The integrated stress response (ISR) is a key cellular signaling pathway activated by environmental alterations that represses protein synthesis to restore homeostasis. To prevent sustained damage, the ISR is counteracted by the upregulation of growth arrest and DNA damage-inducible 34 (GADD34), a stress-induced regulatory subunit of protein phosphatase 1 that mediates translation reactivation and stress recovery. Here, we uncover a novel ISR regulatory mechanism that post-transcriptionally controls the stability of PPP1R15A mRNA encoding GADD34. We establish that the 3\' untranslated region of PPP1R15A mRNA contains an active AU-rich element (ARE) recognized by proteins of the ZFP36 family, promoting its rapid decay under normal conditions and stabilization for efficient expression of GADD34 in response to stress. We identify the tight temporal control of PPP1R15A mRNA turnover as a component of the transient ISR memory, which sets the threshold for cellular responsiveness and mediates adaptation to repeated stress conditions.

Heat-shock protein beta1 (HSPB1) is a member of the small HSP family, downregulated in osteoarthritis (OA) chondrocytes and demonstrated the capacity to serve as an RNA-binding protein (RBP). This work aimed to explore the profile of HSPB1 bound RNA and reveal the potential regulation mechanism of HSPB1 in OA. In this work, we captured an unbiased HSPB1-RNA interaction map in Hela cells using the iRIP-seq. The results demonstrated that HSPB1 interacted with plentiful of mRNAs and genomic location toward the CDS region. Functional enrichment of HSPB1-related peaks showed the involvement in gene expression, translation initiation, cellular protein metabolic process, and nonsense-mediated decay. HOMER software analysis showed that HSPB1 bound peaks were over-represented in GAGGAG sequences. In addition, ABLIRC and CIMS algorithm indicated that HSPB1 bound to AU-rich motifs and the proportion of AU-rich peaks in 3\' UTR were slightly higher than that in other regions. Moreover, HSPB1-binding targets analysis revealed several gens were associated with OA including EGFR, PLEC, COL5A1, and ROR2. The association of OA-related mRNAs to HSPB1 was additionally confirmed in OA tissues by the quantitative RIP-PCR experiments. Further experiment demonstrated the downregulation of HSPB1 in OA tissues. In conclusion, our current study confirmed HSPB1 as an RNA-binding protein and revealed its potential function in the pathological process of OA, providing a reliable insight to further investigate the molecular regulation mechanism of HSPB1 in OA.

Herein, (-)-galiellalactone 1 congeners responsible for the nuclear factor erythroid 2-related factor 2 (Nrf2)-activating neuroprotective effects were elucidated. Additionally, novel congener-based Nrf2 activators were identified using a drug repositioning strategy. (-)-Galiellalactone, which comprises a tricyclic lactone skeleton, significantly activates antioxidant response element (ARE)-mediated transcription in neuroblastoma SH-SY5Y cells. Interestingly, two cyclohexene-truncated [3.3] bicyclic lactone analogs, which possess an exocyclic α-methylene-γ-butyrolactone moiety, exhibited higher Nrf2/ARE transcriptional activities than the parent (-)-galiellalactone. We confirmed that the cyclohexene moiety embedding the [3.3] bicyclic lactone congener does not play the essential role of (-)-galiellalactone for Nrf2/ARE activation. Nrf2/ARE activation by novel analogs resulted in the upregulation of downstream antioxidative and phase II detoxifying enzymes, heme oxygenase-1, and NAD(P)H quinone oxidoreductase 1, which are closely related to the cytoprotective effects on neurodegenerative diseases. (-)-Galiellalactone and its [3.3] bicyclic variants 3l and 3p increased the expression of antioxidant genes and exhibited neuroprotective effects against 6-hydroxydopamine-mediated neurotoxicity in the neuroblastoma SH-SY5Y cell line.

Sedanolide is a bioactive compound with anti-inflammatory and antitumor activities. Although it has been recently suggested that sedanolide activates the nuclear factor E2-related factor 2 (NRF2) pathway, there is little research on its effects on cellular resistance to oxidative stress. The objective of the present study was to investigate the function of sedanolide in suppressing hydrogen peroxide (H2O2)-induced oxidative damage and the underlying molecular mechanisms in human hepatoblastoma cell line HepG2 cells. We found that sedanolide activated the antioxidant response element (ARE)-dependent transcription mediated by the nuclear translocation of NRF2. Pathway enrichment analysis of RNA sequencing data revealed that sedanolide upregulated the transcription of antioxidant enzymes involved in the NRF2 pathway and glutathione metabolism. Then, we further investigated whether sedanolide exerts cytoprotective effects against H2O2-induced cell death. We showed that sedanolide significantly attenuated cytosolic and mitochondrial reactive oxygen species (ROS) generation induced by exposure to H2O2. Furthermore, we demonstrated that pretreatment with sedanolide conferred a significant cytoprotective effect against H2O2-induced cell death probably due to preventing the decrease in the mitochondrial membrane potential and the increase in caspase-3/7 activity. Our study demonstrated that sedanolide enhanced cellular resistance to oxidative damage via the activation of the Kelch-like ECH-associated protein 1 (KEAP1)-NRF2 pathway.

The detoxification of quinones through NAD(P)H:quinone oxidoreductase (NQO1) is a crucial mechanism to maintain cellular homeostasis. The exposure to heavy metals, specifically methylmercury (MeHg), induces several antioxidant enzymes, including NQO1. The nuclear factor erythroid 2-related factor-2 (NRF2) is known to regulate the expression of Nqo1 gene and also the aryl hydrocarbon receptor (AHR) is another Nqo1 gene regulator. This co-regulation prompted us to investigate which transcription factor (NRF2 or AHR) orchestrates the regulation of NQO1 expression upon MeHg exposure. Therefore, we investigated how MeHg can modulate the level of NQO1 expression by exposing Hepa-1c1c7 cells to several concentrations of MeHg with and without the addition of NQO1 inducers, DL-sulforaphane (SUL) and 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). We found that the mRNA expression of Nqo1 is up-regulated by MeHg in time- as well as dose-dependent fashions. Additionally, MeHg increased the NQO1 at all expression levels with and without the presence of its inducers, SUL or TCDD. Furthermore, the MeHg-mediated increase of NQO1 expression was in parallel with a concurrent increase in the nuclear localization of NRF2 protein, but not that of AHR. Mechanistically, the antioxidant response element-driven reporter gene activity was induced by 215% upon MeHg exposure. Also, transfecting Hepa-1c1c7 with Nrf2 siRNA reduced the MeHg-induced NQO1 protein expression by 60%. In conclusion, our findings provide evidence supporting the hypothesis that MeHg upregulates the Nqo1 gene through a transcriptional mechanism at least in part via a NRF2-dependent mechanism.

Flavone is widely found in plants and plays an important role in plant defense against pests. Many pests, such as Helicoverpa armigera, use flavone as a cue to upregulate counter-defense genes for detoxification of flavone. Yet the spectrum of the flavone-inducible genes and their linked cis-regulatory elements remains unclear. In this study, 48 differentially expressed genes (DEGs) were found by RNA-seq. These DEGs were mainly concentrated in the retinol metabolism and drug metabolism-cytochrome P450 pathways. Further in silico analysis of the promoter regions of 24 upregulated genes predicted two motifs through MEME and five previously characterized cis-elements including CRE, TRE, EcRE, XRE-AhR and ARE. Functional analysis of the two predicted motifs and two different versions of ARE (named ARE1 and ARE2) in the promoter region of the flavone-inducible carboxylesterase gene CCE001j verified that the two motifs and ARE2 are not responsible for flavone induction of H. armigera counter-defense genes, whereas ARE1 is a new xenobiotic response element to flavone (XRE-Fla) and plays a decisive role in flavone induction of CCE001j. This study is of great significance for further understanding the antagonistic interaction between plants and herbivorous insects.

UNASSIGNED: Unexplained infertility refers to a diagnosis in which all standard examinations are usually normal and is statistically seen in approximately 30-40% of infertile couples and endometriosis encountered in 25-50% of patients with unexplained infertility. Unexplained infertility is thought to be closely associated with endometriosis and serum and peritoneal fluid levels of Fetuin-A is increased in patients with endometriosis. Fetuin-A is proposed as a diagnostic marker for endometriosis and has anti-inflammatory effects on several diseases. Oxidative stress also is central to the etiopathogenesis of infertility in women. The aim of this study was to evaluate serum Fetuin-A and oxidative stress parameter concentrations impact on unexplained infertility.
UNASSIGNED: In the study, serum Fetuin-A, IL-1β, CA I, TAS, TOS levels, and PON and ARE enzyme activities were measured using the Enzyme-Linked ImmunoSorbent Assay in the sera of diagnosed unexplained infertility females (n=44) and controls (n=41).
UNASSIGNED: There was no statistically significant difference between unexplained infertile patients and control groups in terms of serum IL-1β, CA I, OSI, and PON levels (p>0.05). Serum Fetuin-A and ARE levels were significantly higher in unexplained infertility compared with the control, whereas serum TAS and TOS levels were lower (p<0.05, p<0.01, p<0.05, p<0.05, respectively).
UNASSIGNED: It is thought that increased Fetuin-A levels may be a response to the inflammatory process and increased ARE activity may be a sign of the impaired oxidant-antioxidant balance in unexplained infertility. This may contribute to the pathogenesis of infertility, and the data obtained will make a significant contribution to new works to be done in this sense.

UNASSIGNED: The nuclear factor erythroid 2-related factor 2 (Nrf2) is a pivotal transcription factor that controls the expression of numerous cytoprotective genes and regulates cellular defense system against oxidative insults. Thus, activating the Nrf2 pathway is a promising strategy for the treatment of various chronic diseases characterized by oxidative stress.
UNASSIGNED: This review first discusses the biological effects of Nrf2 and the regulatory mechanism of Kelch-like ECH-associated protein 1-Nrf2-antioxidant response element (Keap1-Nrf2-ARE) pathway. Then, Nrf2 activators (2020-present) are summarized based on the mechanism of action. The case studies consist of chemical structures, biological activities, structural optimization, and clinical development.
UNASSIGNED: Extensive efforts have been devoted to developing novel Nrf2 activators with improved potency and drug-like properties. These Nrf2 activators have exhibited beneficial effects in in vitro and in vivo models of oxidative stress-related chronic diseases. However, some specific problems, such as target selectivity and brain blood barrier (BBB) permeability, still need to be addressed in the future.

The KEAP1-NRF2-ARE signaling pathway plays a central role in mediating the adaptive cellular stress response to oxidative and electrophilic chemicals. This canonical pathway has been extensively studied and reviewed in the past two decades, but rarely was it looked at from a quantitative signaling perspective. Signal amplification, i.e., ultrasensitivity, is crucially important for robust induction of antioxidant genes to appropriate levels that can adequately counteract the stresses. In this review article, we examined a number of well-known molecular events in the KEAP1-NRF2-ARE pathway from a quantitative perspective with a focus on how signal amplification can be achieved. We illustrated, by using a series of mathematical models, that redox-regulated protein sequestration, stabilization, translation, nuclear trafficking, DNA promoter binding, and transcriptional induction - which are embedded in the molecular network comprising KEAP1, NRF2, sMaf, p62, and BACH1 - may generate highly ultrasensitive NRF2 activation and antioxidant gene induction. The emergence and degree of ultrasensitivity depend on the strengths of protein-protein and protein-DNA interaction and protein abundances. A unique, quantitative understanding of signal amplification in the KEAP1-NRF2-ARE pathway will help to identify sensitive targets for the prevention and therapeutics of oxidative stress-related diseases and develop quantitative adverse outcome pathway models to facilitate the health risk assessment of oxidative chemicals.

Organic isothiocyanates (ITCs) are a class of anticancer agents which naturally result from the enzymatic degradation of glucosinolates produced by Brassica vegetables. Previous studies have demonstrated that the structure of an ITC impacts its potency and mode(s) of anticancer properties, opening the way to preparation and evaluation of synthetic, non-natural ITC analogues. This study describes the preparation of a library of 79 non-natural ITC analogues intended to probe further structure-activity relationships for aryl ITCs and second-generation, functionalized biaryl ITC variants. ITC candidates were subjected to bifurcated evaluation of antiproliferative and antioxidant response element (ARE)-induction capacity against human MCF-7 cells. The results of this study led to the identification of (1) several key structure-activity relationships and (2) lead ITCs demonstrating potent antiproliferative properties.