Cells were lysed in lysis buffer (50 mM Tris-HCl, pH 8.0, 150 mM NaCl, 1 mM EDTA, 1% Nonidet P-40) supplemented with EDTA-free protease inhibitor cocktail (Roche Diagnostics) by incubating on ice for 20 min. Aliquots of cleared cell lysates from the same samples were resolved by SDS-PAGE, transferred onto nitrocellulose membranes, and processed in parallel for immunoblotting analyses for multiple proteins. Membranes were immunoblotted with specific antibodies and the bound antibodies were detected with an enhanced chemiluminescence kit (GE Healthcare Life Sciences, Piscataway, NJ, USA).