pLKO.1 - TRC control was a gift from David Root (Addgene plasmid # 10879)76. A bacterial glycerol stock containing the LV-FURIN-shRNA plasmid was purchased from Sigma (SHCLNG-TRCN0000262167). High-titer lentiviral supernatant was generated by co-transfection of shRNA expression vector together with psPAX2 and pMD2.G to package letivirus particles in HEK-293T cells. psPAX2 (Addgene plasmid # 12260) and pMD2.G (Addgene plasmid # 12259) were gifts from Didier Trono. Lentiviral supernatant was concentrated by centrifugation at 19,300 × g for 2hr at 4°C and resuspended in NPC media. Viral titer was determined using a qPCR lentiviral titration kit (Applied Biological Material Inc. - LV900) and TaqMan® RNA-to-Ct™ 1-Step Kit (ThermoFisher Scientific – 4392938). NPC transduction was performed by addition of lentiviral particals to NPCs at an MOI of 0.5–1 followed by centrifugation of plate at 1,000 × g for 1hr at RT then incubation of NPCs at 37°C for an additional 6 hours. 48hr after infection, transduced cells were selected for with 1µg/mL puromycin for 48hr. FURIN knockdown was validated by qPCR using TaqMan® RNA-to-Ct™ 1-Step Kit (ThermoFisher Scientific – 4392938).
