Distinct Roles of Nrf1 and Nrf2 in Monitoring the Reductive Stress Response to Dithiothreitol (DTT)

Abstract

Transcription factor Nrf2 (nuclear factor, erythroid 2-like 2, encoded by Nfe2l2) has been accepted as a key player in redox regulatory responses to oxidative or reductive stresses. However, relatively little is known about the potential role of Nrf1 (nuclear factor, erythroid 2-like 1, encoded by Nfe2l1) in the redox responses, particularly to reductive stress, although this ‘fossil-like’ factor is indispensable for cell homeostasis and organ integrity during the life process. Herein, we examine distinct roles of Nrf1 and Nrf2 in monitoring the defense response to 1,4–dithiothreitol (DTT, serving as a reductive stressor), concomitantly with unfolded protein response being induced by this chemical (also defined as an endoplasmic reticulum stressor). The results revealed that intracellular reactive oxygen species (ROS) were modestly increased in DTT-treated wild-type (WT) and Nrf1α−/− cell lines, but almost unaltered in Nrf2−/−ΔTA or caNrf2ΔN cell lines (with a genetic loss of transactivation or N-terminal Keap1-binding domains, respectively). This chemical treatment also enabled the rate of oxidized to reduced glutathione (i.e., GSSG to GSH) to be amplified in WT and Nrf2−/−ΔTA cells, but diminished in Nrf1α−/− cells, along with to or less extents changes in caNrf2ΔN cells. Consequently, Nrf1α−/−, but not Nrf2−/−ΔTA or caNrf2ΔN, cell viability was reinforced by DTT against its cytotoxicity, as accompanied by decreased apoptosis. Further experiments unraveled that Nrf1 and Nrf2 differentially, and also synergistically, regulated DTT-inducible expression of critical genes for defending against redox stress and endoplasmic reticulum stress. In addition, we also identified that Cys342 and Cys640 of Nrf1 (as redox-sensing sites within its N-glycodomain and DNA-binding domain, respectively) are required for its protein stability and transcription activity.

Redox-dependent condensation and cytoplasmic granulation by human ssDNA binding protein 1 delineate roles in oxidative stress response

“…Menadione was shown to generate ROS through redox cycling while disrupting mitochondrial membrane potential, triggering cytochrome c redistribution to the cytosol and inducing cell death, thus inducing indirect, internal oxidative stress ( 36 ). DTT was used to induce endoplasmic reticulum stress by inhibiting protein folding through the reduction of disulfide bridges ( 37 ). DTT is also responsible for ROS production by influencing cell signaling and the glutathione system ( 37 ).…”

“…DTT was used to induce endoplasmic reticulum stress by inhibiting protein folding through the reduction of disulfide bridges ( 37 ). DTT is also responsible for ROS production by influencing cell signaling and the glutathione system ( 37 ). Treatment with NaAsO 2 , menadione sodium bisulfite, and DTT, similarly to H 2 O 2 , resulted in the decrease of nuclear hSSB1 signal compared to appropriate serum deprivation controls according to treatment time ( Fig.…”

An Intricate Network Involving the Argonaute ALG-1 Modulates Organismal Resistance to Oxidative Stress

“…Importantly, the increased stress resistance promoted by ALG-1 overexpression was specific to oxidative stress, given that ALG-1/OE worms were less resistant to dithiothreitol (DTT) - a reducing agent known to induce endoplasmic reticulum (ER) stress 42 , 43 (Supplementary Fig. 5e ) – and as resistant as the controls to heat stress (Supplementary Fig.…”

Unfolding the Interactions between Endoplasmic Reticulum Stress and Oxidative Stress

“…This phenomenon can be observed by certain thiol-containing compounds such as dithiothreitol (DTT), which disrupts the process of oxidative protein folding. Accordingly, DTT treatment initiates the UPR; however, increased ROS levels were also observed [66]. Similarly, α-lipoic acid has been reported to induce ROS production and UPR activation in hepatoma cells [67].…”