Phosphatidylethanolamine positively regulates autophagy and longevity

Autophagy is a cellular recycling program that retards ageing by efficiently eliminating damaged and potentially harmful organelles and intracellular protein aggregates. Here, we show that the abundance of phosphatidylethanolamine (PE) positively regulates autophagy. Reduction of intracellular PE levels by knocking out either of the two yeast phosphatidylserine decarboxylases (PSD) accelerated chronological ageing-associated production of reactive oxygen species and death. Conversely, the artificial increase of intracellular PE levels, by provision of its precursor ethanolamine or by overexpression of the PE-generating enzyme Psd1, significantly increased autophagic flux, both in yeast and in mammalian cell culture. Importantly administration of ethanolamine was sufficient to extend the lifespan of yeast (Saccharomyces cerevisiae), mammalian cells (U2OS, H4) and flies (Drosophila melanogaster). We thus postulate that the availability of PE may constitute a bottleneck for functional autophagy and that organismal life or healthspan could be positively influenced by the consumption of ethanolamine-rich food.Phosphatidylethanolamine (PE) is a phospholipid found in all living organisms. Together with phosphatidylcholine (PC), phosphatidylserine (PS) and phosphatidylinositol (PI), PE represents the backbone of most biological membranes. PE is the second-most abundant phospholipid in mammalian membranes ranging from 20 to 50%.¹ In yeast, PE is essential for growth and is generated through four different enzymatic pathways:² PE can be produced by decarboxylation of PS, as a first option at the mitochondrial membrane via phosphatidylserine decarboxylase 1 (Psd1)^{3, 4} or, as a second, option at the Golgi and vacuolar membranes through phosphatidylserine decarboxylase 2 (Psd2).⁵ As a third possibility, PE can be produced from actively retrieved extracellular ethanolamine,^{6, 7} which is cytidine 5''-diphosphate-activated⁸ and then coupled to diacylglycerol to generate PE.⁹ The fourth, scarcely employed PE-generating pathway is based on the lysophospholipid acylation of lyso-PE. Importantly, PE does not spontaneously assemble in bilayers and rather incorporates into curved structures, such as the inverted hexagonal phase.¹⁰ The physiological function of non-bilayer lipids in membranes is considered to reside in their interaction with membrane proteins via the membrane lateral pressure¹⁰ and membrane tethering and fusion processes, which are relevant for autophagy.¹¹The term ‘autophagy'' describes a degradation process affecting intracellular components (for a review see, 12 13) which as an important cytoprotective mechanism, is closely linked to ageing. Autophagy mainly differs from the proteasomal pathway, the other major cellular degradation mechanism, in two aspects. First, autophagy can degrade large particles or whole organelles and second, the final degradation occurs in the lysosome/vacuole and not at the proteasome. Prior to the actual degradation, the cargo is gathered in autophagic particles, which are surrounded by a characteristic double-membrane. However, the origin of these autophagosomal membranes is still controversial and might actually depend on the mode of autophagy induction.^{14, 15} Among the discussed membrane sources are the Golgi apparatus, the endosplamic reticulum (ER) or the mitochondrion-associated membrane, which is formed at the interface between the ER.¹⁶ In higher eukaryotes autophagic membranes are enriched in PE with a high degree of unsaturation,¹⁷ similarly to the PE species found in mitochondria.^{14, 18} Moreover, the pre-autophagosomal structure or phagophore assembly site (PAS), which appears at the very beginning of autophagosome formation, already harbours Atg9, an autophagy-related transmembrane protein that shuttles between mitochondria and the PAS structure in yeast.¹⁹Importantly, PE also functions as an anchor to autophagosomal membranes for the autophagy-related protein Atg8 in yeast²⁰ and its mammalian orthologue LC3.^{21, 22} This PE anchor is provided to LC3/Atg8 post-translationally in a process called lipidation. First, LC3/Atg8 is carboxy-terminally cleaved by proteases from the Atg4 family.^{23, 24} Subsequently, the remaining C-terminal glycine is coupled to PE in a series of ubiquitination-like reactions involving diverse Atg-proteins.^{20, 25, 26, 27} In vitro, Atg8-PE causes hemifusion of vesicles, which argues for its potential role in autophagosomal phagophore expansion.^{11, 28} Consistently, semisynthetic LC3-PE has recently been described to stimulate membrane tethering and fusion.²⁹ We thus reasoned that the overall abundance of PE might be critical for PE-lipidation of LC3/Atg8 and could thus regulate autophagosomal membrane formation. Therefore, we tested whether increasing cellular PE levels might have an impact on autophagy and lifespan regulation.Here, we report that knock-out of PSD1 or PSD2 shortens the chronological lifespan of S. cerevisiae, whereas PSD1-overexpression enhances the autophagic capacity and increases longevity. Furthermore, external administration of ethanolamine increases endogenous PE levels, enhances autophagic flux and extends the lifespan of yeast, mammalian cells in culture and flies (Drosophila melanogaster).