Ana I. Rojo

Bio: Ana I. Rojo is an academic researcher from Spanish National Research Council. The author has contributed to research in topics: Transcription factor & Neuroprotection. The author has an hindex of 39, co-authored 56 publications receiving 6448 citations. Previous affiliations of Ana I. Rojo include Autonomous University of Madrid & Complutense University of Madrid.

Guidelines for the use and interpretation of assays for monitoring autophagy (4th edition)

Abstract: In 2008, we published the first set of guidelines for standardizing research in autophagy. Since then, this topic has received increasing attention, and many scientists have entered the field. Our knowledge base and relevant new technologies have also been expanding. Thus, it is important to formulate on a regular basis updated guidelines for monitoring autophagy in different organisms. Despite numerous reviews, there continues to be confusion regarding acceptable methods to evaluate autophagy, especially in multicellular eukaryotes. Here, we present a set of guidelines for investigators to select and interpret methods to examine autophagy and related processes, and for reviewers to provide realistic and reasonable critiques of reports that are focused on these processes. These guidelines are not meant to be a dogmatic set of rules, because the appropriateness of any assay largely depends on the question being asked and the system being used. Moreover, no individual assay is perfect for every situation, calling for the use of multiple techniques to properly monitor autophagy in each experimental setting. Finally, several core components of the autophagy machinery have been implicated in distinct autophagic processes (canonical and noncanonical autophagy), implying that genetic approaches to block autophagy should rely on targeting two or more autophagy-related genes that ideally participate in distinct steps of the pathway. Along similar lines, because multiple proteins involved in autophagy also regulate other cellular pathways including apoptosis, not all of them can be used as a specific marker for bona fide autophagic responses. Here, we critically discuss current methods of assessing autophagy and the information they can, or cannot, provide. Our ultimate goal is to encourage intellectual and technical innovation in the field.

Therapeutic targeting of the NRF2 and KEAP1 partnership in chronic diseases

Abstract: The transcription factor NF-E2 p45-related factor 2 (NRF2; encoded by NFE2L2) and its principal negative regulator, the E3 ligase adaptor Kelch-like ECH-associated protein 1 (KEAP1), are critical in the maintenance of redox, metabolic and protein homeostasis, as well as the regulation of inflammation. Thus, NRF2 activation provides cytoprotection against numerous pathologies including chronic diseases of the lung and liver; autoimmune, neurodegenerative and metabolic disorders; and cancer initiation. One NRF2 activator has received clinical approval and several electrophilic modifiers of the cysteine-based sensor KEAP1 and inhibitors of its interaction with NRF2 are now in clinical development. However, challenges regarding target specificity, pharmacodynamic properties, efficacy and safety remain.

SCF/{beta}-TrCP promotes glycogen synthase kinase 3-dependent degradation of the Nrf2 transcription factor in a Keap1-independent manner.

Abstract: A disadvantage of aerobic life is the constant generation of potentially damaging reactive oxygen species (ROS). The intracellular levels of such species need to be tightly controlled in order to avoid oxidative stress. Transcription factor Nrf2 (NF-E2-related factor 2) plays a critical role in redox homeostasis since it increases the expression of many antioxidant and drug-metabolizing genes, including those encoding heme oxygenase 1 (HO-1), NADPH:quinone oxidoreductase 1, glutathione S-transferases, glutamate-cysteine ligase, and glutathione peroxidases, in response to oxidative and electrophile stressors (13). These genes all contain a common promoter enhancer called the antioxidant response element (ARE) and are transactivated by Nrf2. Because ROS play a role as intracellular signaling molecules for many physiological processes, Nrf2 can have an impact on numerous cell functions, ranging from differentiation and development to proliferation and inflammation. Therefore, Nrf2 activity influences neurodegenerative disease, cardiovascular disease, and cancer (3, 4, 14, 16, 17, 49, 53). While increased Nrf2 transcriptional activity enhances cellular antioxidant defenses and increases the capacity to detoxify drugs, it may also lead to unwanted side effects. For instance, in tumors, high levels of Nrf2 activity have been correlated with a poor prognosis (41). Indeed, high Nrf2 activity has not been favored during evolution (25), but its levels are restricted via both redox-dependent and redox-independent pathways in normal healthy cells (29). In normal cells, Keap1 (Kelch-like ECH-associated protein 1), an E3 ubiquitin ligase substrate adaptor, regulates the level of Nrf2 protein in a redox-dependent fashion (5, 20, 51). The interaction between Nrf2 and Keap1 occurs via a “two-site tethering” process, otherwise called the “hinge and latch” mechanism. In this model, two motifs, a high-affinity ETGE motif and a low-affinity DLG motif, within the N-terminal Neh2 domain of Nrf2 each interact with a separate Kelch repeat domain present in the Keap1 homodimer (40). Both the ETGE motif and the DLG motif are required for the transcription factor to be repressed by Keap1 (28). In addition to its interaction with Nrf2, Keap1 also binds Cullin 3 (Cul3), which forms a core E3 ubiquitin ligase complex through an association with Ring-box1 protein (Rbx1, also called Roc1) (5, 10, 20, 51). The Keap1-Cul3-Rbx1 complex is able to ubiquitinate Nrf2 and target it for proteasomal degradation only under normal redox conditions, and upon exposure to oxidants or electrophiles, Cys-151, Cys-273, and Cys-288 in Keap1 become modified, leading to disturbance of the interaction between Nrf2 and Keap1 (8, 21, 49, 50). Failure of Nrf2 to dock simultaneously onto both Kelch repeat domains enables it to escape ubiquitination by Cul3-Rbx1 (21, 32, 47, 50). Thus, stress-related modification of Keap1 results in Nrf2 stabilization, accumulation of the transcription factor in the nucleus, and upregulation of ARE-driven genes. Perturbation of the Nrf2-Keap1 complex by oxidants and electrophiles is considered the principal mechanism by which Nrf2 accumulates and induces the ARE-gene battery. However, other regulatory mechanisms must exist in order to explain the following: (i) how Nrf2 contributes to the basal expression of certain ARE-driven genes under normal homeostatic conditions, (ii) how Nrf2 activity returns to its low basal levels after the intracellular redox balance has been restored, and (iii) how Nrf2 activity is limited during oxidative and electrophile stress. Conventional cell signaling studies have suggested that Nrf2 might be regulated by protein phosphorylation (2, 6, 15, 18, 36, 44). Previously, we presented data suggesting that GSK-3β (glycogen synthase kinase 3β) influences the nuclear exclusion and inactivation of Nrf2 (37-39). However, the mechanistic connection between GSK-3 and Nrf2 remains largely unexplored. A number of studies have demonstrated that GSK-3 directs the ubiquitination and proteasomal degradation of various transcription factors and other proteins by SCF/β-TrCP; these include Snail (54), β-catenin (1, 22, 34), Gli2 and Gli3 (33, 48), Xom (55), Cdc 25a (19), FGD1 and -3 (11, 12), Mcl-1 (7), securin (24), prolactin receptor (46), and the phosphatase PHLPP1 (23). In these instances, GSK-3 phosphorylates a cluster of Ser/Thr residues in target proteins, which are then recognized by SCF/β-TrCP. In turn, the complex formed by SCF/β-TrCP binds the Cullin-1 (Cul1) scaffold protein to form a complete E3 ligase by association with a linker protein called Skp1 and with Rbx1. Therefore, β-TrCP is an adapter protein that contains a Skp1-binding site called F-box and a WD recognition domain for phosphorylated substrates in the consensus motif DpSGX(1-4)pS (9, 42). To date, the existence of a phosphodegron in Nrf2 has not been explored. In the present article, we report that Nrf2 is destabilized as a consequence of its phosphorylation by GSK-3 and subsequent ubiquitination by SCF/β-TrCP. This pathway represents an alternative mechanism to the Keap1-dependent degradation of Nrf2 and provides a means by which this transcription factor can be regulated in a redox-independent manner.

疟原虫var基因转换速率变化导致抗原变异[英]／Paul H, Robert P, Christodoulou Z, et al//Proc Natl Acad Sci U S A

Abstract: 抗原变异可使得多种致病微生物易于逃避宿主免疫应答。表达在感染红细胞表面的恶性疟原虫红细胞表面蛋白1（PfPMP1）与感染红细胞、内皮细胞、树突状细胞以及胎盘的单个或多个受体作用，在黏附及免疫逃避中起关键的作用。每个单倍体基因组var基因家族编码约60种成员，通过启动转录不同的var基因变异体为抗原变异提供了分子基础。

Role of nrf2 in oxidative stress and toxicity.

Abstract: Organismal life encounters reactive oxidants from internal metabolism and environmental toxicant exposure. Reactive oxygen and nitrogen species cause oxidative stress and are traditionally viewed as being harmful. On the other hand, controlled production of oxidants in normal cells serves useful purposes to regulate signaling pathways. Reactive oxidants are counterbalanced by complex antioxidant defense systems regulated by a web of pathways to ensure that the response to oxidants is adequate for the body's needs. A recurrent theme in oxidant signaling and antioxidant defense is reactive cysteine thiol–based redox signaling. The nuclear factor erythroid 2–related factor 2 (Nrf2) is an emerging regulator of cellular resistance to oxidants. Nrf2 controls the basal and induced expression of an array of antioxidant response element–dependent genes to regulate the physiological and pathophysiological outcomes of oxidant exposure. This review discusses the impact of Nrf2 on oxidative stress and toxicity and how...

Crosstalk of reactive oxygen species and NF-κB signaling.

Abstract: NF-κB proteins are a family of transcription factors that are of central importance in inflammation and immunity. NF-κB also plays important roles in other processes, including development, cell growth and survival, and proliferation, and is involved in many pathological conditions. Reactive Oxygen Species (ROS) are created by a variety of cellular processes as part of cellular signaling events. While certain NF-κB-regulated genes play a major role in regulating the amount of ROS in the cell, ROS have various inhibitory or stimulatory roles in NF-κB signaling. Here we review the regulation of ROS levels by NF-κB targets and various ways in which ROS have been proposed to impact NF-κB signaling pathways.

Heme oxygenase-1/carbon monoxide: from basic science to therapeutic applications.

Abstract: The heme oxygenases, which consist of constitutive and inducible isozymes (HO-1, HO-2), catalyze the rate-limiting step in the metabolic conversion of heme to the bile pigments (i.e., biliverdin and bilirubin) and thus constitute a major intracellular source of iron and carbon monoxide (CO). In recent years, endogenously produced CO has been shown to possess intriguing signaling properties affecting numerous critical cellular functions including but not limited to inflammation, cellular proliferation, and apoptotic cell death. The era of gaseous molecules in biomedical research and human diseases initiated with the discovery that the endothelial cell-derived relaxing factor was identical to the gaseous molecule nitric oxide (NO). The discovery that endogenously produced gaseous molecules such as NO and now CO can impart potent physiological and biological effector functions truly represented a paradigm shift and unraveled new avenues of intense investigations. This review covers the molecular and biochemical characterization of HOs, with a discussion on the mechanisms of signal transduction and gene regulation that mediate the induction of HO-1 by environmental stress. Furthermore, the current understanding of the functional significance of HO shall be discussed from the perspective of each of the metabolic by-products, with a special emphasis on CO. Finally, this presentation aspires to lay a foundation for potential future clinical applications of these systems.

The Endocannabinoid System as an Emerging Target of Pharmacotherapy

Abstract: The recent identification of cannabinoid receptors and their endogenous lipid ligands has triggered an exponential growth of studies exploring the endocannabinoid system and its regulatory functions in health and disease. Such studies have been greatly facilitated by the introduction of selective cannabinoid receptor antagonists and inhibitors of endocannabinoid metabolism and transport, as well as mice deficient in cannabinoid receptors or the endocannabinoid-degrading enzyme fatty acid amidohydrolase. In the past decade, the endocannabinoid system has been implicated in a growing number of physiological functions, both in the central and peripheral nervous systems and in peripheral organs. More importantly, modulating the activity of the endocannabinoid system turned out to hold therapeutic promise in a wide range of disparate diseases and pathological conditions, ranging from mood and anxiety disorders, movement disorders such as Parkinson9s and Huntington9s disease, neuropathic pain, multiple sclerosis and spinal cord injury, to cancer, atherosclerosis, myocardial infarction, stroke, hypertension, glaucoma, obesity/metabolic syndrome, and osteoporosis, to name just a few. An impediment to the development of cannabinoid medications has been the socially unacceptable psychoactive properties of plant-derived or synthetic agonists, mediated by CB 1 receptors. However, this problem does not arise when the therapeutic aim is achieved by treatment with a CB 1 receptor antagonist, such as in obesity, and may also be absent when the action of endocannabinoids is enhanced indirectly through blocking their metabolism or transport. The use of selective CB 2 receptor agonists, which lack psychoactive properties, could represent another promising avenue for certain conditions. The abuse potential of plant-derived cannabinoids may also be limited through the use of preparations with controlled composition and the careful selection of dose and route of administration. The growing number of preclinical studies and clinical trials with compounds that modulate the endocannabinoid system will probably result in novel therapeutic approaches in a number of diseases for which current treatments do not fully address the patients9 need. Here, we provide a comprehensive overview on the current state of knowledge of the endocannabinoid system as a target of pharmacotherapy.