Cell Membrane Homeostasis in Mammals - The roles of ADIPOR2 and TLCD1 & 2

Sammanfattning: Ten nanometers. That is the approximate thickness of the plasma membrane that separates the extracellular space from the entire cellular machinery. An asymmetrical bilayer consisting of predominantly phospholipids in which proteins and carbohydrates are anchored and exert their function. Together, these components determine the viscosity, thickness, fluidity, permeability and packing of the plasma membrane, which is constantly maintained within a near-optimal chemical composition for proper function, a phenomenon also known as membrane homeostasis. Dietary fatty acids become the building block for the major component of the membrane bilayer, namely the phospholipids, and its composition can reflect the composition in the food, giving some validity to the term “you are what you eat”. To mitigate the huge variation in dietary fatty acids ending up in the membrane, it is reasonable to believe that regulatory mechanisms exist that adjust the fatty acid composition when required. Surprisingly, not much is known regarding how such adaptive responses regulate membrane fatty acid composition on a molecular level. Within this thesis, novel insights into such regulatory mechanisms are presented. Founded on genetic modifications using CRISPR/Cas9, lipidomic analyses, and membrane fluidity measurements, we build upon previous work and provide further evidence implicating the mammalian adiponectin receptor 2 (ADIPOR2) as a master regulator of membrane homeostasis. Human cells that lack ADIPOR2 show a dramatic vulnerability to saturated fat (SFA) exposure, leading to a defective translational response, increased ER stress, impaired mitochondrial respiration, and importantly, a massive increase in SFA-containing phospholipids. By exploiting this phenotype, genetic ablation of ADIPOR2 in human and mouse pre-adipocytes led to cells with drastic elevation in SFA-containing phospholipids. These were deployed to study the effect of phospholipid saturation on insulin signaling in vitro and in vivo. Surprisingly, both human and mouse adipocytes showed normal insulin signaling despite excess SFA content in their phospholipids, thus highlighting the robustness of adipocytes in lipid handling. Separately, genetic suppressors of SFA vulnerability in PAQR-2 (homolog to mammalian ADIPOR2) deficient C. elegans worms, revealed a novel protein dubbed FLD-1 that influences the amount of fluidizing, long-chain polyunsaturated fatty acids (PUFAs) in membrane phospholipids. Further studies in transgenic mice, lacking the mammalian homologs TLCD1 and TLCD2, led to elucidating the mechanism of action of these proteins, namely that they regulate the incorporation of monounsaturated fatty acids (MUFAs) at the sn-1 position of phosphatidylethanolamines. In parallel, another genetic suppressor to the SFA vulnerability of PAQR-2 knockouts was discovered, namely ACS-13, that was shown to regulate mitochondrial activation of long-chain PUFAs, a function conserved in the human homolog ACSL1. Taken together, this thesis provides new insight into the mechanisms that regulate the fatty acid composition of cellular membranes and that are crucial for the ability of cells to maintain fluid membranes in the face of fluctuating levels of dietary fatty acids.