title: Membrane localization and topology of leukotriene C4 synthase.
abstract: Leukotriene C(4) (LTC(4)) synthase conjugates LTA(4) with GSH to form LTC(4). Determining the site of LTC(4) synthesis and the topology of LTC(4) synthase may uncover unappreciated intracellular roles for LTC(4), as well as how LTC(4) is transferred to its export carrier, the multidrug resistance protein-1. We have determined the membrane localization of LTC(4) synthase by immunoelectron microscopy. In contrast to the closely related five-lipoxygenase-activating protein, LTC(4) synthase is distributed in the outer nuclear membrane and peripheral endoplasmic reticulum but is excluded from the inner nuclear membrane. We have combined immunofluorescence with differential membrane permeabilization to determine the topology of LTC(4) synthase. The active site of LTC(4) synthase is localized in the lumen of the nuclear envelope and endoplasmic reticulum. These results indicate that the synthesis of LTB(4) and LTC(4) occurs in different subcellular locations and suggests that LTC(4) must be returned to the cytoplasmic side of the membrane for export by multidrug resistance protein-1. The differential localization of two very similar integral membrane proteins suggests that mechanisms other than size-dependent exclusion regulate their passage to the inner nuclear membrane.
authors.affiliation: Renal Unit and Program in Membrane Biology, Department of Medicine, Harvard Medical School and Massachusetts General Hospital (East), Charlestown, Massachusetts 02129, USA.
authors.forename: Peter
authors.initials: P
authors.lastname: Christmas
authors.forename: Brittany M
authors.initials: BM
authors.lastname: Weber
authors.forename: Mary
authors.initials: M
authors.lastname: McKee
authors.forename: Dennis
authors.initials: D
authors.lastname: Brown
authors.forename: Roy J
authors.initials: RJ
authors.lastname: Soberman
mesh_terms: D000818:Animals; D015703:Antigens, CD / Q000378:metabolism; D016466:CHO Cells; D019556:COS Cells; D002462:Cell Membrane / Q000201:enzymology* / Q000378:metabolism; D006224:Cricetinae; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism; D000072470:Cytochrome P450 Family 4; D003593:Cytoplasm / Q000378:metabolism; D018076:DNA, Complementary / Q000378:metabolism; D004721:Endoplasmic Reticulum / Q000378:metabolism; D005978:Glutathione / Q000378:metabolism; D005982:Glutathione Transferase / Q000096:biosynthesis*; D007975:Leukotriene B4 / Q000378:metabolism; D017997:Leukotriene C4 / Q000378:metabolism; D008562:Membrane Glycoproteins; D008856:Microscopy, Fluorescence; D016253:Microscopy, Immunoelectron; D006899:Mixed Function Oxygenases / Q000378:metabolism; D008954:Models, Biological; D010957:Plasmids / Q000378:metabolism; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D060245:Tetraspanin 29; D014162:Transfection
chemical_list: D015703:Antigens, CD; D018076:DNA, Complementary; D008562:Membrane Glycoproteins; D060245:Tetraspanin 29; D007975:Leukotriene B4; D017997:Leukotriene C4; D003577:Cytochrome P-450 Enzyme System; D006899:Mixed Function Oxygenases; D000072470:Cytochrome P450 Family 4; C040780:leukotriene B4 20-hydroxylase; D005982:Glutathione Transferase; C041116:leukotriene-C4 synthase; D005978:Glutathione