Ddit3 chop

Immunoblots were performed using standard protocols. Cells were lysed directly in Laemmli loading buffer, proteins were resolved by SDS-PAGE and transferred to PVDF membranes (EMD Millipore, Billerica, MA). Membranes were blocked in Tris buffered saline (TBS) containing 0.1% Tween and 5% non-fat powdered milk (TBS-T, 5% milk). Primary antibody incubation was performed in TBS-T, 5% milk overnight at 4°C. Proteins were visualized using a species-specific HRP-conjugated secondary antibody and the ECL Plus chemifluorescent detection system (Thermo Fisher Scientific Inc., Waltham, MA) on a STORM scanner ((GE Healthcare, Piscataway, NJ). Signal was quantified using ImageJ software (http://rsb.info.nih.gov/ij/). The following primary antibodies were used: ACC (#3676, Cell Signaling), phospho-ACC (Ser79, #3661, Cell Signaling), AMPKα (#2603, Cell Signaling), phospho-AMPKα (Thr172, #2535, Cell Signaling), DDIT3/CHOP (#2895, Cell Signaling), HER2 (#4290, Cell Signaling), HER3 (#4754, Cell Signaling), α-tubulin (MCA78G, AbD Serotec, Oxford, UK). Secondary antibodies: goat-anti-rabbit-HRP (#7074, Cell Signaling), horse-anti-mouse-HRP (#7076, Cell Signaling), goat-anti-rat-HRP (sc-2303), Santa Cruz Biotechnology, Inc., Santa Cruz, CA).