Phospholipid flippases.
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ATP-Binding Cassette Transporters↗
Biomedical subjects
David L Daleke
Publications and source records attributed to David L Daleke.
Lipid specific activation of the murine P4-ATPase Atp8a1 (ATPase II).
The asymmetric transbilayer distribution of phosphatidylserine (PS) in the mammalian plasma membrane and secretory vesicles is maintained, in part, by an ATP-dependent transporter. This aminophospholipid "flippase" selectively transports PS to the cytosolic leaflet of the bilayer and is sensitive to vanadate, Ca(2+), and modification by sulfhydryl reagents. Although the flippase has not been positively identified, a subfamily of P-type ATPases has been proposed to function as transporters of amphipaths, including PS and other phospholipids. A candidate PS flippase ATP8A1 (ATPase II), originally isolated from bovine secretory vesicles, is a member of this subfamily based on sequence homology to the founding member of the subfamily, the yeast protein Drs2, which has been linked to ribosomal assembly, the formation of Golgi-coated vesicles, and the maintenance of PS asymmetry. To determine if ATP8A1 has biochemical characteristics consistent with a PS flippase, a murine homologue of this enzyme was expressed in insect cells and purified. The purified Atp8a1 is inactive in detergent micelles or in micelles containing phosphatidylcholine, phosphatidic acid, or phosphatidylinositol, is minimally activated by phosphatidylglycerol or phosphatidylethanolamine (PE), and is maximally activated by PS. The selectivity for PS is dependent upon multiple elements of the lipid structure. Similar to the plasma membrane PS transporter, Atp8a1 is activated only by the naturally occurring sn-1,2-glycerol isomer of PS and not the sn-2,3-glycerol stereoisomer. Both flippase and Atp8a1 activities are insensitive to the stereochemistry of the serine headgroup. Most modifications of the PS headgroup structure decrease recognition by the plasma membrane PS flippase. Activation of Atp8a1 is also reduced by these modifications; phosphatidylserine-O-methyl ester, lysophosphatidylserine, glycerophosphoserine, and phosphoserine, which are not transported by the plasma membrane flippase, do not activate Atp8a1. Weakly translocated lipids (PE, phosphatidylhydroxypropionate, and phosphatidylhomoserine) are also weak Atp8a1 activators. However, N-methyl-phosphatidylserine, which is transported by the plasma membrane flippase at a rate equivalent to PS, is incapable of activating Atp8a1 activity. These results indicate that the ATPase activity of the secretory granule Atp8a1 is activated by phospholipids binding to a specific site whose properties (PS selectivity, dependence upon glycerol but not serine, stereochemistry, and vanadate sensitivity) are similar to, but distinct from, the properties of the substrate binding site of the plasma membrane flippase.
Adenosine Triphosphatases↗
Interaction between phosphatidylserine and the phosphatidylserine receptor inhibits immune responses in vivo.
Phosphatidylserine (PS) on apoptotic cells promotes their uptake and induces anti-inflammatory responses in phagocytes, including TGF-beta release. Little is known regarding the effects of PS on adaptive immune responses. We therefore investigated the effects of PS-containing liposomes on immune responses in mice in vivo. PS liposomes specifically inhibited responses to Ags as determined by decreased draining lymph node tissue mass, with reduced numbers of total leukocytes and Ag-specific CD4(+) T cells. There was also a decrease in formation and size of germinal centers in spleen and lymph nodes, accompanied by decreased levels of Ag-specific IgG in blood. Many of these effects were mimicked by an agonistic Ab-specific for the PS receptor. TGF-beta appears to play a critical role in this inhibition, as the inhibitory effects of PS were reversed by in vivo administration of anti-TGF-beta Ab. PS-containing liposomes did not appear to directly inhibit dendritic cell maturation in vitro in response to a variety of stimuli, nor did it prevent their migration to regional lymph nodes in vivo, suggesting that the inhibitory effects may have resulted from complicated interactions between tissue cells and dendritic cells, subsequently inhibiting their ability to productively activate T lymphocytes.
Adoptive Transfer↗
The role of oxidative stress in diabetic complications.
The morbidity and mortality associated with diabetes is the result of the myriad complications related to the disease. One of the most explored hypotheses to explain the onset of complications is a hyperglycemia-induced increase in oxidative stress. Reactive oxygen species (ROS) are produced by oxidative phosphorylation, nicotinamide adenine dinucleotide phosphate oxidase (NADPH), xanthine oxidase, the uncoupling of lipoxygenases, cytochrome P450 monooxygenases, and glucose autoxidation. Once formed, ROS deplete antioxidant defenses, rendering the affected cells and tissues more susceptible to oxidative damage. Lipid, DNA, and protein are the cellular targets for oxidation, leading to changes in cellular structure and function. Recent evidence suggests ROS are also important as second messengers in the regulation of intracellular signaling pathways and, ultimately, gene expression. This review explores the production of ROS and the propagation and consequences of oxidative stress in diabetes.
Animals↗
Purification of the human erythrocyte PS-stimulated Mg2+-ATPase: a putative PS flippase.
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Ca(2+) Mg(2+)-ATPase↗
Regulation of transbilayer plasma membrane phospholipid asymmetry.
Lipids in biological membranes are asymmetrically distributed across the bilayer; the amine-containing phospholipids are enriched on the cytoplasmic surface of the plasma membrane, while the choline-containing and sphingolipids are enriched on the outer surface. The maintenance of transbilayer lipid asymmetry is essential for normal membrane function, and disruption of this asymmetry is associated with cell activation or pathologic conditions. Lipid asymmetry is generated primarily by selective synthesis of lipids on one side of the membrane. Because passive lipid transbilayer diffusion is slow, a number of proteins have evolved to either dissipate or maintain this lipid gradient. These proteins fall into three classes: 1) cytofacially-directed, ATP-dependent transporters ("flippases"); 2) exofacially-directed, ATP-dependent transporters ("floppases"); and 3) bidirectional, ATP-independent transporters ("scramblases"). The flippase is highly selective for phosphatidylserine and functions to keep this lipid sequestered from the cell surface. Floppase activity has been associated with the ABC class of transmembrane transporters. Although they are primarily nonspecific, at least two members of this class display selectivity for their substrate lipid. Scramblases are inherently nonspecific and function to randomize the distribution of newly synthesized lipids in the endoplasmic reticulum or plasma membrane lipids in activated cells. It is the combined action of these proteins and the physical properties of the membrane bilayer that generate and maintain transbilayer lipid asymmetry.
Animals↗