Si pkr

ON-TARGET plus SMART pool small-interfering RNA (siRNA) for si-Control was bought from Dharmacon Research. si-PKR was bought from Santa Cruz. Transfections of si-PKR and si-Control were performed using INTERFERin siRNA transfection kit (Polyplus Transfection, Huntingdon, UK) according to the protocol.

U87, A172, and HEK293T (293T) cells were purchased from American Type Culture Collection. HeLa cells stably expressing empty vector (EV), 6His-SUMO 1 and 6His-SUMO 2, were obtained from Dr. RT Hay and have been described previously [37 (link)]. Cells were cultured in DMEM supplemented with 10% FBS and 1% penicillin-streptomycin (ThermoFisher Scientific). GBM34 and GBM44 GSCs obtained from Dr. Mariano Viapiano (Brigham and Women’s Hospital, Boston, MA) were maintained as neurospheres as described [27 ]. All cell lines were screened for mycoplasma using the ATCC Universal Mycoplasma Detection Kit (catalogue # 30- 121 1012K) every 4 months. Early passage primary MEFs from wild-type, MyD88^−/−, Rig-I^−/−, Mda5^−/−, Lgp2^−/−, MAVS^−/−, Tmem173^−/−, and Trif^−/− mice were cultured as previously described [51 (link), 52 (link)]. All cell lines were authenticated by routine morphological and growth analysis and by western blotting. TMZ was obtained from Sigma-Aldrich. DNA transfection was performed using TransIT LT1 (Mirus) and siRNA transfection with Oligofectamine (Invitrogen). Cycloheximide (CHX) and MG132 were from Cayman Chemical. The following siRNAs were used: siGENOME non-targeting siRNA#3 (Dharmacon), siGENOME Smartpool si-LGP2 (Dharmacon), si-TLR3 (sc-36685, Santa Cruz), and si-PKR (sc-36263, Santa Cruz).