Role of phosphatidic acid phosphatase 2a in uptake of extracellular lipid phosphate mediators.
Phosphatidic acid (PA) phosphatase 2a (PAP2a) is an integral membrane glycoprotein that hydrolyzes a number of structurally related lipid phosphate substrates when presented in mixed phospholipid and detergent micelles. The physiological substrate specificity and functions of this enzyme are unclear. Using reconstitution studies we demonstrate that PAP2a hydrolyses both PA and LysoPA substrates in a lipid bilayer. To investigate the activity of PAP2a against cellular substrates we generated HEK293 cell variants stably overexpressing the enzyme. Although one of these lines exhibited a 27-fold increase in PAP2 activity measured in vitro, levels of PA were not significantly reduced in comparison with control cells. Cell surface labeling and activity measurements demonstrate that a portion of the enzyme was localized to the cell surface. Pagano and Longmuir (J. Biol. Chem. 260 (1985) 1909) described the rapid uptake of PA by cultured cells, but the mechanisms and proteins involved were not identified. We found that overexpression of PAP2a was accompanied by a 2.1-fold increase in uptake of a fluorescent PA analog but that uptake of other phospholipids and diacylglycerols was unaltered. The increase in lipid uptake was completely dependent on PAP activity and unaffected by endocytosis inhibitors. Our results indicate that PAP2a is a cell surface enzyme that plays an active role in the hydrolysis and uptake of lipids from the extracellular space.
Roberts
R Z
RZ
Department of Pharmacological Sciences, Institute for Cell and Developmental Biology, Stony Brook Health Sciences Center, Stony Brook, NY, USA.
Morris
A J
AJ
D002460:Cell Line; D002462:Cell Membrane / Q000201:enzymology; D004337:Drug Carriers; D004705:Endocytosis; D005822:Genetic Vectors; D006801:Humans; D008051:Lipid Bilayers / Q000378:metabolism; D008081:Liposomes; D008856:Microscopy, Fluorescence; D008956:Models, Chemical; D010711:Phosphatidate Phosphatase / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D010712:Phosphatidic Acids / Q000378:metabolism* / Q000494:pharmacology; D010743:Phospholipids / Q000378:metabolism; D011994:Recombinant Proteins / Q000378:metabolism; D013379:Substrate Specificity; D014162:Transfection
D004337:Drug Carriers; D008051:Lipid Bilayers; D008081:Liposomes; D010712:Phosphatidic Acids; D010743:Phospholipids; D011994:Recombinant Proteins; C406242:phosphatidic acid phosphatase type 2; D010711:Phosphatidate Phosphatase