Thierry Ferreira

Lipid-Induced ER Stress: Synergistic Effects of Sterols and Saturated Fatty Acids

Stress within the endoplasmic reticulum (ER) induces a coordinated response, namely the unfolded protein response (UPR), devoted to helping the ER cope with the accumulation of misfolded proteins. Failure of the UPR plays an important role in several human diseases. Recent studies report that intracellular accumulation of saturated fatty acids (SFAs) and cholesterol, seen in diseases of high incidence, such as obesity or atherosclerosis, results in ER stress. In the present study, we evaluated the effects of perturbations to lipid homeostasis on ER stress/UPR induction in the model eukaryote Saccharomyces cerevisiae. We show that SFA originating from either endogenous (preclusion of fatty acid desaturation) or exogenous (feeding with extracellular SFA) sources trigger ER stress and that ergosterol, the major sterol in yeast, acts synergistically with SFA in this process. This latter effect is connected to ergosterol accumulation within microsomal fractions from SFA‐accumulating cells, which display highly saturated phospholipid content. Moreover, treating the cells with the molecular chaperone 4‐phenyl butyrate abolishes UPR induction, suggesting that lipid‐induced ER stress leads to an overload of misfolded protein that acts, in turn, as the molecular signal for induction of the UPR. The present data are discussed in the context of human diseases that involve lipid deregulation.

Polyunsaturated phospholipids facilitate membrane deformation and fission by endocytic proteins

Bending the benefits of polyunsaturates We have often heard that it is beneficial to eat polyunsaturated fatty acids. We also know that some organelles such as synaptic vesicles are extremely rich in polyunsaturated lipids. However, what polyunsaturated lipids do in our body is unclear. Using cell biology, biochemical reconstitutions, and molecular dynamics, Pinot et al. show that polyunsaturated phospholipids can change the response of membranes to proteins involved in membrane curvature sensing, membrane shaping, and membrane fission. Polyunsaturated phospholipids make the plasma membrane more amenable to deformation; facilitate endocytosis; and, in reconstitution experiments, increased membrane fission by the dynamin-endophilin complex. Science, this issue p. 693 Certain membrane lipids adapt their conformation to membrane curvature, facilitating membrane deformation and fission. Phospholipids (PLs) with polyunsaturated acyl chains are extremely abundant in a few specialized cellular organelles such as synaptic vesicles and photoreceptor discs, but their effect on membrane properties is poorly understood. Here, we found that polyunsaturated PLs increased the ability of dynamin and endophilin to deform and vesiculate synthetic membranes. When cells incorporated polyunsaturated fatty acids into PLs, the plasma membrane became more amenable to deformation by a pulling force and the rate of endocytosis was accelerated, in particular, under conditions in which cholesterol was limiting. Molecular dynamics simulations and biochemical measurements indicated that polyunsaturated PLs adapted their conformation to membrane curvature. Thus, by reducing the energetic cost of membrane bending and fission, polyunsaturated PLs may help to support rapid endocytosis.

SUT1-promoted sterol uptake involves the ABC transporter Aus1 and the mannoprotein Dan1 whose synergistic action is sufficient for this process

Efficient sterol influx in the yeast Saccharomyces cerevisiae is restricted to anaerobiosis or to haem deficiency resulting from mutations. Constitutive expression of SUT1, an hypoxic gene encoding a transcriptional regulator, induces sterol uptake in aerobiosis. A genome-wide approach using DNA microarray was used to identify the mediators of SUT1 effects on aerobic sterol uptake. A total of 121 ORFs (open reading frames) were significantly and differentially expressed after SUT1 overexpression, 61 down-regulated and 60 up-regulated. Among these genes, the role of the putative ABC transporter (ATP-binding-cassette transporter) Aus1, and of the cell-wall mannoprotein Dan1, was characterized better. These two genes play an essential role in aerobic sterol uptake, since their deletion compromised the SUT1 effects, but individual overexpression of either of these genes in a wild-type background was not sufficient for this process. However, constitutive co-expression of AUS1 and DAN1 in a wild-type background resulted in sterol influx in aerobiosis. These results suggest that the corresponding proteins may act synergistically in vivo to promote sterol uptake.

ERGOSTEROL BIOSYNTHESIS: A FUNGAL PATHWAY FOR LIFE ON LAND?

Sterols, essential lipids of most eukaryotic cells, ensure important structural and signaling functions. The selection pressure that has led to different dominant sterols in the three eukaryotic kingdoms remains unknown. Here, we investigated the influence of the progression in the different steps of the ergosterol biosynthetic pathway (EBP) on the yeast resistance to transitions from aqueous to aerial media, typical perturbations of the higher fungi habitats. Five mutants of the EBP (ergΔ), accumulating different sterol intermediates in the EBP, and the wild‐type (WT) strain were exposed to drying under atmospheric air or nitrogen and wetting. Results show that the progression in the EBP parallels an increase in the yeast resistance to air‐drying with a maximal survival rate for the WT strain. When drying/wetting was performed under nitrogen, yeast survival was higher, particularly for the earlier mutants of the EBP. Thus, ergosterol, through its protective role against mechanical and oxidative stress, might have been selected by the pressure induced by drying/wetting cycles occurring in the fungi habitats. These results support the Bloch hypothesis, which postulates that the properties of sterols are gradually optimized for function along the biosynthetic pathway and provide a response to the enduring question “why ergosterol in fungi?”.

Modulation of Lipid-Induced ER Stress by Fatty Acid Shape

Exposure of pancreatic β cells to long‐chain saturated fatty acids (SFA) induces a so‐called endoplasmic reticulum (ER) stress that can ultimately lead to cell death. This process is believed to participate in insulin deficiency associated with type 2 diabetes, via a decrease in β‐cell mass. By contrast, some unsaturated fatty acid species appear less toxic to the cells and can even alleviate SFA‐induced ER stress. In the present study, we took advantage of a simple yeast‐based model, which brings together most of the trademarks of lipotoxicity in human cells, to screen fatty acids of various structures for their capacity to counter ER stress. Here we demonstrate that the tendency of a free fatty acid (FFA) to reduce SFA toxicity depends on a complex conjunction of parameters, including chain length, level of unsaturation, position of the double bonds and nature of the isomers (cis or trans). Interestingly, potent FFA act as building blocks for phospholipid synthesis and help to restore an optimal membrane organization, compatible with ER function and normal protein trafficking.

Is fat so bad? Modulation of endoplasmic reticulum stress by lipid droplet formation

LDs (lipid droplets) have long been considered as inert particles used by the cells to store fatty acids and sterols as esterified non‐toxic lipid species (i.e. triacylglycerols and steryl esters). However, accumulating evidence suggests that LDs behave as a dynamic compartment, which is involved in the regulation of several aspects of the homoeostasis of their originating organelle, namely the ER (endoplasmic reticulum). The ER is particularly sensitive to physiological/pathological stimuli, which can ultimately induce ER stress. In the present review, after considering the basic mechanisms of LD formation and the signal cascades leading to ER stress, we focus on the connections between these two pathways. Taking into consideration recent data from the literature, we will try to draw possible mechanisms for the role of LDs in the regulation of ER homoeostasis and in ER‐stress‐related diseases.

Nature of sterols affects plasma membrane behavior and yeast survival during dehydration.

The plasma membrane (PM) is a main site of injury during osmotic perturbation. Sterols, major lipids of the PM structure in eukaryotes, are thought to play a role in ensuring the stability of the lipid bilayer during physicochemical perturbations. Here, we investigated the relationship between the nature of PM sterols and resistance of the yeast Saccharomyces cerevisiae to hyperosmotic treatment. We compared the responses to osmotic dehydration (viability, sterol quantification, ultrastructure, cell volume, and membrane permeability) in the wild-type (WT) strain and the ergosterol mutant erg6Δ strain. Our main results suggest that the nature of membrane sterols governs the mechanical behavior of the PM during hyperosmotic perturbation. The mutant strain, which accumulates ergosterol precursors, was more sensitive to osmotic fluctuations than the WT, which accumulates ergosterol. The hypersensitivity of erg6Δ was linked to modifications of the membrane properties, such as stretching resistance and deformation, which led to PM permeabilization during the volume variation during the dehydration-rehydration cycles. Anaerobic growth of erg6Δ strain with ergosterol supplementation restored resistance to osmotic treatment. These results suggest a relationship between hydric stress resistance and the nature of PM sterols. We discuss this relationship in the context of the evolution of the ergosterol biosynthetic pathway.

Lipid dynamics in yeast under haem-induced unsaturated fatty acid and/or sterol depletion

In the yeast Saccharomyces cerevisiae, UFA (unsaturated fatty acids) and ergosterol syntheses are aerobic processes that require haem. We took advantage of a strain affected in haem synthesis ( hem1 Delta) to starve specifically for one or the other of these essential lipids in order to examine the consequences on the overall lipid composition. Our results demonstrate that reserve lipids (i.e. triacylglycerols and steryl esters) are depleted independently of haem availability and that their UFA and sterol content is not crucial to sustain residual growth under lipid depletion. In parallel to UFA starvation, a net accumulation of SFA (saturated fatty acids) is observed as a consequence of haem biosynthesis preclusion. Interestingly, the excess SFA are not mainly stored within triacylglycerols and steryl esters but rather within specific phospholipid species, with a marked preference for PtdIns. This results in an increase in the cellular PtdIns content. However, neutral lipid homoeostasis is perturbed under haem starvation. The contribution of two lipid particle-associated proteins (namely Tgl1p and Dga1p) to this process is described.

From zero to six double bonds: phospholipid unsaturation and organelle function

Cellular phospholipids (PLs) differ by the nature of their polar heads as well as by the length and unsaturation level of their fatty acyl chains. We discuss how the ratio between saturated, monounsaturated, and polyunsaturated PLs impacts on the functions of such organelles as the endoplasmic reticulum, synaptic vesicles, and photoreceptor discs. Recent experiments and simulations suggest that polyunsaturated PLs respond differently to mechanical stress, including membrane bending, than monounsaturated PLs owing to their unique conformational plasticity. These findings suggest a rationale for PL acyl chain remodeling by acyltransferases and a molecular explanation for the importance of a balanced fatty acid diet.

Lipid-induced ER stress in yeast and β cells: parallel trails to a common fate

Exposure to long-chain saturated fatty acids (SFAs; e.g. palmitate) induces apoptosis in pancreatic β cells, a process that may contribute to the development of type 2 diabetes. Under palmitate treatment, β cells undergo a so-called endoplasmic reticulum (ER) stress that can be counteracted by the unfolded protein response (UPR). The UPR is a coordinated response, which is primarily devoted to helping the ER to cope with the accumulation of misfolded proteins. Sustained SFA exposure may ultimately overwhelm the UPR, resulting in cell death. By contrast, unsaturated fatty acids (e.g. oleate) are much less harmful to the cells and can even alleviate palmitate toxicity. Surprisingly, recent evidences indicate that a simple unicellular eukaryote, the budding yeast Saccharomyces cerevisiae, which is not routinely exposed to high-fat diets, also undergoes ER stress under lipotoxic conditions. This suggests that the mechanisms of SFA toxicity are largely conserved throughout eukaryotes and are not specific of a given cell type. The present review discusses the mechanisms of SFA toxicity in yeast and β cells, with a main emphasis on their potential impacts on ER-membrane organization/function and ER-based processes.