Abstract
Mechanisms underlying therapy resistance of tumor cells include protein kinase Akt. Putative Akt targets include store-operated Ca(2+)-entry (SOCE) accomplished by pore forming ion channel unit Orai1 and its regulator STIM1. We explored whether therapy resistant (A2780cis) differ from therapy sensitive (A2780) ovary carcinoma cells in Akt, Orai1, and STIM1 expression, Ca(2+)-signaling and cell survival following cisplatin (100 µM) treatment. Transcript levels were quantified with RT-PCR, protein abundance with Western blotting, cytosolic Ca(2+)-activity ([Ca(2+)]i) with Fura-2-fluorescence, SOCE from increase of [Ca(2+)]i following Ca(2+)-readdition after Ca(2+)-store depletion, and apoptosis utilizing flow cytometry. Transcript levels of Orai1 and STIM1, protein expression of Orai1, STIM1, and phosphorylated Akt, as well as SOCE were significantly higher in A2780cis than A2780 cells. SOCE was decreased by Akt inhibitor III (SH-6, 10 µM) in A2780cis but not A2780 cells and decreased in both cell lines by Orai1 inhibitor 2-aminoethoxydiphenyl borate (2-ABP, 50 µM). Phosphatidylserine exposure and late apoptosis following cisplatin treatment were significantly lower in A2780cis than A2780 cells, a difference virtually abolished by SH-6 or 2-ABP. In conclusion, Orai1/STIM1 expression and function are increased in therapy resistant ovary carcinoma cells, a property at least in part due to enhanced Akt activity and contributing to therapy resistance in those cells.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Antineoplastic Agents / pharmacology
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Apoptosis / drug effects
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Apoptosis / genetics
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Benzamides / pharmacology
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Blotting, Western
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Boron Compounds / pharmacology
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Calcium / metabolism*
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Calcium Channels / genetics
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Calcium Channels / metabolism*
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Cell Line, Tumor
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Cell Survival / drug effects
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Cell Survival / genetics
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Cisplatin / pharmacology
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Drug Resistance, Neoplasm / genetics
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Female
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Gene Expression Regulation, Neoplastic / drug effects
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Humans
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Hydrazines / pharmacology
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Immediate-Early Proteins / antagonists & inhibitors
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Immediate-Early Proteins / metabolism
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Membrane Proteins / genetics
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Membrane Proteins / metabolism*
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Neoplasm Proteins / genetics
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Neoplasm Proteins / metabolism*
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ORAI1 Protein
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Ovarian Neoplasms / genetics
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Ovarian Neoplasms / metabolism
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Ovarian Neoplasms / pathology
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Phosphatidylinositols / pharmacology
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Phosphorylation / drug effects
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Protein Serine-Threonine Kinases / antagonists & inhibitors
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Protein Serine-Threonine Kinases / metabolism
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Proto-Oncogene Proteins c-akt / antagonists & inhibitors
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Proto-Oncogene Proteins c-akt / metabolism
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Reverse Transcriptase Polymerase Chain Reaction
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Stromal Interaction Molecule 1
Substances
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Antineoplastic Agents
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Benzamides
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Boron Compounds
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Calcium Channels
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EMD 638683
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Hydrazines
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Immediate-Early Proteins
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Membrane Proteins
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Neoplasm Proteins
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ORAI1 Protein
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ORAI1 protein, human
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Phosphatidylinositols
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SH-6 compound
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STIM1 protein, human
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Stromal Interaction Molecule 1
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2-aminoethoxydiphenyl borate
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Protein Serine-Threonine Kinases
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Proto-Oncogene Proteins c-akt
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serum-glucocorticoid regulated kinase
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Cisplatin
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Calcium