Nuclear factor erythroid 2 related factor-1 is stimulated by the nutraceutical celastrol at proteasome inhibiting doses

The mechanism by which many nutraceuticals promote health is incompletely understood. Some act by inhibiting the proteasome. Nuclear factor erythroid 2 related factor 1 (NRF1) is a stress-protective transcription factor that is stimulated by suppressed proteasome activity, such as upon exposure to the proteasome inhibitor bortezomib. Here, we investigated whether nutraceutical compounds with proteasome inhibitory properties can stimulate NRF1 to regulate stress-protective genes. Using a luciferase-based promoter reporting the activity of NRF1 and its homolog NRF2 in Hepa 1-6 cells, we performed a small nutraceutical screen and identified that 2 and 4 μM treatment with celastrol exhibited robust stimulatory effect. Using the same reporter in wild type and gene deleted murine embryonic fibroblasts, we found that Nrf1 deficiency reduced celastrol-stimulated luciferase activity. In Hep3B cells, we found 2 μM celastrol, but not lower concentrations, increased NRF1 processing and nuclear accumulation while also suppressing the trypsin-like, chymotrypsin-like, and caspase-like activities of the proteasome, similar to proteasome inhibiting drugs. However, using control and NRF1 deficient Hep3B cells treated with 2 μM celastrol, MG132, bortezomib, or vehicle, we found distinct effects by celastrol versus MG132 and bortezomib on NRF1-dependent regulation of stress-protective genes. Hence, our results show that celastrol can stimulate NRF1 at concentrations that suppress proteasome activity, but its effect on gene regulation appear distinct compared to proteasome inhibitor. Overall, our findings reveal that that NRF1 may play a role in the health promoting effects of celastrol and possibly other nutraceuticals with proteasome inhibitory properties.