title: Silencing overexpression of FXYD3 protein in breast cancer cells amplifies effects of doxorubicin and γ-radiation on Na(+)/K(+)-ATPase and cell survival.
abstract: FXYD3, also known as mammary tumor protein 8, is overexpressed in several common cancers, including in many breast cancers. We examined if such overexpression might protect Na(+)/K(+)-ATPase and cancer cells against the high levels of oxidative stress characteristic of many tumors and often induced by cancer treatments. We measured FXYD3 expression, Na(+)/K(+)-ATPase activity and glutathionylation of the β1 subunit of Na(+)/K(+)-ATPase, a reversible oxidative modification that inhibits the ATPase, in MCF-7 and MDA-MB-468 cells. Expression of FXYD3 was suppressed by transfection with FXYD3 siRNA. A colorimetric end-point assay was used to estimate cell viability. Apoptosis was estimated by caspase 3/7 (DEVDase) activation using a Caspase fluorogenic substrate kit. Expression of FXYD3 in MCF-7 breast cancer cells was ~eightfold and ~twofold higher than in non-cancer MCF-10A cells and MDA-MB-468 cancer cells, respectively. A ~50 % reduction in FXYD3 expression increased glutathionylation of the β1 Na(+)/K(+)-ATPase subunit and reduced Na(+)/K(+)-ATPase activity by ~50 %, consistent with the role of FXYD3 to facilitate reversal of glutathionylation of the β1 subunit of Na(+)/K(+)-ATPase and glutathionylation-induced inhibition of Na(+)/K(+)-ATPase. Treatment of MCF-7 and MDA-MB- 468 cells with doxorubicin or γ-radiation decreased cell viability and induced apoptosis. The treatments upregulated FXYD3 expression in MCF-7 but not in MDA-MB-468 cells and suppression of FXYD3 in MCF-7 but not in MDA-MB-468 cells amplified effects of treatments on Na(+)/K(+)-ATPase activity and treatment-induced cell death and apoptosis. Overexpression of FXYD3 may be a marker of resistance to cancer treatments and a potentially important therapeutic target.
authors.affiliation: North Shore Heart Research Group, Kolling Institute of Medical Research, University of Sydney, Sydney, Australia.
authors.forename: Chia-Chi
authors.initials: CC
authors.lastname: Liu
authors.affiliation: North Shore Heart Research Group, Kolling Institute of Medical Research, University of Sydney, Sydney, Australia.
authors.forename: Rachel
authors.initials: R
authors.lastname: Teh
authors.affiliation: North Shore Heart Research Group, Kolling Institute of Medical Research, University of Sydney, Sydney, Australia.
authors.forename: Christine A
authors.initials: CA
authors.lastname: Mozar
authors.affiliation: Hormones and Cancer Laboratories, Kolling Institute of Medical Research, University of Sydney, Sydney, Australia.
authors.forename: Robert C
authors.initials: RC
authors.lastname: Baxter
authors.affiliation: North Shore Heart Research Group, Kolling Institute of Medical Research, University of Sydney, Sydney, Australia. helge.rasmussen@sydney.edu.au.|The Department of Cardiology, Royal North Shore Hospital, St Leonards, NSW, 2065, Australia. helge.rasmussen@sydney.edu.au.
authors.forename: Helge H
authors.initials: HH
authors.lastname: Rasmussen
mesh_terms: D017209:Apoptosis / Q000187:drug effects / Q000528:radiation effects; D001943:Breast Neoplasms / Q000188:drug therapy / Q000235:genetics* / Q000532:radiotherapy; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects / Q000235:genetics* / Q000528:radiation effects; D004317:Doxorubicin / Q000494:pharmacology*; D005260:Female; D005720:Gamma Rays / Q000627:therapeutic use*; D015870:Gene Expression / Q000187:drug effects / Q000528:radiation effects; D020868:Gene Silencing / Q000187:drug effects*; D006801:Humans; D061986:MCF-7 Cells; D008565:Membrane Proteins / Q000235:genetics*; D009363:Neoplasm Proteins / Q000235:genetics*; D000254:Sodium-Potassium-Exchanging ATPase / Q000235:genetics*
chemical_list: C091777:FXYD3 protein, human; D008565:Membrane Proteins; D009363:Neoplasm Proteins; D004317:Doxorubicin; D000254:Sodium-Potassium-Exchanging ATPase
keywords: Breast cancer; Doxorubicin; FXYD3 protein; Gamma irradiation; Glutathionylation; Sodium–potassium (Na+/K+) ATPase