c-abl control sirnas  (Thermo Fisher)

Journal: The Journal of Biological Chemistry

Article Title: AMP-dependent Kinase Inhibits Oxidative Stress-induced Caveolin-1 Phosphorylation and Endocytosis by Suppressing the Dissociation between c-Abl and Prdx1 Proteins in Endothelial Cells *

doi: 10.1074/jbc.M113.460832

Figure Lengend Snippet: Both AMPKα1 and -α2 isoforms are required for the inhibition of caveolin-1 phosphorylation under oxidative stress. A and D, the amounts of p-caveolin-1 and p-c-Abl in HUVEC were examined by Western blotting. Cells were transfected with siRNA against AMPKα1 (A) or α2 (D). Three days after transfection cells were stimulated with 2 mm AICAR for 2 h followed by H2O2 (2 mm) stimulation for 30 min. B, densitometry of p-caveolin-1 in A is shown. C, densitometry of p-c-Abl in A is shown. E, densitometry of p-caveolin-1 in D is shown. F, densitometry of p-c-Abl in D is shown. A and B, representative blots are shown. *, p < 0.01.

Article Snippet: Imatinib mesylate, c-Abl inhibitor, was purchased from Cayman Chemicals (Ann Arbor, MI). siRNAs targeting c-Abl, AMPKα1, AMPKα2, and Prdx1 and control siRNA were purchased from Thermoscientific (Rockford, IL).

Techniques: Inhibition, Western Blot, Transfection

Journal: The Journal of Biological Chemistry

Article Title: AMP-dependent Kinase Inhibits Oxidative Stress-induced Caveolin-1 Phosphorylation and Endocytosis by Suppressing the Dissociation between c-Abl and Prdx1 Proteins in Endothelial Cells *

doi: 10.1074/jbc.M113.460832

Figure Lengend Snippet: AICAR inhibits caveolin-1 phosphorylation under oxidative stress by suppressing the dissociation between Prdx1 and c-Abl. A, cells were transfected with siRNA against Prdx1. Three days after transfection cells were stimulated with 2 mm AICAR for 2 h followed by H2O2 (2 mm) stimulation for 30 min. The amounts of p-c-Abl, p-caveolin-1 were examined by Western blotting. B, densitometry of p-caveolin-1 in A is shown. C, densitometry of p-c-Abl in A is shown. D, cells were stimulated with 2 mm AICAR for 2 h followed by H2O2 (2 mm) stimulation for 30 min. After total cell lysates of each group were collected, the interaction between Prdx1 and c-Abl was examined by immunoprecipitation with anti-Prdx1 antibody. Immunoprecipitates were then subjected to immunoblotting using anti-c-Abl antibody. E, densitometry of p-c-Abl in D is shown. F, cells were transfected with siRNA against AMPKα1 or -α2. Three days after transfection cells were stimulated with 2 mm AICAR for 2 h followed by H2O2 (2 mm) stimulation for 30 min. After total cell lysates of each group were collected, the interaction between c-Abl and Prdx1 was examined by immunoprecipitation (IP) with anti-Prdx1 antibody. Immunoprecipitates were then subjected to immunoblotting using anti-c-Abl antibody. G, densitometry of c-Abl in F. H, cells were transfected with siRNA against AMPKα1 or -α2. Three days after transfection cells were stimulated with 2 mm AICAR for 2 h followed by H2O2 (2 mm) stimulation for 30 min. After total cell lysates of each group were collected, the interaction between total AMPK and Prdx1 or c-Abl was examined by immunoprecipitation with anti-total AMPK antibody. Immunoprecipitates were then subjected to immunoblotting using anti-c-Abl and Prdx1 antibody. A, D, and F, representative blots are shown. *, p < 0.01; NS, not significant.

Article Snippet: Imatinib mesylate, c-Abl inhibitor, was purchased from Cayman Chemicals (Ann Arbor, MI). siRNAs targeting c-Abl, AMPKα1, AMPKα2, and Prdx1 and control siRNA were purchased from Thermoscientific (Rockford, IL).

Techniques: Transfection, Western Blot, Immunoprecipitation