Heme oxygenase-1 promotes survival of renal cancer cells through modulation of apoptosis- and autophagy-regulating molecules - PubMed (original) (raw)

HO-1, RAPA, and sorafenib modulate the association of Beclin-1 with Rubicon, and HO-1 inhibits RAPA-induced autophagy of renal cancer cells. A, Caki-1 and 786-O cells were transfected with different concentrations of HO-1 overexpression plasmid or empty vector for 24 h. B, Caki-1 cells were transfected with either HO-1 overexpression plasmid (0.5 μg) or empty vector, and treated with 10 ng/ml RAPA, 20 μ

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sorafenib or vehicle alone for 24 h. C, Caki-1 cells were transfected with either 50 n

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HO-1 siRNA or control siRNA. After 48 h of siRNA transfection the cells were treated with either 20 μ

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sorafenib or vehicle for another 24 h. A–C, cell lysates were immunoprecipitated (IP) with anti-Rubicon. Immunoprecipitates were captured by protein A-Sepharose beads, boiled in SDS buffer and separated by SDS-PAGE. The expression of Beclin-1 and Rubicon in the immunoprecipitate, and β-actin in the cell lysate was analyzed by Western blot. Data shown are representative of three independent experiments. D, 786–0 cells were treated with different combinations of 50 ng/ml RAPA, 10 μ

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(left panel) or 20 μ

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(right panel) CoPP, or vehicle for 72 h. The cells were stained with Cyto ID Green Autophagy Detection Reagent as described in “Experimental Procedures,” and analyzed by flow cytometry. Data shown are representative of three independent experiments.