Erf3 gspt1

Cells were lysed in M-PER buffer (Thermo Scientific) containing protease/phosphatase inhibitor cocktail (Roche). After assessing protein concentration by BCA assay (Pierce), equal amounts of protein for each sample were loaded into 4–12% Bis-Tris gels (Invitrogen), transferred to nitrocellulose membranes, and immunoblotted with antibodies against Ikaros, Helios, cleaved Caspase-3, and Actin (Cell Signaling); and eRF3/GSPT1 (Abcam). Membranes were detected on an Odyssey detection system (LI-COR Biosciences) after incubation with IRDye®800-labeled goat anti-rabbit IgG and IRDye®680-labeled goat anti-mouse IgG (LI-COR) secondary antibodies.

Cells were washed with PBS before being lysed with Cell Lysis Buffer (Cell Signaling) supplemented with protease and phosphatase inhibitor cocktails (Roche) at 4°C for 15 minutes. The cell lysate vortexed before being centrifuged at 14,000 x g for 20 min at 4°C. Protein in cell lysate was quantified by BCA assay (Pierce). Primary antibodies used in this study include β-Actin (Cell Signaling Technology, 3700s), CK1α (Abcam, ab206652), CRBN (Novus Biologicals, NBP1-91810), eRF3/GSPT1 (Abcam, ab49878), IKZF1 (Cell Signaling Technology, 5443S), IKZF3 (Cell Signaling Technology, 15103S), and vinculin (Abcam, ab130007). Blot quantification was performed using Image Studio 4.0 software, normalizing to loading controls.