Leukotriene synthesis in response to A23187 is inhibited by methyl- beta-cyclodextrin in RBL-2H3 cells.
Leukotrienes (LTs) are produced by several biosynthetic enzymes including cytosolic phospholipase A2 (cPLA2), 5-lipoxygenase (5-LO), and 5-lipoxygenase activating protein (FLAP) in the perinuclear area. In the present study, we showed that pretreatment with methyl-beta-cyclodextrin (MbetaCD), a cholesterol-depleting agent, dramatically reduced the synthesis of LTs in response to A23187 in mast cells. A23187-induced LT synthesis was inhibited by pretreatment with MbetaCD, and this effect was reversed when cholesterol was added. In an approach to identifying the MbetaCD-sensitive protein(s), we observed that FLAP co-localized with flotillin-1, a lipid raft marker protein, in the lipid raft-rich low-density region of sucrose gradients. In addition, electron microscopic analysis revealed that FLAP co-localized with flotillin-1. Together, these results suggest that FLAP is present in cholesterol-rich lipid raft-like domains and that its localization in these domains is critical for LT synthesis.
You
Hye Jin
HJ
School of Life Sciences and Biotechnology, Korea University, Seoul 136-701, Korea.
Seo
Ji-Min
JM
Moon
Ji-Young
JY
Han
Sung-Sik
SS
Ko
Young-Gyu
YG
Kim
Jae-Hong
JH
D058927:5-Lipoxygenase-Activating Proteins; D000818:Animals; D016718:Arachidonic Acid / Q000378:metabolism; D000001:Calcimycin / Q000494:pharmacology*; D002118:Calcium / Q000378:metabolism; D002352:Carrier Proteins / Q000648:ultrastructure; D045744:Cell Line, Tumor; D003545:Cysteine / Q000096:biosynthesis; D015289:Leukotrienes / Q000096:biosynthesis*; D021962:Membrane Microdomains / Q000187:drug effects / Q000648:ultrastructure; D008565:Membrane Proteins / Q000648:ultrastructure; D021381:Protein Transport / Q000187:drug effects; D051381:Rats; D047392:beta-Cyclodextrins / Q000494:pharmacology*
D058927:5-Lipoxygenase-Activating Proteins; D002352:Carrier Proteins; D015289:Leukotrienes; D008565:Membrane Proteins; D047392:beta-Cyclodextrins; C112381:cysteinyl-leukotriene; C106354:flotillins; C108732:methyl-beta-cyclodextrin; D016718:Arachidonic Acid; D000001:Calcimycin; D003545:Cysteine; D002118:Calcium