Diet suppresses glioblastoma initiation in mice by maintaining quiescence of mutation-bearing neural stem cells.

Glioblastoma is thought to originate from neural stem cells (NSCs) of the subventricular zone that acquire genetic alterations. In the adult brain, NSCs are largely quiescent, suggesting that deregulation of quiescence maintenance may be a prerequisite for tumor initiation. Although inactivation of the tumor suppressor p53 is a frequent event in gliomagenesis, whether or how it affects quiescent NSCs (qNSCs) remains unclear. Here, we show that p53 maintains quiescence by inducing fatty-acid oxidation (FAO) and that acute p53 deletion in qNSCs results in their premature activation to a proliferative state. Mechanistically, this occurs through direct transcriptional induction of PPARGC1a, which in turn activates PPARα to upregulate FAO genes. Dietary supplementation with fish oil containing omega-3 fatty acids, natural PPARα ligands, fully restores quiescence of p53-deficient NSCs and delays tumor initiation in a glioblastoma mouse model. Thus, diet can silence glioblastoma driver mutations, with important implications for cancer prevention.

The dietary sweetener sucralose is a negative modulator of T cell-mediated responses.

Artificial sweeteners are used as calorie-free sugar substitutes in many food products and their consumption has increased substantially over the past years1. Although generally regarded as safe, some concerns have been raised about the long-term safety of the consumption of certain sweeteners2-5. In this study, we show that the intake of high doses of sucralose in mice results in immunomodulatory effects by limiting T cell proliferation and T cell differentiation. Mechanistically, sucralose affects the membrane order of T cells, accompanied by a reduced efficiency of T cell receptor signalling and intracellular calcium mobilization. Mice given sucralose show decreased CD8+ T cell antigen-specific responses in subcutaneous cancer models and bacterial infection models, and reduced T cell function in models of T cell-mediated autoimmunity. Overall, these findings suggest that a high intake of sucralose can dampen T cell-mediated responses, an effect that could be used in therapy to mitigate T cell-dependent autoimmune disorders.

Functions of Stress-Induced Lipid Droplets in the Nervous System.

Lipid droplets are highly dynamic intracellular organelles that store neutral lipids such as cholesteryl esters and triacylglycerols. They have recently emerged as key stress response components in many different cell types. Lipid droplets in the nervous system are mostly observed in vivo in glia, ependymal cells and microglia. They tend to become more numerous in these cell types and can also form in neurons as a consequence of ageing or stresses involving redox imbalance and lipotoxicity. Abundant lipid droplets are also a characteristic feature of several neurodegenerative diseases. In this minireview, we take a cell-type perspective on recent advances in our understanding of lipid droplet metabolism in glia, neurons and neural stem cells during health and disease. We highlight that a given lipid droplet subfunction, such as triacylglycerol lipolysis, can be physiologically beneficial or harmful to the functions of the nervous system depending upon cellular context. The mechanistic understanding of context-dependent lipid droplet functions in the nervous system is progressing apace, aided by new technologies for probing the lipid droplet proteome and lipidome with single-cell type precision.

Metabolic decisions in development and disease-a Keystone Symposia report.

There is an increasing appreciation for the role of metabolism in cell signaling and cell decision making. Precise metabolic control is essential in development, as evident by the disorders caused by mutations in metabolic enzymes. The metabolic profile of cells is often cell-type specific, changing as cells differentiate or during tumorigenesis. Recent evidence has shown that changes in metabolism are not merely a consequence of changes in cell state but that metabolites can serve to promote and/or inhibit these changes. Metabolites can link metabolic pathways with cell signaling pathways via several mechanisms, for example, by serving as substrates for protein post-translational modifications, by affecting enzyme activity via allosteric mechanisms, or by altering epigenetic markers. Unraveling the complex interactions governing metabolism, gene expression, and protein activity that ultimately govern a cell's fate will require new tools and interactions across disciplines. On March 24 and 25, 2021, experts in cell metabolism, developmental biology, and human disease met virtually for the Keystone eSymposium, "Metabolic Decisions in Development and Disease." The discussions explored how metabolites impact cellular and developmental decisions in a diverse range of model systems used to investigate normal development, developmental disorders, dietary effects, and cancer-mediated changes in metabolism.

Adipose triglyceride lipase protects renal cell endocytosis in a Drosophila dietary model of chronic kidney disease.

Obesity-related renal lipotoxicity and chronic kidney disease (CKD) are prevalent pathologies with complex aetiologies. One hallmark of renal lipotoxicity is the ectopic accumulation of lipid droplets in kidney podocytes and in proximal tubule cells. Renal lipid droplets are observed in human CKD patients and in high-fat diet (HFD) rodent models, but their precise role remains unclear. Here, we establish a HFD model in Drosophila that recapitulates renal lipid droplets and several other aspects of mammalian CKD. Cell type-specific genetic manipulations show that lipid can overflow from adipose tissue and is taken up by renal cells called nephrocytes. A HFD drives nephrocyte lipid uptake via the multiligand receptor Cubilin (Cubn), leading to the ectopic accumulation of lipid droplets. These nephrocyte lipid droplets correlate with endoplasmic reticulum (ER) and mitochondrial deficits, as well as with impaired macromolecular endocytosis, a key conserved function of renal cells. Nephrocyte knockdown of diglyceride acyltransferase 1 (DGAT1), overexpression of adipose triglyceride lipase (ATGL), and epistasis tests together reveal that fatty acid flux through the lipid droplet triglyceride compartment protects the ER, mitochondria, and endocytosis of renal cells. Strikingly, boosting nephrocyte expression of the lipid droplet resident enzyme ATGL is sufficient to rescue HFD-induced defects in renal endocytosis. Moreover, endocytic rescue requires a conserved mitochondrial regulator, peroxisome proliferator-activated receptor-gamma coactivator 1α (PGC1α). This study demonstrates that lipid droplet lipolysis counteracts the harmful effects of a HFD via a mitochondrial pathway that protects renal endocytosis. It also provides a genetic strategy for determining whether lipid droplets in different biological contexts function primarily to release beneficial or to sequester toxic lipids.

Quantification of fetal organ sparing in maternal low-protein dietary models.

Background: Maternal malnutrition can lead to fetal growth restriction. This is often associated with organ sparing and long-lasting physiological dysfunctions during adulthood, although the underlying mechanisms are not yet well understood. Methods: Low protein (LP) dietary models in C57BL/6J mice were used to investigate the proximal effects of maternal malnutrition on fetal organ weights and organ sparing at embryonic day 18.5 (E18.5). Results:  Maternal 8% LP diet induced strikingly different degrees of fetal growth restriction in different animal facilities, but adjustment of dietary protein content allowed similar fetal body masses to be obtained. A maternal LP diet that restricted fetal body mass by 40% did not decrease fetal brain mass to the same extent, reflecting positive growth sparing of this organ. Under these conditions, fetal pancreas and liver mass decreased by 60-70%, indicative of negative organ sparing. A series of dietary swaps between LP and standard diets showed that the liver is capable of efficient catch-up growth from as late as E14.5 whereas, after E10.5, the pancreas is not. Conclusions: This study highlights that the reproducibility of LP fetal growth restriction studies between laboratories can be improved by careful calibration of maternal dietary protein content. LP diets that induce 30-40% restriction of prenatal growth provide a good model for fetal organ sparing. For the liver, recovery of growth following protein restriction is efficient throughout fetal development but, for the pancreas, transient LP exposures spanning the progenitor expansion phase lead to an irreversible fetal growth deficit.

Cryogenic OrbiSIMS Localizes Semi-Volatile Molecules in Biological Tissues.

OrbiSIMS is a recently developed instrument for label-free imaging of chemicals with micron spatial resolution and high mass resolution. We report a cryogenic workflow for OrbiSIMS (Cryo-OrbiSIMS) that improves chemical detection of lipids and other biomolecules in tissues. Cryo-OrbiSIMS boosts ionization yield and decreases ion-beam induced fragmentation, greatly improving the detection of biomolecules such as triacylglycerides. It also increases chemical coverage to include molecules with intermediate or high vapor pressures, such as free fatty acids and semi-volatile organic compounds (SVOCs). We find that Cryo-OrbiSIMS reveals the hitherto unknown localization patterns of SVOCs with high spatial and chemical resolution in diverse plant, animal, and human tissues. We also show that Cryo-OrbiSIMS can be combined with genetic analysis to identify enzymes regulating SVOC metabolism. Cryo-OrbiSIMS is applicable to high resolution imaging of a wide variety of non-volatile and semi-volatile molecules across many areas of biomedicine.

An Improved Method for Measuring Absolute Metabolite Concentrations in Small Biofluid or Tissue Samples.

The measurement of absolute metabolite concentrations in small samples remains a significant analytical challenge. This is particularly the case when the sample volume is only a few microliters or less and cannot be determined accurately via direct measurement. We previously developed volume determination with two standards (VDTS) as a method to address this challenge for biofluids. As a proof-of-principle, we applied VDTS to NMR spectra of polar metabolites in the hemolymph (blood) of the tiny yet powerful genetic model Drosophila melanogaster. This showed that VDTS calculation of absolute metabolite concentrations in fed versus starved Drosophila larvae is more accurate than methods utilizing normalization to total spectral signal. Here, we introduce paired VDTS (pVDTS), an improved VDTS method for biofluids and solid tissues that implements the statistical power of paired control and experimental replicates. pVDTS utilizes new equations that also include a correction for dilution errors introduced by the variable surface wetness of solid samples. We then show that metabolite concentrations in Drosophila larvae are more precisely determined and logically consistent using pVDTS than using the original VDTS method. The refined pVDTS workflow described in this study is applicable to a wide range of different tissues and biofluids.

Histidine is selectively required for the growth of Myc-dependent dedifferentiation tumours in the Drosophila CNS.

Rewired metabolism of glutamine in cancer has been well documented, but less is known about other amino acids such as histidine. Here, we use Drosophila cancer models to show that decreasing the concentration of histidine in the diet strongly inhibits the growth of mutant clones induced by loss of Nerfin-1 or gain of Notch activity. In contrast, changes in dietary histidine have much less effect on the growth of wildtype neural stem cells and Prospero neural tumours. The reliance of tumours on dietary histidine and also on histidine decarboxylase (Hdc) depends upon their growth requirement for Myc. We demonstrate that Myc overexpression in nerfin-1 tumours is sufficient to switch their mode of growth from histidine/Hdc sensitive to resistant. This study suggests that perturbations in histidine metabolism selectively target neural tumours that grow via a dedifferentiation process involving large cell size increases driven by Myc.

Two Negatives Make a Positive for Insulin Secretion and Growth.

The confusingly named growth-blocking peptides are nutrient-dependent adipokines that stimulate insulin secretion and boost growth in developing flies. In this issue of Developmental Cell, Meschi et al. (2018) show that these adipose tissue-derived factors regulate insulin secretion by silencing a pair of inhibitory neurons that synapse with insulin-producing cells.

Sex-lethal in neurons controls female body growth in Drosophila.

Sexual size dimorphism (SSD), a sex difference in body size, is widespread throughout the animal kingdom, raising the question of how sex influences existing growth regulatory pathways to bring about SSD. In insects, somatic sexual differentiation has long been considered to be controlled strictly cell-autonomously. Here, we discuss our surprising finding that in Drosophila larvae, the sex determination gene Sex-lethal (Sxl) functions in neurons to non-autonomously specify SSD. We found that Sxl is required in specific neuronal subsets to upregulate female body growth, including in the neurosecretory insulin producing cells, even though insulin-like peptides themselves appear not to be involved. SSD regulation by neuronal Sxl is also independent of its known splicing targets, transformer and msl-2, suggesting that it involves a new molecular mechanism. Interestingly, SSD control by neuronal Sxl is selective for larval, not imaginal tissue types, and operates in addition to cell-autonomous effects of Sxl and Tra, which are present in both larval and imaginal tissues. Overall, our findings add to a small but growing number of studies reporting non-autonomous, likely hormonal, control of sex differences in Drosophila, and suggest that the principles of sexual differentiation in insects and mammals may be more similar than previously thought.

Early-life exposure to low-dose oxidants can increase longevity via microbiome remodelling in Drosophila.

Environmental stresses experienced during development exert many long-term effects upon health and disease. For example, chemical oxidants or genetic perturbations that induce low levels of reactive oxygen species can extend lifespan in several species. In some cases, the beneficial effects of low-dose oxidants are attributed to adaptive protective mechanisms such as mitohormesis, which involve long-term increases in the expression of stress response genes. Here we show in Drosophila that transient exposure to low concentrations of oxidants during development leads to an extension of adult lifespan. Surprisingly, this depends upon oxidants acting in an antibiotic-like manner to selectively deplete the microbiome of Acetobacter proteobacteria. We demonstrate that the presence of Acetobacter species, such as A. aceti, in the indigenous microbiota increases age-related gut dysfunction and shortens lifespan. This study demonstrates that low-dose oxidant exposure during early life can extend lifespan via microbiome remodelling rather than mitohormesis.

Developmental diet regulates Drosophila lifespan via lipid autotoxins.

Early-life nourishment exerts long-term influences upon adult physiology and disease risk. These lasting effects of diet are well established but the underlying mechanisms are only partially understood. Here we show that restricting dietary yeast during Drosophila development can, depending upon the subsequent adult environment, more than double median lifespan. Developmental diet acts via a long-term influence upon the adult production of toxic molecules, which we term autotoxins, that are shed into the environment and shorten the lifespan of both sexes. Autotoxins are synthesised by oenocytes and some of them correspond to alkene hydrocarbons that also act as pheromones. This study identifies a mechanism by which the developmental dietary history of an animal regulates its own longevity and that of its conspecific neighbours. It also has important implications for the design of lifespan experiments as autotoxins can influence the regulation of longevity by other factors including diet, sex, insulin signalling and population density.

The sex of specific neurons controls female body growth in Drosophila.

Sexual dimorphisms in body size are widespread throughout the animal kingdom but their underlying mechanisms are not well characterized. Most models for how sex chromosome genes specify size dimorphism have emphasized the importance of gonadal hormones and cell-autonomous influences in mammals versus strictly cell-autonomous mechanisms in Drosophila melanogaster. Here, we use tissue-specific genetics to investigate how sexual size dimorphism (SSD) is established in Drosophila. We find that the larger body size characteristic of Drosophila females is established very early in larval development via an increase in the growth rate per unit of body mass. We demonstrate that the female sex determination gene, Sex-lethal (Sxl), functions in central nervous system (CNS) neurons as part of a relay that specifies the early sex-specific growth trajectories of larval but not imaginal tissues. Neuronal Sxl acts additively in 2 neuronal subpopulations, one of which corresponds to 7 median neurosecretory cells: the insulin-producing cells (IPCs). Surprisingly, however, male-female differences in the production of insulin-like peptides (Ilps) from the IPCs do not appear to be involved in establishing SSD in early larvae, although they may play a later role. These findings support a relay model in which Sxl in neurons and Sxl in local tissues act together to specify the female-specific growth of the larval body. They also reveal that, even though the sex determination pathways in Drosophila and mammals are different, they both modulate body growth via a combination of tissue-autonomous and nonautonomous inputs.

Lipid droplet functions beyond energy storage.

Lipid droplets are cytoplasmic organelles that store neutral lipids and are critically important for energy metabolism. Their function in energy storage is firmly established and increasingly well characterized. However, emerging evidence indicates that lipid droplets also play important and diverse roles in the cellular handling of lipids and proteins that may not be directly related to energy homeostasis. Lipid handling roles of droplets include the storage of hydrophobic vitamin and signaling precursors, and the management of endoplasmic reticulum and oxidative stress. Roles of lipid droplets in protein handling encompass functions in the maturation, storage, and turnover of cellular and viral polypeptides. Other potential roles of lipid droplets may be connected with their intracellular motility and, in some cases, their nuclear localization. This diversity highlights that lipid droplets are very adaptable organelles, performing different functions in different biological contexts. This article is part of a Special Issue entitled: Recent Advances in Lipid Droplet Biology edited by Rosalind Coleman and Matthijs Hesselink.

Stable isotope analysis of dynamic lipidomics.

Metabolic pathway flux is a fundamental element of biological activity, which can be quantified using a variety of mass spectrometric techniques to monitor incorporation of stable isotope-labelled substrates into metabolic products. This article contrasts developments in electrospray ionisation mass spectrometry (ESI-MS) for the measurement of lipid metabolism with more established gas chromatography mass spectrometry and isotope ratio mass spectrometry methodologies. ESI-MS combined with diagnostic tandem MS/MS scans permits the sensitive and specific analysis of stable isotope-labelled substrates into intact lipid molecular species without the requirement for lipid hydrolysis and derivatisation. Such dynamic lipidomic methodologies using non-toxic stable isotopes can be readily applied to quantify lipid metabolic fluxes in clinical and metabolic studies in vivo. However, a significant current limitation is the absence of appropriate software to generate kinetic models of substrate incorporation into multiple products in the time domain. Finally, we discuss the future potential of stable isotope-mass spectrometry imaging to quantify the location as well as the extent of lipid synthesis. This article is part of a Special Issue entitled: BBALIP_Lipidomics Opinion Articles edited by Sepp Kohlwein.

Drosophila Spidey/Kar Regulates Oenocyte Growth via PI3-Kinase Signaling.

Cell growth and proliferation depend upon many different aspects of lipid metabolism. One key signaling pathway that is utilized in many different anabolic contexts involves Phosphatidylinositide 3-kinase (PI3K) and its membrane lipid products, the Phosphatidylinositol (3,4,5)-trisphosphates. It remains unclear, however, which other branches of lipid metabolism interact with the PI3K signaling pathway. Here, we focus on specialized fat metabolizing cells in Drosophila called larval oenocytes. In the presence of dietary nutrients, oenocytes undergo PI3K-dependent cell growth and contain very few lipid droplets. In contrast, during starvation, oenocytes decrease PI3K signaling, shut down cell growth and accumulate abundant lipid droplets. We now show that PI3K in larval oenocytes, but not in fat body cells, functions to suppress lipid droplet accumulation. Several enzymes of fatty acid, triglyceride and hydrocarbon metabolism are required in oenocytes primarily for lipid droplet induction rather than for cell growth. In contrast, a very long chain fatty-acyl-CoA reductase (FarO) and a putative lipid dehydrogenase/reductase (Spidey, also known as Kar) not only promote lipid droplet induction but also inhibit oenocyte growth. In the case of Spidey/Kar, we show that the growth suppression mechanism involves inhibition of the PI3K signaling pathway upstream of Akt activity. Together, the findings in this study show how Spidey/Kar and FarO regulate the balance between the cell growth and lipid storage of larval oenocytes.

Antioxidant Role for Lipid Droplets in a Stem Cell Niche of Drosophila.

Stem cells reside in specialized microenvironments known as niches. During Drosophila development, glial cells provide a niche that sustains the proliferation of neural stem cells (neuroblasts) during starvation. We now find that the glial cell niche also preserves neuroblast proliferation under conditions of hypoxia and oxidative stress. Lipid droplets that form in niche glia during oxidative stress limit the levels of reactive oxygen species (ROS) and inhibit the oxidation of polyunsaturated fatty acids (PUFAs). These droplets protect glia and also neuroblasts from peroxidation chain reactions that can damage many types of macromolecules. The underlying antioxidant mechanism involves diverting PUFAs, including diet-derived linoleic acid, away from membranes to the core of lipid droplets, where they are less vulnerable to peroxidation. This study reveals an antioxidant role for lipid droplets that could be relevant in many different biological contexts.

Hox proteins drive cell segregation and non-autonomous apical remodelling during hindbrain segmentation.

Hox genes encode a conserved family of homeodomain transcription factors regulating development along the major body axis. During embryogenesis, Hox proteins are expressed in segment-specific patterns and control numerous different segment-specific cell fates. It has been unclear, however, whether Hox proteins drive the epithelial cell segregation mechanism that is thought to initiate the segmentation process. Here, we investigate the role of vertebrate Hox proteins during the partitioning of the developing hindbrain into lineage-restricted units called rhombomeres. Loss-of-function mutants and ectopic expression assays reveal that Hoxb4 and its paralogue Hoxd4 are necessary and sufficient for cell segregation, and for the most caudal rhombomere boundary (r6/r7). Hox4 proteins regulate Eph/ephrins and other cell-surface proteins, and can function in a non-cell-autonomous manner to induce apical cell enlargement on both sides of their expression border. Similarly, other Hox proteins expressed at more rostral rhombomere interfaces can also regulate Eph/ephrins, induce apical remodelling and drive cell segregation in ectopic expression assays. However, Krox20, a key segmentation factor expressed in odd rhombomeres (r3 and r5), can largely override Hox proteins at the level of regulation of a cell surface target, Epha4. This study suggests that most, if not all, Hox proteins share a common potential to induce cell segregation but in some contexts this is masked or modulated by other transcription factors.

The development and functions of oenocytes.

Oenocytes have intrigued insect physiologists since the nineteenth century. Many years of careful but mostly descriptive research on these cells highlights their diverse sizes, numbers, and anatomical distributions across Insecta. Contemporary molecular genetic studies in Drosophila melanogaster and Tribolium castaneum support the hypothesis that oenocytes are of ectodermal origin. They also suggest that, in both short and long germ-band species, oenocytes are induced from a Spalt major/Engrailed ectodermal zone by MAPK signaling. Recent glimpses into some of the physiological functions of oenocytes indicate that they involve fatty acid and hydrocarbon metabolism. Genetic studies in D. melanogaster have shown that larval oenocytes synthesize very-long-chain fatty acids required for tracheal waterproofing and that adult oenocytes produce cuticular hydrocarbons required for desiccation resistance and pheromonal communication. Exciting areas of future research include the evolution of oenocytes and their cross talk with other tissues involved in lipid metabolism such as the fat body.