Ceapin a7

The RNeasy Kit, RT² First Strand Kit, RT² SYBR Green ROX qPCR Mastermix, and RT² lncRNA qPCR Assay for Human ZFAS1 were purchased from Qiagen (Valencia, CA, USA). The BrdU Cell Proliferation Assay Kit was purchased from BioVision (Mountain View, CA, USA). The sorafenib was purchased from LC Laboratories (Woburn, MA, USA). The GSK-2606414 was purchased from APExBIO (Boston, MA, USA). The trans-ISRIB was purchased from Tocris Bioscience (Ellisville, MO, USA). Ceapin-A7 and 4μ8c were purchased from Sigma-Aldrich (St. Louis, MO, USA). Silencer Select pre-designed siRNA for human ZFAS1, negative control siRNA, and RNAiMAX transfection reagent were purchased from Thermo Fisher Scientific (Wilmington, DE, USA).

BC3 (ATCC, CRL-2277) and BCBL1 (kindly provided by Prof. P. Monini, National AIDS Center, Istituto Superiore di Sanità, Rome, Italy) are human B-cell lines derived from Primary effusion Lymphoma (PEL). Cells were cultured in RPMI 1640 (Sigma-Aldrich, St. Louis, MO, USA, R0883) with 10% fetal calf serum (Euroclone, Milano, Italy, ECLS0180L), l-glutamine (2 mM) streptomycin (100 µg/mL) and penicillin (100 U/mL) (Gibco, Gaithersburg, MD, USA, 10378-016) in 5% CO₂ at 37 °C. Cells were treated with the following drugs: 4μ8C (IRE1 RNAse inhibitor) provided by Sigma (Sigma-Aldrich, MO, USA, cat n. SML0949), Ceapin-A7 (ATF6a signaling blocker) provided by Sigma-Aldrich (cat n. SML2330), GSK2606414 (PERK inhibitor) provided by Selleckem, USA (cat. n. S7307). Chloroquine (CQ) (inhibitor of autophagic protein degradation) (Sigma-Aldrich, MO, USA, cat. n. C6628). Chemicals were added to cell cultures at the final concentrations of 10 and 20 μM (4μ8C), 6 and 12 μM (Ceapin) and 10 and 20 ng/mL (GSK2606414) for 24 h; Cloroquine was used at final concentration of 10 μM for 18 h. After treatments, cells were collected, counted by trypan blue exclusion assay using a hemocytometer and used for further analysis. Each experiment was performed in triplicate and repeated at least three times.