Click here to close
Hello! We notice that you are using Internet Explorer, which is not supported by Xenbase and may cause the site to display incorrectly.
We suggest using a current version of Chrome,
FireFox, or Safari.
Oncotarget
2016 Jun 07;723:35327-40. doi: 10.18632/oncotarget.9205.
Show Gene links
Show Anatomy links
Zinc- and bicarbonate-dependent ZIP8 transporter mediates selenite uptake.
McDermott JR
,
Geng X
,
Jiang L
,
Gálvez-Peralta M
,
Chen F
,
Nebert DW
,
Liu Z
.
???displayArticle.abstract???
Selenite (HSeO3-) is a monovalent anion of the essential trace element and micronutrient selenium (Se). In therapeutic concentrations, HSeO3- has been studied for treating certain cancers, serious inflammatory disorders, and septic shock. Little is known, however, about HSeO3- uptake into mammalian cells; until now, no mammalian HSeO3- uptake transporter has been identified. The ubiquitous mammalian ZIP8 divalent cation transporter (encoded by the SLC39A8 gene) is bicarbonate-dependent, moving endogenous substrates (Zn2+, Mn2+, Fe2+ or Co2+) and nonessential metals such as Cd2+ into the cell. Herein we studied HSeO3- uptake in: human and mouse cell cultures, shRNA-knockdown experiments, Xenopus oocytes, wild-type mice and two transgenic mouse lines having genetically altered ZIP8 expression, and mouse erythrocytes ex vivo. In mammalian cell culture, excess Zn2+ levels and/or ZIP8 over-expression can be associated with diminished viability in selenite-treated cells. Intraperitoneal HSeO3- causes the largest ZIP8-dependent increases in intracellular Se content in liver, followed by kidney, heart, lung and spleen. In every model system studied, HSeO3- uptake is tightly associated with ZIP8 protein levels and sufficient Zn2+ and HCO3- concentrations, suggesting that the ZIP8-mediated electroneutral complex transported contains three ions: Zn2+/(HCO3-)(HSeO3-). Transporters having three different ions in their transport complex are not without precedent. Although there might be other HSeO3- influx transporters as yet undiscovered, data herein suggest that mammalian ZIP8 plays a major role in HSeO3- uptake.
Figure 1. Intracellular Se content and toxicity; ZIP8 protein levels in human prostate cancer cellsA. effect of Zn2+ onSe content in selenite-treated DU145 cells; Zn2+ added concomitantly with HSeO3− for 30 min. B. effect of Zn2+ on selenite-mediated viability of DU145 cells following 12-h treatment, varying HSeO3− amounts. Untreated cultures were given a ratio value of 1.0., and other ratios of experimental regimens are expressed, relative to 1.0. C. effect of EDTA and Zn2+ onSe content in selenite-treated cells; EDTA (5 mM) + Zn2+ (200 μM) pretreatment was 5 min; at far right, Zn2+ was washed out prior to HSeO3− treatment for 30 min. D. Western blots of human ZIP8 in five prostate cancer lines (left lanes) and five prostate non-cancer cell lines (right lanes). GAPDH, glyceraldehyde 3-phosphate dehydrogenase, lane-loading control. E. cell viability of Zn2+-treated ZIP8-MEFs versus LUC-MEFs, as a function of selenite concentration; treatment (5 μM Zn2+ and 25 mM HCO3−) was 16 h. For panels B and E, “relative viability” of cells not treated with selenite is given a value of 1.0, and viability of all other experimental regimens are expressed relative to that control.
Figure 2. Se uptake as a function of time and Zn2+ concentration in oocytes and mouse cell cultures; localization of ZIP8 to plasma membraneA. Se content following addition of Zn2+ (500 μM), or Zn2+ + HCO3− (3.5 mM) in selenite-treated Xenopus laevis oocytes expressing mouse ZIP8 cDNA (closed bar) versus control oocytes carrying vector only (open bar). Without added HCO3−, note that a basal level of HCO3− exists in the ND96 medium due to pCO2 in solution, as can be calculated by the Henderson-Hasselbach equation [30]. B. Se uptake kinetics in stably-transfected high-ZIP8-expressing ZIP8-MEFs vs control low-ZIP8-expressing LUC-MEFs during 20-min exposure to 50 μM HSeO3− + 100 μM Zn2+. C. Intracellular Se content in ZIP8-MEFs as a function of HSeO3− concentration; cells were treated with Zn2+ (100 μM) plus 5 to 100 μM HSeO3−. D. Intracellular Se content in ZIP8-MEFs (treated with 50 μM HSeO3− for 20 min) as a function of Zn2+ concentration. E. Intracellular Se content as a function of HSeO3− concentration (20-min treatment) in wild-type (WT) vs Slc39a8(neo/neo) knockdown mouse primary embryonic fibroblast (MEF) cultures; Zn2+ (100 μM) and HCO3− (25 mM) were constant. F. confocal microscopy of co-localization of ZIP8 (green) with the membrane marker Na+/K+-ATPase subunit (red) in ZIP8-MEF versus LUC-MEF cultures. DAPI, blue stain for DNA, i.e. nucleus. For all mouse culture experiments, HCO3− was present in culture medium at 25 mM. Addition of the HCO3− concentrations used in these experiments in either frog oocytes or mouse cultures did not significantly alter pH (7.5) of the medium. Brackets denote S.D. *P <0.05, **P <0.01.
Figure 3. ZIP8 knockdown by shRNA causes decreased ZIP8 protein expression and Se and Zn uptakeA. Western blot of human ZIP8 in TAs cells stably expressing shRNA against ZIP8 mRNA (shRNA-1 and shRNA-2) vs scrambled shRNA (sc-shRNA) control; GAPDH, glyceraldehyde 3-phosphate dehydrogenase, lane-loading control. B. Se content and C. Zn content in TAs cells, ZIP8-knockdown vs control, after exposure to HSeO3− + Zn2+ (both 200 μM) for 30 min. TAs_ZIP8-shRNA-1 and TAs_ZIP8-shRNA-2 represent two different stably-expressing cell lines carrying two different shRNAs. For all experiments, HCO3− (25 mM) was present in the culture medium. Intracellular Se and Zn content was determined by ICP-MS. Brackets denote S.E.M. *P <0.05, **P <0.01.
Figure 4. Se content in ten mouse tissues as a function of ZIP8 concentrations in three mouse lines having different Slc39a8 genotypesA. Intracellular Se content, 2 h after IP selenite (2.5 mg/kg) administration to the ZIP8-over-expressing BTZIP8-3 mouse line (solid red bar; N=6), the ZIP8-normal-expressing wild-type (WT; pink bar; N=9), and the hypomorph ZIP8 Slc39a8(+/neo) line; (open bar; N=9). Red brackets (S.E.M.) depict statistical differences between BTZIP8-3 and Slc39a8(+/neo); black brackets (S.E.M.) signify statistical differences between BTZIP8-3 and WT. *P <0.05, **P <0.01. Animals (2-3 months of age) included similar numbers of males and females; we first had determined that no sex differences existed. B–F. ZIP8 immunofluorescence in 10-μm tissue sections from WT vs BTZIP8-3 mice. White bar (in panel B) = 50 μm for panels B through E; white bar (in panel F) = 10 μm.
Figure 5. Se content in mouse whole blood or washed red blood cells (RBCs)A. relative Se content in WT whole blood, following HSeO3− treatment (20 μM; 15 min at 37°C), with, versus without, Zn2+ (75 μM); where indicated, whole blood was pretreated for 5 min with EDTA (4 mM) or TPEN (100 μM). Value for selenite-treated whole blood without added Zn2+ was chosen as “1.0” and all other values are relative to that. B. Intracellular Se content in isolated WT RBCs resuspended in PBS containing HCO3− (10 mM; 15 min at 37°C), as a function of Zn2+ concentration following 5-min pretreatment with HSeO3− (20 μM, 37°C). C. Intracellular Se content in isolated RBCs from the three mouse genotypes resuspended in PBS with HCO3− (25 mM) + 75 μM Zn2+ (closed red bars) versus 25 mM HCO3− alone (open bars). Cells were incubated with HSeO3− (20 μM) for 5 min at 37°C. **P <0.01, comparing BTZIP8-3 with, vs without, added Zn2+. D. comparison of rate of Se uptake in RBCs (also resuspended in PBS), as a function of incubation time (37°C); RBCs were isolated from BTZIP8-3, WT and Slc39a8(+/neo) mice; RBCs were treated with HSeO3− (20 μM). **P <0.01, comparing BTZIP8-3 with both WT and Slc39a8(+/neo). Brackets denote S.E.M.
Figure 6. Intracellular Se content of RBCs (resuspended in PBS, without any exogenous Zn2+ or HCO3−), in response to thiol, HSeO3−, and/or EDTARBCs were incubated for 2 h with HSeO3− or its reduced products (20 or 100 μM): first two bars denote first being reduced for 15 min by GSH (100 μM) alone, causing build-up of extracellular reduced selenium product(s) (derived from 20 μM HSeO3−, or with GSH plus EDTA (5 mM); second two bars indicate no reduction by GSH first, but higher concentrations of HSeO3− treatment (100 μM), comparing GSH (800 μM) with or without 5-min EDTA pretreatment (5 mM); last two bars designate HSeO3− treatment (20 μM) alone or with 5-min EDTA (5 mM) pretreatment, but without GSH. *P < 0.05, **P < 0.01.
Figure 7. The two proposed models of selenite transportA. ZIP8-mediated HSeO3− uptake model, which is both zinc- and bicarbonate-dependent. Right, the extracellular HSeO3− conversion model. In the former, ZIP8-mediated HSeO3− uptake can be prevented with prior chelation of Zn2+ by EDTA. Intracellular HSeO3− reacts spontaneously with proteins containing coordinated thiol groups−−modifying signaling pathways dependent on zinc-finger proteins, or thiol proteins. Functional downstream effects on multiple targets are dependent on HSeO3− concentration and ZIP8 levels. HSeO3− reactions with zinc-finger proteins might release free Zn2+, which, in addition to Zn2+ transported by ZIP8, is proposed to elevate labile Zn2+ concentrations and participate in subsequent zinc-signaling functions. Concentration-dependent effects (bottom) range from anti-cancer and anti-inflammatory, to apoptosis and oxidative stress, to cytotoxicity. B. In the alternative model, HSeO3− uptake is proposed to be regulated by extracellular thiol reduction of selenite–in the extracellular milieu–followed by uptake of unknown reduced Se product(s) [22, 35].
Bernotiene,
The effects of cadmium chloride and sodium selenite on protein synthesis in mouse liver.
2013, Pubmed
Bernotiene,
The effects of cadmium chloride and sodium selenite on protein synthesis in mouse liver.
2013,
Pubmed
Bhattacharyya,
Selenite treatment inhibits LAPC-4 tumor growth and prostate-specific antigen secretion in a xenograft model of human prostate cancer.
2008,
Pubmed
Boehler,
Deletion of thioredoxin reductase and effects of selenite and selenate toxicity in Caenorhabditis elegans.
2013,
Pubmed
Boycott,
Autosomal-Recessive Intellectual Disability with Cerebellar Atrophy Syndrome Caused by Mutation of the Manganese and Zinc Transporter Gene SLC39A8.
2015,
Pubmed
Carrera,
Association study of nonsynonymous single nucleotide polymorphisms in schizophrenia.
2012,
Pubmed
Cavalieri,
Selenite (75Se) as a tumor-localizing agent in man.
1966,
Pubmed
Cavalieri,
Sodium selenite Se 75. A more specific agent for scanning tumors.
1968,
Pubmed
Chen,
Protective role of sodium selenite on histopathological lesions, decreased T-cell subsets and increased apoptosis of thymus in broilers intoxicated with aflatoxin B₁.
2013,
Pubmed
Cui,
Interaction of glutathione and sodium selenite in vitro investigated by electrospray ionization tandem mass spectrometry.
2008,
Pubmed
Gasparian,
Selenium compounds inhibit I kappa B kinase (IKK) and nuclear factor-kappa B (NF-kappa B) in prostate cancer cells.
2002,
Pubmed
Gazi,
Sodium selenite inhibits interleukin-6-mediated androgen receptor activation in prostate cancer cells via upregulation of c-Jun.
2007,
Pubmed
Gálvez-Peralta,
ZIP8 zinc transporter: indispensable role for both multiple-organ organogenesis and hematopoiesis in utero.
2012,
Pubmed
He,
ZIP8, member of the solute-carrier-39 (SLC39) metal-transporter family: characterization of transporter properties.
2006,
Pubmed
He,
Discovery of ZIP transporters that participate in cadmium damage to testis and kidney.
2009,
Pubmed
Huang,
Assignment of the human GABA transporter gene (GABATHG) locus to chromosome 3p24-p25.
1995,
Pubmed
Husbeck,
Inhibition of androgen receptor signaling by selenite and methylseleninic acid in prostate cancer cells: two distinct mechanisms of action.
2006,
Pubmed
Jiang,
ATF4 activation by the p38MAPK-eIF4E axis mediates apoptosis and autophagy induced by selenite in Jurkat cells.
2013,
Pubmed
Jiang,
Distinct effects of methylseleninic acid versus selenite on apoptosis, cell cycle, and protein kinase pathways in DU145 human prostate cancer cells.
2002,
Pubmed
Jiang,
Heat shock protein 90-mediated inactivation of nuclear factor-κB switches autophagy to apoptosis through becn1 transcriptional inhibition in selenite-induced NB4 cells.
2011,
Pubmed
Jiang,
Sodium selenite-induced activation of DAPK promotes autophagy in human leukemia HL60 cells.
2012,
Pubmed
Knoell,
Impact of zinc metabolism on innate immune function in the setting of sepsis.
2010,
Pubmed
Koolen,
Genomic microarrays in mental retardation: a practical workflow for diagnostic applications.
2009,
Pubmed
Li,
Recent Positive Selection Drives the Expansion of a Schizophrenia Risk Nonsynonymous Variant at SLC39A8 in Europeans.
2016,
Pubmed
Li,
Downregulation of protein kinase Cα was involved in selenite-induced apoptosis of NB4 cells.
2010,
Pubmed
Liu,
Cd2+ versus Zn2+ uptake by the ZIP8 HCO3--dependent symporter: kinetics, electrogenicity and trafficking.
2008,
Pubmed
,
Xenbase
Liu,
ZIP8 regulates host defense through zinc-mediated inhibition of NF-κB.
2013,
Pubmed
Lopez,
Structurally based, selective interaction of arsenite with steroid receptors.
1990,
Pubmed
Majno,
Apoptosis, oncosis, and necrosis. An overview of cell death.
1995,
Pubmed
Manoharan,
The role of charged residues in the transmembrane helices of monocarboxylate transporter 1 and its ancillary protein basigin in determining plasma membrane expression and catalytic activity.
2006,
Pubmed
,
Xenbase
Maret,
Thiolate ligands in metallothionein confer redox activity on zinc clusters.
1998,
Pubmed
Markadieu,
Physiology and pathophysiology of SLC12A1/2 transporters.
2014,
Pubmed
McDermott,
Pentavalent methylated arsenicals are substrates of human AQP9.
2010,
Pubmed
,
Xenbase
Olm,
Extracellular thiol-assisted selenium uptake dependent on the x(c)- cystine transporter explains the cancer-specific cytotoxicity of selenite.
2009,
Pubmed
Park,
SLC39A8 Deficiency: A Disorder of Manganese Transport and Glycosylation.
2015,
Pubmed
Ren,
Autophagy inhibition through PI3K/Akt increases apoptosis by sodium selenite in NB4 cells.
2009,
Pubmed
Ryan-Harshman,
The relevance of selenium to immunity, cancer, and infectious/inflammatory diseases.
2005,
Pubmed
Sarveswaran,
Selenite triggers rapid transcriptional activation of p53, and p53-mediated apoptosis in prostate cancer cells: Implication for the treatment of early-stage prostate cancer.
2010,
Pubmed
Schroeder,
Effect of selenite combined with chemotherapeutic agents on the proliferation of human carcinoma cell lines.
2004,
Pubmed
Simons,
Arsenite and cadmium(II) as probes of glucocorticoid receptor structure and function.
1990,
Pubmed
Soleimani,
Ionic mechanism of Na+-HCO3- cotransport in rabbit renal basolateral membrane vesicles.
1989,
Pubmed
Sunde,
Incorporation of selenium from selenite and selenocystine into glutathione peroxidase in the isolated perfused rat liver.
1980,
Pubmed
Tarze,
Extracellular production of hydrogen selenide accounts for thiol-assisted toxicity of selenite against Saccharomyces cerevisiae.
2007,
Pubmed
Tian,
Sodium selenite radiosensitizes hormone-refractory prostate cancer xenograft tumors but not intestinal crypt cells in vivo.
2010,
Pubmed
Wang,
ZIP8 is an iron and zinc transporter whose cell-surface expression is up-regulated by cellular iron loading.
2012,
Pubmed
,
Xenbase
Wang,
Generation of a Slc39a8 hypomorph mouse: markedly decreased ZIP8 Zn²⁺/(HCO₃⁻)₂ transporter expression.
2011,
Pubmed
Wang,
Enhanced cadmium-induced testicular necrosis and renal proximal tubule damage caused by gene-dose increase in a Slc39a8-transgenic mouse line.
2007,
Pubmed
Wang,
Dose-dependent effects of selenite (Se(4+)) on arsenite (As(3+))-induced apoptosis and differentiation in acute promyelocytic leukemia cells.
2015,
Pubmed
Waterworth,
Genetic variants influencing circulating lipid levels and risk of coronary artery disease.
2010,
Pubmed
Zhu,
Combined microarray analysis uncovers self-renewal related signaling in mouse embryonic stem cells.
2007,
Pubmed