title: Mobilization of iron from endocytic vesicles. The effects of acidification and reduction.
abstract: The factors necessary to dissociate iron from transferrin in endocytic vesicles and to mobilize the iron across the vesicle membrane were studied in a preparation of endocytic vesicles markedly enriched in transferrin-transferrin receptor complexes isolated from rabbit reticulocytes. Vesicles were prepared with essentially fully saturated transferrin by incubating the reticulocytes with the protonophore carbonyl cyanide 4-(trifluoromethoxy)phenylhydrazone prior to incubation with 59Fe, 125I-transferrin with or without fluorescein isothiocyanate labeling. Initiation of acidification by the addition of ATP was sufficient to achieve dissociation of 59Fe from transferrin with a rate constant of 0.054 +/- 0.06 s-1. Mobilization of 59Fe out of the vesicles required, besides ATP, the addition of a reductant with 1 mM ascorbate, allowing approximately 60% mobilization at 10 min with a rate constant of 0.0038 +/- 0.0006 s-1. An NADH:ferricyanide reductase activity could be demonstrated in the vesicles with an activity of 7.1 x 10(-9) mol of NADH reduced per min/mg of vesicle protein. Both dissociation and mobilization were inhibited by N-ethylmaleimide, carbonyl cyanide 4-(trifluoromethoxy)phenylhydrazone, and monensin. Mobilization, but not dissociation, was inhibited by the permeant Fe(II) chelator alpha,alpha'-dipyridyl. The Fe(III) chelators deferoxamine, diethylenetriaminepentaacetic acid, and apotransferrin did not promote mobilization of dissociated iron in the absence of a reductant. This study establishes the basis for the cellular incorporation of iron through the endocytic pathway in which the endocytic vesicle membrane utilizes, in a sequential way, an acidification system, an iron reduction system, and an Fe(II) transporter system.
authors.affiliation: Departamento de Biologia, Facultad de Ciencias, Universidad de Chile, Santiago.
authors.forename: M T
authors.initials: MT
authors.lastname: Núñez
authors.forename: V
authors.initials: V
authors.lastname: Gaete
authors.forename: J A
authors.initials: JA
authors.lastname: Watkins
authors.forename: J
authors.initials: J
authors.lastname: Glass
mesh_terms: D000818:Animals; D004705:Endocytosis / Q000187:drug effects; D006863:Hydrogen-Ion Concentration; D007501:Iron / Q000097:blood*; D007502:Iron Chelating Agents / Q000494:pharmacology; D007700:Kinetics; D008954:Models, Biological; D015388:Organelles / Q000187:drug effects / Q000378:metabolism*; D010084:Oxidation-Reduction; D011817:Rabbits; D012156:Reticulocytes / Q000378:metabolism*
chemical_list: D007502:Iron Chelating Agents; D007501:Iron