The mechanism of docosahexaenoic acid-induced phospholipase D activation in human lymphocytes involves exclusion of the enzyme from lipid rafts.
Docosahexaenoic acid (DHA), an n-3 polyunsaturated fatty acid that inhibits T lymphocyte activation, has been shown to stimulate phospholipase D (PLD) activity in stimulated human peripheral blood mononuclear cells (PBMC). To elucidate the mechanisms underlying the DHA-induced PLD activation, we first characterized the PLD expression pattern of PBMC. We show that these cells express PLD1 and PLD2 at the protein and mRNA level and are devoid of oleate-dependent PLD activity. DHA enrichment of PBMC increased the DHA content of cell phospholipids, which was directly correlated with the extent of PLD activation. The DHA-induced PLD activation was independent of conventional protein kinase C but inhibited by brefeldin A, which suggests ADP-ribosylation factor (ARF)-dependent mechanism. Furthermore, DHA enrichment dose-dependently stimulated ARF translocation to cell membranes. Whereas 50% of the guanosine 5'-3-O-(thio)triphosphate plus ARF-dependent PLD activity and a substantial part of PLD1 protein were located to the detergent-insoluble membranes, so-called rafts, of non-enriched PBMC, DHA treatment strongly displaced them toward detergent-soluble membranes where ARF is present. Collectively, these results suggest that the exclusion of PLD1 from lipid rafts, due to their partial disorganization by DHA, and its relocalization in the vicinity of ARF, is responsible for its activation. This PLD activation might be responsible for the immunosuppressive effect of DHA because it is known to transmit antiproliferative signals in lymphoid cells.
Unité INSERM 352, Laboratoire de Biochimie et Pharmacologie, INSA de Lyon, 69621 Villeurbanne, France.
Olivier
O
Diaz
Alexandre
A
Berquand
Madeleine
M
Dubois
Silvia
S
Di Agostino
Claudio
C
Sette
Sylvain
S
Bourgoin
Michel
M
Lagarde
Georges
G
Nemoz
Annie-France
AF
Prigent
D020727:ADP-Ribosylation Factors / Q000378:metabolism; D015153:Blotting, Western; D020126:Brefeldin A / Q000494:pharmacology; D002455:Cell Division; D002474:Cell-Free System; D002478:Cells, Cultured; D003902:Detergents / Q000494:pharmacology; D004281:Docosahexaenoic Acids / Q000378:metabolism*; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation; D006160:Guanosine Triphosphate / Q000378:metabolism; D006801:Humans; D007963:Leukocytes, Mononuclear / Q000378:metabolism; D008214:Lymphocytes / Q000201:enzymology / Q000378:metabolism*; D021962:Membrane Microdomains / Q000378:metabolism*; D010739:Phospholipase D / Q000378:metabolism*; D010743:Phospholipids / Q000378:metabolism; D011233:Precipitin Tests; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D021381:Protein Transport; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013997:Time Factors
D003902:Detergents; D010743:Phospholipids; D020126:Brefeldin A; D004281:Docosahexaenoic Acids; D006160:Guanosine Triphosphate; D010739:Phospholipase D; D020727:ADP-Ribosylation Factors