Antioxidants & Redox Signaling

Pepper fruit is a horticultural product worldwide consumed that has great nutritional and economic relevance. Besides the phenotypical changes that undergo pepper fruit during ripening, there are many associated modifications at transcriptomic, proteomic, biochemical, and metabolic levels. Nitric oxide (NO) and hydrogen sulfide (H₂S) are recognized signal molecules that can exert regulatory functions in diverse plant processes. This study aims at analyzing the interrelationship between NO and H₂S during fruit ripening.

Our data indicate that the H₂S-generating cytosolic L-cysteine desulfhydrase (LCD) and the mitochondrial D-cysteine desulfhydrase (DCD) activities are downregulated during ripening but this effect was reverted after NO treatment of fruits.

Using as a model the non-climacteric pepper fruits at different ripening stages and under an NO-enriched atmosphere, the activity of the H₂S-generating LCD and DCD was analyzed. LCD and DCD activities were downregulated during ripening, but this effect was reverted after NO treatment of fruits. The analysis of LCD activity by non-denaturing polyacrylamide gel electrophoresis (PAGE) allowed identifying three isozymes designated CaLCD I to CaLCD III, which were differentially modulated by NO and strictly dependent on pyridoxal 5′-phosphate (PLP). In vitro analyses of green fruit samples in the presence of different compounds including NO donors, peroxynitrite (ONOO⁻), and reducing agents such as reduced glutathione (GSH) and L-cysteine (L-Cys) triggered an almost 100% inhibition of CaLCD II and CaLCD III. This redox adaptation process of both enzymes could be cataloged as a hormesis phenomenon. The protein tyrosine (Tyr) nitration (an NO-promoted post-translational modification) of the recombinant LCD was corroborated by immunoblot and by mass spectrometry (MS) analyses. Among the 11 Tyr residues present in this enzyme, MS of the recombinant LCD enabled us to identify that Tyr82 and Tyr254 were nitrated by ONOO⁻, this occurring near the active center on the enzyme, where His237 and Lys260 together with the cofactor PLP are involved. These data support the relationship between NO and H₂S during pepper fruit ripening, since LCD and DCD are regulated by NO during this physiological event, and this could also be extrapolated to other plant species.

Protein persulfidation (the formation of RSSH), an evolutionarily conserved oxidative posttranslational modification in which thiol groups in cysteine residues are converted into persulfides, has emerged as one of the main mechanisms through which hydrogen sulfide (H₂S) conveys its signaling.

New methodological advances in persulfide labeling started unraveling the chemical biology of this modification and its role in (patho)physiology. Some of the key metabolic enzymes are regulated by persulfidation. RSSH levels are important for the cellular defense against oxidative injury, and they decrease with aging, leaving proteins vulnerable to oxidative damage. Persulfidation is dysregulated in many diseases.

A relatively new field of signaling by protein persulfidation still has many unanswered questions: the mechanism(s) of persulfide formation and transpersulfidation and the identification of “protein persulfidases,” the improvement of methods to monitor RSSH changes and identify protein targets, and understanding the mechanisms through which this modification controls important (patho)physiological functions.

Deep mechanistic studies using more selective and sensitive RSSH labeling techniques will provide high-resolution structural, functional, quantitative, and spatiotemporal information on RSSH dynamics and help with better understanding how H₂S-derived protein persulfidation affects protein structure and function in health and disease. This knowledge could pave the way for targeted drug design for a wide variety of pathologies. Antioxid. Redox Signal. 39, 19–39.

Hydrogen sulfide (H₂S) is a multitasking potent regulator that facilitates plant growth, development, and responses to environmental stimuli.

The important beneficial effects of H₂S in various aspects of plant physiology aroused the interest of this chemical for agriculture. Protein cysteine persulfidation has been recognized as the main reduction–oxidation (redox) regulatory mechanism of H₂S signaling. An increasing number of studies, including large-scale proteomic analyses and functional characterizations, have revealed that H₂S-mediated persulfidations directly regulate protein functions, altering downstream signaling in plants. To date, the importance of H₂S-mediated persulfidation in several abscisic acid signaling-controlling key proteins has been assessed as well as their role in stomatal movements, largely contributing to the understanding of the plant H₂S-regulatory mechanism.

The molecular mechanisms of the H₂S sensing and transduction in plants remain elusive. The correlations of H₂S-mediated persulfidation with other oxidative post-translational modifications of cysteines are still to be explored.

Implementation of advanced detection approaches for the spatiotemporal monitoring of H₂S levels in cells and the current proteomic profiling strategies for the identification and quantification of the cysteine site-specific persulfidation will provide insight into the H₂S signaling in plants. Antioxid. Redox Signal. 39, 40–58.

Ferroptosis is featured by the accumulation of polyunsaturated-lipid peroxidation on cellular membranes in an iron-dependent manner. Ferroptosis has been implicated in various pathophysiological processes, including cancer, neurodegeneration, and ischemia-reperfusion tissue injury. However, our understanding about the dynamic and context-specific regulation of ferroptosis remains incomplete.

As the major substrate for peroxidation, the cellular lipidome regulates ferroptosis sensitivity and execution by controlling the abundance and availability of polyunsaturated-lipids for peroxidative modifications. In turn, the cellular lipidome is regulated by a complex network of enzymes and transporters, as well as upstream layers of receptors, kinases, and transcription factors. A number of research has shed light on the link between lipid metabolism and ferroptosis. Here, we summarize our current knowledge on the role of the lipidome and associated protein regulators in various stages of ferroptosis, ranging from initiation, execution to cell death evasion by cells experiencing ferroptotic stress.

This review provides an overview of the mechanisms underlying lipid peroxidation and ferroptosis by discussing the lipid species that directly contribute to lipid peroxidation and ferroptosis, how cells regulate the abundances of these pro-ferroptosis lipids, how lipid peroxidation causes cell death, and how cells prevent and repair membrane lipid damage under ferroptotic conditions.

Cell fate regulation in vivo could be different from in vitro culture settings. We envision that a comprehensive and detailed understanding about these important questions in the dynamic regulation of ferroptosis in vivo will accelerate our development of ferroptosis-targeted therapies to improve human health.

Autophagy is a self-degrading process that determines cell fate in response to various environmental stresses. In contrast to autophagy-mediated cell survival, the signals, mechanisms, and effects of autophagy-dependent cell death remain obscure. The discovery of autophagy-dependent ferroptosis provides a paradigm for understanding the relationship between aberrant degradation pathways and excessive lipid peroxidation in driving regulated cell death.

Ferroptosis was originally described as an autophagy-independent and iron-mediated nonapoptotic cell death. Current studies reveal that the level of intracellular autophagy is positively correlated with ferroptosis sensitivity. Selective autophagic degradation of proteins (e.g., ferritin, SLC40A1, ARNTL, GPX4, and CDH2) or organelles (e.g., lipid droplets or mitochondria) promotes ferroptosis by inducing iron overload and/or lipid peroxidation. Several upstream autophagosome regulators (e.g., TMEM164), downstream autophagy receptors (e.g., HPCAL1), or danger signals (e.g., DCN) are selectively required for ferroptosis-related autophagy, but not for starvation-induced autophagy. The induction of autophagy-dependent ferroptosis is an effective approach to eliminate drug-resistant cancer cells.

How different organelles selectively activate autophagy to modulate ferroptosis sensitivity is not fully understood. Identifying direct protein effectors of ferroptotic cell death remains a challenge.

Further understanding of the molecular mechanics and immune consequences of autophagy-dependent ferroptosis is critical for the development of precision antitumor therapies. Antioxid. Redox Signal. 39, 79–101.

Cholesterol is a type of lipid that plays a crucial role in building and maintaining cell membranes, producing certain hormones, and aiding in digestion. The two main types of cholesterol are low-density lipoprotein and high-density lipoprotein, and maintaining a healthy balance between them is essential for cellular function and organism health.

Cholesterol metabolism is a complex and dynamic process that involves biosynthesis, uptake, efflux, transport, and esterification. Disruptions in cholesterol metabolism are implicated in all stages of cancer, contributing to drug resistance, immune evasion, and autophagy dysfunction. These disruptions have also been linked to various types of regulated cell death, such as apoptosis, anoikis, lysosome-dependent cell death, pyroptosis, NETosis, necroptosis, entosis, ferroptosis, alkaliptosis, immunogenic cell death, and paraptosis.

Understanding the complex interplay between cholesterol metabolism and cell death and their impact on cancer development and progression is still a significant challenge. In addition, there is currently a lack of reliable biomarkers that can accurately reflect cholesterol metabolism dysregulation in cancer.

To develop more specific and effective cholesterol metabolism-targeted therapies, a better understanding of the mechanisms by which cholesterol metabolism dysregulation contributes to cell death and cancer progression is needed. In addition, improving the accuracy and reliability of biomarkers will be crucial for monitoring and diagnosing cholesterol-related cancer subtypes and evaluating the effectiveness of cholesterol metabolism-targeted therapies. These efforts will require ongoing research and collaboration among multidisciplinary teams of scientists and clinicians. Antioxid. Redox Signal. 39, 102–140.

The lack of disease-modifying treatments for Alzheimer’s disease (AD) that substantially alter the course of the disease highlights the need for new biological models of disease progression and neurodegeneration. Oxidation of macromolecules within the brain, including lipids, proteins, and DNA, is believed to contribute to AD pathophysiology, concomitant with dysregulation of redox-active metals, such as iron. Creating a unified model of pathogenesis and progression underpinned by iron dysregulation and redox dysregulation in AD could lead to new therapeutic targets with disease-modifying potential.

Ferroptosis, which was named in 2012, is a necrotic form of regulated cell death that depends on both iron and lipid peroxidation. While it is distinct from other types of regulated cell death, ferroptosis is regarded as being mechanistically synonymous with oxytosis. The ferroptosis paradigm has great explanatory potential in describing how neurons degenerate and die in AD. At the molecular level, ferroptosis is executed by the lethal accumulation of phospholipid hydroperoxides generated by the iron-dependent peroxidation of polyunsaturated fatty acids, while the major defensive protein against ferroptosis is the selenoenzyme, glutathione peroxidase 4 (GPX4). An expanding network of protective proteins and pathways have also been identified to complement GPX4 in the protection of cells against ferroptosis, with a central role emerging for nuclear factor erythroid 2-related factor 2 (NRF2).

In this review, we provide a critical overview of the utility of ferroptosis and NRF2 dysfunction in understanding the iron- and lipid peroxide-associated neurodegeneration of AD.

Finally, we discuss how the ferroptosis paradigm in AD is providing a new spectrum of therapeutic targets. Antioxid. Redox Signal. 39, 141–161.

Ferroptosis, a type of oxidative cell death driven by unlimited lipid peroxidation, is emerging as a target for cancer therapy. Although mitochondrial dysfunction may lead to ferroptosis, the underlying molecular mechanisms and metabolic pathways for ferroptosis are incompletely understood. Here, we identify solute carrier family 25 member 22 (SLC25A22), a mitochondrial glutamate transporter, as a driver of ferroptosis resistance in pancreatic ductal adenocarcinoma (PDAC) cells.

The downregulation of SLC25A22 expression was associated with increased sensitivity to ferroptosis, but not to apoptosis. Mechanistically, on the one hand, SLC25A22-dependent NAPDH synthesis blocks ferroptotic cell death in PDAC cells through mediating the production of glutathione (GSH), the most important hydrophilic antioxidant. On the other hand, SLC25A22 promotes the expression of stearoyl-CoA desaturase in PDAC cells in an AMP-activated protein kinase-dependent manner, resulting in the production of antiferroptotic monounsaturated fatty acids (MUFAs). The animal study further confirms that SLC25A22 inhibits ferroptosis-mediated tumor suppression.

SLC25A22 is a novel metabolic repressor of ferroptosis by producing GSH and MUFAs.

These findings establish a previously unrecognized metabolic defense pathway to limit ferroptotic cell death in vitro and in vivo. Antioxid. Redox Signal. 39, 166–185.

Reactive oxygen species (ROS) are critical to normal cellular function with redox homeostasis achieved by balancing ROS production with removal through detoxification mechanisms. Many of the conventional chemotherapies used to treat colorectal cancer (CRC) derive a proportion of their cytotoxicity from ROS generation, and resistance to chemotherapy is associated with elevated detoxification mechanisms. Furthermore, cancer stem cells demonstrate elevated detoxification mechanisms making definitive treatment with existing chemotherapy challenging. In this article, we review the roles of ROS in normal and malignant colonic cell biology and how existing and emerging therapies might harness ROS for therapeutic benefit.

Recent publications have elucidated the contribution of ROS to the cytotoxicity of conventional chemotherapy alongside the emerging approaches of photodynamic therapy (PDT), sonodynamic therapy (SDT), and radiodynamic therapy (RDT), in which ROS are generated in response to excitatory light, sound, or X-ray stimuli to promote cancer cell apoptosis.

The majority of patients with metastatic CRC have a very poor prognosis with a 5-year survival of ∼13% making the need for new or more effective treatments an imperative.

Modulation of ROS through a combination of new and emerging therapies may improve the efficacy of current chemotherapy providing novel approaches to treat the otherwise resistant disease. Antioxid. Redox Signal. 39, 186–205.

The significance of ferroptosis in cancer therapeutics has now been unveiled. Specific ferroptosis inducers are expected as a promising strategy for cancer treatment, especially in cancers with epithelial mesenchymal transition and possibly in cancers with activated Hippo signaling pathways, both of which cause resistance to traditional chemotherapy but tend to show ferroptosis susceptibility.

Ferroptosis is a new form of regulated non-apoptotic cell death, which is characterized by iron-dependent lipid peroxidation, leading eventually to plasma membrane rupture. Its core mechanisms have been elucidated, consisting of a driving force as catalytic Fe(II)-dependent Fenton reaction and an incorporation of polyunsaturated fatty acids to membrane phospholipids via peroxisome-dependent and -independent pathways, and suppressing factors as prevention of lipid peroxidation with glutathione peroxidase 4 and direct membrane repair via coenzyme Q10 and ESCRT-III pathways.

Developments of ferroptosis inducers are in progress by nanotechnology-based drugs or by innovative engineering devices. Especially, low-temperature (non-thermal) plasma is a novel technology at the preclinical stage. The exposure can induce ferroptosis selectively in cancer cells rich in catalytic Fe(II).

We also summarize and discuss the recently uncovered responsible molecular mechanisms in association with iron metabolism, ferroptosis and cancer therapeutics. Targeting ferroptosis in addition to the current therapeutic modalities would be important to cure advanced-stage cancer. Antioxid. Redox Signal. 39, 206–223.