Good Fat, Essential Cellular Requirements for Triacylglycerol Synthesis to Maintain Membrane Homeostasis in Yeast

Abstract

Storage triacylglycerols (TAG) and membrane phospholipids share common precursors, i.e. phosphatidic acid and diacylglycerol, in the endoplasmic reticulum. In addition to providing a biophysically rather inert storage pool for fatty acids, TAG synthesis plays an important role to buffer excess fatty acids (FA). The inability to incorporate exogenous oleic acid into TAG in a yeast mutant lacking the acyltransferases Lro1p, Dga1p, Are1p, and Are2p contributing to TAG synthesis results in dysregulation of lipid synthesis, massive proliferation of intracellular membranes, and ultimately cell death. Carboxypeptidase Y trafficking from the endoplasmic reticulum to the vacuole is severely impaired, but the unfolded protein response is only moderately up-regulated, and dispensable for membrane proliferation, upon exposure to oleic acid. FA-induced toxicity is specific to oleic acid and much less pronounced with palmitoleic acid and is not detectable with the saturated fatty acids, palmitic and stearic acid. Palmitic acid supplementation partially suppresses oleic acid-induced lipotoxicity and restores carboxypeptidase Y trafficking to the vacuole. These data show the following: (i) FA uptake is not regulated by the cellular lipid requirements; (ii) TAG synthesis functions as a crucial intracellular buffer for detoxifying excess unsaturated fatty acids; (iii) membrane lipid synthesis and proliferation are responsive to and controlled by a balanced fatty acid composition.In the aqueous cellular environment, fatty acyl chains esterified in glycerophospholipids constitute the hydrophobic barrier of biological membranes. Thus, fatty acid (FA) 3 composition is a crucial determinant of cellular membrane function. Establishment of the specific FA profiles in lipid species of various organelle membranes (1) relies on an intricate balance between endogenous FA synthesis, recycling of FA from lipid breakdown, and perhaps uptake from the exterior. Glycerophospholipids and triacylglycerols (TAG), which serve as the major storage form of FA, share the similar precursors phosphatidic acid (PA) and diacylglycerol (DAG), both generated in the endoplasmic reticulum (ER) membrane. TAG are packaged into lipid droplets and are thus sequestered away from the ER membrane by a mechanism not yet understood. In addition, membranes and lipid storage pools (2, 3) undergo significant turnover and intracellular flux, e.g. during secretion or endocytosis and cellular growth, which must be accounted for by mechanisms that establish and maintain lipid homeostasis in these dynamic membrane systems (4). We have recently shown that TAG degradation provides metabolites that are critical for efficient cell cycle progression at the G 1

Arabidopsis Lipins, PDAT1 Acyltransferase, and SDP1 Triacylglycerol Lipase Synergistically Direct Fatty Acids toward β-Oxidation, Thereby Maintaining Membrane Lipid Homeostasis

“…The increase in membrane lipid content was also observed in tgd1-1 disrupted in PDAT1 (Fan et al, 2013a) and in yeast mutants defective in TAG synthesis (Petschnigg et al, 2009;Fakas et al, 2011). Such an increase has been regarded as an alternative means to protect against lipotoxicity due to lipid overload (Petschnigg et al, 2009;Fakas et al, 2011).…”

“…The increase in membrane lipid content was also observed in tgd1-1 disrupted in PDAT1 (Fan et al, 2013a) and in yeast mutants defective in TAG synthesis (Petschnigg et al, 2009;Fakas et al, 2011). Such an increase has been regarded as an alternative means to protect against lipotoxicity due to lipid overload (Petschnigg et al, 2009;Fakas et al, 2011). Because phospholipid levels are elevated in both tgd1-1 sdp1-4, in which TAG hydrolysis is impaired, and tgd1-1 pxa1-2, in which FA degradation is impaired, it is unlikely that this increase is due to the augmented recycling to TAG into membrane lipids as a result of blocked FA turnover.…”

“…Under conditions of acute lipid overload, excess FAs are converted into biologically inert neutral lipids and packaged in specific subcellular organelles named lipid droplets (LDs) (Listenberger et al, 2003;Petschnigg et al, 2009;Kohlwein, 2010;Murphy, 2012;Kohlwein et al, 2013). In yeast (Saccharomyces cerevisiae), a critical enzyme involved in TAG synthesis (Han et al, 2006) and LD formation (Adeyo et al, 2011) is the Mg 2+ -dependent phosphatidic acid phosphohydrolase (Pah1), the homolog of the mammalian lipin protein family.…”

Three Acyltransferases and Nitrogen-responsive Regulator Are Implicated in Nitrogen Starvation-induced Triacylglycerol Accumulation in Chlamydomonas

“…We tested the biochemical activities of DGTT1 and PDAT1 in a heterologous system by complementation of an S. cerevisiae mutant carrying a quadruple deletion in four acyltransferases (encoded by DGA1, LRO1, ARE1, and ARE2) that contribute to TAG synthesis (75). The resulting strain is sensitive to oleic acid, and introduction of either a DGAT-or PDAT-encoding sequence will rescue the strain (76).…”

Combinatorial Engineering of Yarrowia lipolytica as a Promising Cell Biorefinery Platform for the de novo Production of Multi-Purpose Long Chain Dicarboxylic Acids

“…Our genetic engineering did not involve the disruption of TAG biosynthesis. This is because the generation of free fatty acids, particularly unsaturated ones, causes toxicity in the cell with deficient lipid body formation [87]. This occurs for several reasons: the destabilizing effect of fatty acid overproduction on the cellular membrane [80,88], the harmful effects of oxidative species derived from the oxidation of unsaturated fatty acids [89], and a lack of protection against excess lipid intermediates [90].…”