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The phagocyte NADPH oxidase depends on cholesterol‐enriched membrane microdomains for assembly
Author(s) -
Vilhardt Frederik,
van Deurs Bo
Publication year - 2004
Publication title -
the embo journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 7.484
H-Index - 392
eISSN - 1460-2075
pISSN - 0261-4189
DOI - 10.1038/sj.emboj.7600066
Subject(s) - nadph oxidase , lipid raft , cytosol , superoxide , microbiology and biotechnology , biochemistry , phagocyte , chemistry , lipid microdomain , oxidase test , membrane , biology , enzyme , phagocytosis
The superoxide‐producing phagocyte NADPH oxidase consists of a membrane‐bound flavocytochrome b 558 complex, and cytosolic factors p47phox, p67phox and the small GTPase Rac, which translocate to the membrane to assemble the active complex following cell activation. We here show that insolubility of NADPH oxidase subunits in nonionic detergents TX‐100, Brij‐58, and Brij‐98 is a consequence of inclusion into cholesterol‐enriched membrane microdomains (lipid rafts). Thus, flavocytochrome b 558 , in a cholesterol‐dependent manner, segregated to the bouyant low‐density detergent‐resistant membrane (DRM) fraction, and the cytosolic NADPH oxidase factors associated dynamically with low‐density DRM. Further, superoxide production following cholesterol depletion was severely compromised in intact cells or in a cell‐free reconstituted system, correlating with a reduced translocation of cytosolic phox subunits to the membrane. In analogy with the widely accepted role of lipid rafts as signaling platforms, our data indicate that cholesterol‐enriched microdomains act to recruit and/or organize the cytosolic NADPH oxidase factors in the assembly of the active NADPH oxidase.