Colocalization of leukotriene C synthase and microsomal glutathione S-transferase elucidated by indirect immunofluorescence analysis

We have previously shown that the two membrane bound enzymes leukotriene C synthase and microsomal glutathione S-transferase interact in vitro and in vivo, Rat basophilic leukemia cells and murine mastocytoma cells, two well- known sources of leukotriene C synthase, both expressed microsomal glutathi one S-transferase as determined by Western blot analyses, Several human tis sues were found to contain both leukotriene C synthase and microsomal gluta thione S-transferase mRNA, These data suggest that the interaction may be p hysiologically important, To study this further, expression vectors encodin g the two enzymes were cotransfected into mammalian cells and the subcellul ar localization of the enzymes was determined by indirect immunofluorescenc e using confocal laser scanning microscopy, The results showed that leukotr iene C synthase and microsomal glutathione S-transferase were both localize d on the nuclear envelope and adjacent parts of the endoplasmic reticulum, Image overlay demonstrated virtually identical localization. We also observ ed that coexpression substantially reduced the catalytic activity of each e nzyme suggesting that a mechanism involving protein-protein interaction may contribute to the regulation of LTC4 production. (C) 2000 Federation of Eu ropean Biochemical Societies. Published by Elsevier Science B.V. All rights reserved.