Lys inhibits Nrf2 protein levels in HCT116 and SW480 cells. (a) HCT116 and SW480 cells were treated with varying doses of Lys (ranging from 0 to 30 μM) over a period of 24 hours, and western blot analysis was done to determine the amount of the Nrf2 protein produced. (b) HCT116 and SW480 cells were treated with 30 μM of Lys for 6 hours, 12 hours, and 24 hours, and western blot analysis was done to determine how much Nrf2 protein was produced. qRT-PCR was performed to quantify the mRNA levels of Nrf2 and Keap1 in CRC cells after HCT116 and SW480 cells were treated with 30 μM of Lys for a duration of 6 hours and 12 hours (c, d). (e) HCT116 and SW480 cells were treated with 30 μM of Lys, 10 μM of MG132, or a combination of Lys and MG132 for a period of 12 hours, and then, the protein levels of Nrf2 were analyzed by western blotting. (f) The ubiquitination test was used to determine the levels of ubiquitination of Nrf2 in HCT116 and SW480 cells after they were treated with or without 30 μM of Lys for a period of 12 hours. The data are presented as the mean standard deviation, with a sample size of three.