Cell lysates were boiled in sample buffer containing 100 mM DTT for 5 min, resolved by 4 to 12% gradient SDS-PAGE, and transferred to nitrocellulose. Primary antibodies were used at 1:1000 dilution in tris-buffered saline with Tween 20 (0.1% Tween 20) plus 5% bovine serum albumin, with the exception of anti-HSP90, which was used at 1:2000 dilution. Secondary antibodies [goat anti-rabbit immunoglobulin G (IgG) horseradish peroxidase (HRP), goat anti-mouse IgG HRP (both from Bio-Rad), and goat anti-guinea pig IgG HRP (Jackson ImmunoResearch)] were used at 1:5000 dilution. Imaging was performed after incubation with the Clarity Western ECL Substrate (Bio-Rad no. 1705061) according to the manufacturer’s instructions.

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