occupationally relevant low-doses may alter both the proteome and the lipidome profiles of the target epithelial cells in the lung.

The loss of lipid homeostasis can lead to lipid overload and is associated with a variety of disease states. However, little is known as to how the disruption of lipid regulation or lipid overload affects cell survival. In this study we investigated how excess diacylglycerol (DG), a cardinal metabolite suspected to mediate lipotoxicity, compromises the survival of yeast cells. We reveal that increased DG achieved by either genetic manipulation or pharmacological administration of 1,2-dioctanoyl-sn-glycerol (DOG) triggers necrotic cell death. The toxic effects of DG are linked to glucose metabolism and require a functional Rim101 signaling cascade involving the Rim21-dependent sensing complex and the activation of a calpain-like protease. The Rim101 cascade is an established pathway that triggers a transcriptional response to alkaline or lipid stress. We propose that the Rim101 pathway senses DG-induced lipid perturbation and conducts a signaling response that either facilitates cellular adaptation or triggers lipotoxic cell death. Using established models of lipotoxicity, i.e., high-fat diet in Drosophila and palmitic acid administration in cultured human endothelial cells, we present evidence that the core mechanism underlying this calpain-dependent lipotoxic cell death pathway is phylogenetically conserved.

Elevated circulating fatty acids (FAs) contribute to the development of obesity-associated metabolic complications such as insulin resistance (IR) and non-alcoholic fatty liver disease (NAFLD). Hence, reducing adipose tissue lipolysis to diminish the mobilization of FAs and lower their respective plasma concentrations represents a potential treatment strategy to counteract obesity-associated disorders. Here we show that specific inhibition of adipose triglyceride lipase (Atgl) with the chemical inhibitor, Atglistatin, effectively reduces adipose tissue lipolysis, weight gain, IR and NAFLD in mice fed a high-fat diet. Importantly, even long-term treatment does not lead to lipid accumulation in ectopic tissues such as the skeletal muscle or heart. Thus, the severe cardiac steatosis and cardiomyopathy that is observed in genetic models of Atgl deficiency does not occur in Atglistatin-treated mice. Our data validate the pharmacological inhibition of Atgl as a potentially powerful therapeutic strategy to treat obesity and associated metabolic disorders.

Clinical lipidomics is an emerging biomarker discovery approach that compares lipid profiles under pathologically and physiologically normal conditions. Here we describe a method for the absolute (molar) quantification of more than 200 molecules from 14 major lipid classes from 5 μL of human blood plasma using high-resolution top-down shotgun mass spectrometry. Because of its technical simplicity and robustness, the protocol lends itself for high-throughput clinical lipidomics screens.

Aging is associated with an increased risk of cardiovascular disease and death. Here we show that oral supplementation of the natural polyamine spermidine extends the lifespan of mice and exerts cardioprotective effects, reducing cardiac hypertrophy and preserving diastolic function in old mice. Spermidine feeding enhanced cardiac autophagy, mitophagy and mitochondrial respiration, and it also improved the mechano-elastical properties of cardiomyocytes in vivo, coinciding with increased titin phosphorylation and suppressed subclinical inflammation. Spermidine feeding failed to provide cardioprotection in mice that lack the autophagy-related protein Atg5 in cardiomyocytes. In Dahl salt-sensitive rats that were fed a high-salt diet, a model for hypertension-induced congestive heart failure, spermidine feeding reduced systemic blood pressure, increased titin phosphorylation and prevented cardiac hypertrophy and a decline in diastolic function, thus delaying the progression to heart failure. In humans, high levels of dietary spermidine, as assessed from food questionnaires, correlated with reduced blood pressure and a lower incidence of cardiovascular disease. Our results suggest a new and feasible strategy for protection against cardiovascular disease.

α/β Hydrolase domain-containing 6 (ABHD6) can act as monoacylglycerol hydrolase and is believed to play a role in endocannabinoid signaling as well as in the pathogenesis of obesity and liver steatosis. However, the mechanistic link between gene function and disease is incompletely understood. Here we aimed to further characterize the role of ABHD6 in lipid metabolism. We show that mouse and human ABHD6 degrade bis(monoacylglycero)phosphate (BMP) with high specific activity. BMP, also known as lysobisphosphatidic acid, is enriched in late endosomes/lysosomes, where it plays a key role in the formation of intraluminal vesicles and in lipid sorting. Up to now, little has been known about the catabolism of this lipid. Our data demonstrate that ABHD6 is responsible for ∼ 90% of the BMP hydrolase activity detected in the liver and that knockdown of ABHD6 increases hepatic BMP levels. Tissue fractionation and live-cell imaging experiments revealed that ABHD6 co-localizes with late endosomes/lysosomes. The enzyme is active at cytosolic pH and lacks acid hydrolase activity, implying that it degrades BMP exported from acidic organelles or de novo-formed BMP. In conclusion, our data suggest that ABHD6 controls BMP catabolism and is therefore part of the late endosomal/lysosomal lipid-sorting machinery., (© 2015 by The American Society for Biochemistry and Molecular Biology, Inc.)

Background: Membrane protein research is frequently hampered by the low natural abundance of these proteins in cells and typically relies on recombinant gene expression. Different expression systems, like mammalian cells, insect cells, bacteria and yeast are being used, but very few research efforts have been directed towards specific host cell customization for enhanced expression of membrane proteins. Here we show that by increasing the intracellular membrane production by interfering with a key enzymatic step of lipid synthesis, enhanced expression of membrane proteins in yeast is achieved., Results: We engineered the oleotrophic yeast, Yarrowia lipolytica, by deleting the phosphatidic acid phosphatase, PAH1, which led to massive proliferation of endoplasmic reticulum (ER) membranes. For all eight tested representatives of different integral membrane protein families, we obtained enhanced protein accumulation levels and in some cases enhanced proteolytic integrity in the ∆pah1 strain. We analysed the adenosine A2AR G-protein coupled receptor case in more detail and found that concomitant induction of the unfolded protein response in the ∆pah1 strain enhanced the specific ligand binding activity of the receptor. These data indicate an improved quality control mechanism for membrane proteins accumulating in yeast cells with proliferated ER., Conclusions: We conclude that redirecting the metabolic flux of fatty acids away from triacylglycerol- and sterylester-storage towards membrane phospholipid synthesis by PAH1 gene inactivation, provides a valuable approach to enhance eukaryotic membrane protein production. Complementary to this improvement in membrane protein quantity, UPR co-induction further enhances the quality of the membrane protein in terms of its proper folding and biological activity. Importantly, since these pathways are conserved in all eukaryotes, it will be of interest to investigate similar engineering approaches in other cell types of biotechnological interest, such as insect cells and mammalian cells.

The 'discovery' of lipid droplets as a metabolically highly active subcellular organelle has sparked great scientific interest in its research in recent years. The previous view of a rather inert storage pool of neutral lipids--triacylglycerol and sterols or steryl esters--has markedly changed. Driven by the endemic dimensions of lipid-associated disorders on the one hand, and the promising biotechnological application to generate oils ('biodiesel') from single-celled organisms on the other, multiple model organisms are exploited in basic and applied research to develop a better understanding of biogenesis and metabolism of this organelle. This article summarizes the current status of LD research in yeast and experimental approaches to obtain insight into the regulatory and structural components driving lipid droplet formation and their physiological and pathophysiological roles in lipid homeostasis.

Numerous studies in humans link a nonsynonymous genetic polymorphism (I148M) in adiponutrin (ADPN) to various forms of fatty liver disease and liver cirrhosis. Despite its high clinical relevance, the molecular function of ADPN and the mechanism by which I148M variant affects hepatic metabolism are unclear. Here we show that ADPN promotes cellular lipid synthesis by converting lysophosphatidic acid (LPA) into phosphatidic acid. The ADPN-catalyzed LPA acyltransferase (LPAAT) reaction is specific for LPA and long-chain acyl-CoAs. Wild-type mice receiving a high-sucrose diet exhibit substantial upregulation of Adpn in the liver and a concomitant increase in LPAAT activity. In Adpn-deficient mice, this diet-induced increase in hepatic LPAAT activity is reduced. Notably, the I148M variant of human ADPN exhibits increased LPAAT activity leading to increased cellular lipid accumulation. This gain of function provides a plausible biochemical mechanism for the development of liver steatosis in subjects carrying the I148M variant., (Copyright © 2012 Elsevier Inc. All rights reserved.)