Long Isoforms of NRF1 Contribute to Arsenic-Induced Antioxidant Response in Human Keratinocytes

Background: Human exposure to inorganic arsenic (iAs), a potent oxidative Stressor, causes various dermal disorders, including hyperkeratosis and skin cancer. Nuclear factor—erythroid 2—related factor 1 (NRF1, also called NFE2L1) plays a critical role in regulating the expression of many antioxidant response element (ARE)-dependent genes. Objectives: We investigated the role of NRF1 in arsenic-induced antioxidant response and cytotoxicity in human keratinocytes. Results: In cultured human keratinocyte HaCaT cells, inorganic arsenite (iAs³⁺) enhanced the protein accumulation of long isoforms (120—140 kDa) of NRF1 in a dose-and time-dependent fashion. These isoforms accumulated mainly in the nuclei of HaCaT cells. Selective deficiency of NRF1 by lentiviral short-hairpin RNAs in HaCaT cells [NRT1-knockdown (KD)] led to decreased expression of γ-glutamate cysteine ligase catalytic subunit (GCLC) and regulatory subunit (GCLM) and a reduced level of intracellular glutathione. In response to acute iAs³⁺ exposure, induction of some ARE-dependent genes, including NAD(PH:quinone oxidoreductase 1 (NQO1), GCLC and GCLM, was significantly attenuated in NRF1-KD cells. However, the iAs³⁺ -induced expression of heme oxygenase 1 (HMOX-1) was unaltered by silencing NRF1, suggesting that HMOX-1 is not regulated by NRF1. In addition, the lack of NRF1 in HaCaT cells did not disturb iAs³⁺ -induced NRF2 accumulation but noticeably decreased Kelch-like ECH -associated protein 1 (KEAP1) levels under basal and iAs³⁺-exposed conditions, suggesting a potential interaction between NRF1 and KEAP1. Consistent with the critical role of NRF1 in the transcriptional regulation of some ARE-bearing genes, knockdown of NRF1 significantly increased iAs³⁺-induced cytotoxicity and apoptosis. Conclusions: Here, we demonstrate for the first time that long isoforms of NRF1 contribute to arsenic-induced antioxidant response in human keratinocytes and protect the cells from acute arsenic cytotoxicity.