title: Heavy metal detoxification in crustacean epithelial lysosomes: role of anions in the compartmentalization process.
abstract: Crustacean hepatopancreatic lysosomes are organelles of heavy metal sequestration and detoxification. Previous studies have shown that zinc uptake by lysosomal membrane vesicles (LMV) occurred by a vanadate- and thapsigargin-sensitive ATPase that was stimulated by a transmembrane proton gradient established by a co-localized V-ATPase associated with this organelle. In the present study, hepatopancreatic LMV from the American lobster Homarus americanus were prepared by standard centrifugation methods and 65Zn2+, 36Cl-, 35SO(4)2- and 14C-oxalate2- were used to characterize the interactions between the metal and anions during vesicular detoxification events. Vesicles loaded with SO4(2-) or PO(4)3- led to a threefold greater steady-state accumulation of Zn2+ than similar vesicles loaded with mannitol, Cl- or oxalate2-. The stimulation of 65Zn2+ uptake by intravesicular sulfate was SO(4)2- concentration dependent with a maximal enhancement at 500 micromol l(-1). Zinc uptake in the presence of ATP was proton-gradient enhanced and electrogenic, exhibiting an apparent exchange stoichiometry of 1Zn+/3H+. 35SO4(2-) and 14C-oxalate2- uptakes were both enhanced in vesicles loaded with intravesicular Cl- compared to vesicles containing mannitol, suggesting the presence of anion countertransport. 35SO4(2-) influx was a sigmoidal function of external [SO(4)2-] with 25 mmol l(-1) internal [Cl-], or with several intravesicular pH values (e.g. 7.0, 8.0 and 9.0). In all instances Hill coefficients of approximately 2.0 were obtained, suggesting that 2 sulfate ions exchange with single Cl- or OH- ions. 36Cl- influx was a sigmoidal function of external [Cl-] with intravesicular pH of 7.0 and 9.0. A Hill coefficient of 2.0 was also obtained, suggesting the exchange of 2 Cl- for 1 OH-. 14C-oxalate influx was a hyperbolic function of external [oxalate2-] with 25 mmol l(-1) internal [Cl-], suggesting a 1:1 exchange of oxalate2- for Cl-. As a group, these experiments suggest the presence of an anion exchange mechanism exchanging monovalent for polyvalent anions. Polyvalent inorganic anions (SO4(2-) and PO4(3-)) are known to associate with metals inside vesicles and a detoxification model is presented that suggests how these anions may contribute to concretion formation through precipitation with metals at appropriate vesicular pH.
authors.affiliation: Department of Biology, University of North Florida, 4567 St Johns Bluff Road, S., Jacksonville, FL 32224, USA.
authors.forename: Kenneth M
authors.initials: KM
authors.lastname: Sterling
authors.forename: Prabir K
authors.initials: PK
authors.lastname: Mandal
authors.forename: Barbara A
authors.initials: BA
authors.lastname: Roggenbeck
authors.forename: Sean E
authors.initials: SE
authors.lastname: Ahearn
authors.forename: George A
authors.initials: GA
authors.lastname: Gerencser
authors.forename: Gregory A
authors.initials: GA
authors.lastname: Ahearn
mesh_terms: D000818:Animals; D000838:Anions / Q000378:metabolism*; D001693:Biological Transport, Active; D002712:Chlorides; D004847:Epithelial Cells / Q000166:cytology / Q000378:metabolism*; D043143:Hepatopancreas / Q000166:cytology / Q000378:metabolism; D006878:Hydroxides; D008247:Lysosomes / Q000378:metabolism*; D019216:Metals, Heavy / Q000378:metabolism*; D008121:Nephropidae / Q000378:metabolism*; D013431:Sulfates
chemical_list: D000838:Anions; D002712:Chlorides; D006878:Hydroxides; D019216:Metals, Heavy; D013431:Sulfates; C031356:hydroxide ion