FIG 1.

Design and validation of the ARE-BSD-PEST reporter. (A) ARE-BSD-PEST consists of a transcriptional pause site (TPS) followed by a synthetic sequence consisting of a triple antioxidant response sequence (4× ARE), a minimal promoter, an open reading frame for a blasticidin resistance gene with a PEST degron sequence (BSD-PEST), and a terminator; the BSD-PEST transcription is under the control of the NRF2 transcription factor. (B) Western blot showing effective KEAP1 knockout. The Western blot shows activation of NRF2 as well as NRF2 transcriptional targets NQO1 and FTL. β-Actin (ACTB) was used as a loading control. (C) HK2-BSD cells harboring KEAP1 knockout (HK2-BSD-sgKEAP1) are more resistant to blasticidin than control cells.