Person: Martínez Ruiz, Antonio
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First Name
Antonio
Last Name
Martínez Ruiz
Affiliation
Universidad Complutense de Madrid
Faculty / Institute
Farmacia
Department
Bioquímica y Biología Molecular
Area
Bioquímica y Biología Molecular
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6 results
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Now showing 1 - 6 of 6
Item Signalling by NO-induced protein S-nitrosylation and S-glutathionylation: Convergences and divergences(Cardiovascular research, 2007) Martínez Ruiz, Antonio; Lamas, SantiagoThe role of nitric oxide in several signalling routes has been clearly established. In recent years increasing attention has been paid to its ability to produce covalent protein post-translational modifications in conjunction with other reactive oxygen and nitrogen species. Among these, the modification of cysteine residues has been shown to be of particular importance due to the functional relevance of many of them. In this review, we focus on the modification of the cysteine thiol by incorporation of a NO moiety (S-nitrosylation) or of a glutathione moiety (S-glutathionylation). Both modifications are produced by different reactions induced by nitric oxide-related species. We discuss the differences and similarities of both modifications, and their relationships, in regard to the biochemical mechanisms that produce them, including the enzymatic activities that may catalyze some of them and their subcellular compartmentalization. Even when biochemical knowledge is one step ahead of the demonstration of their pathophysiological relevance, we also describe the potential role of both modifications in several processes in which both post-translational modifications are involved. © 2007 European Society of Cardiology. Published by Elsevier B.V. All rights reserved.Item Nitric Oxide Down-regulates Caveolin-3 Levels through the Interaction with Myogenin, Its Transcription Factor(Journal of biological chemistry, 2007) Martínez-Moreno, Mónica; Martínez Ruiz, Antonio; Álvarez-Barrientos, Alberto; Gavilanes Franco, Francisco; Lamas, Santiago; Rodríguez Crespo, José IgnacioCertain patients suffering from chronic diseases such as AIDS or cancer experience a constant cellular secretion of tumor necrosis factor and other pro-inflammatory cytokines that results in a continuous release of nitric oxide ( NO) to the bloodstream. One immediate consequence of the deleterious action of NO is weight loss and the progressive destruction of muscular mass in a process known as cachexia. We have previously reported that caveolin-3, a specific marker of muscle cells, becomes down-regulated by the action of NO on muscular myotubes. We describe herein that the changes observed in caveolin-3 levels are due to the alteration of the DNA binding activity of the muscular transcription factor myogenin. In the presence of NO, the binding of transcription factors from cell nuclear extracts of muscular tissues to the E boxes present in the caveolin-3 promoter become substantially reduced.When we purified recombinant myogenin and treated it with NO donors, we could detect its S-nitrosylation by three independent methods, suggesting that very likely one of the cysteine residues of the molecule is being modified. Given the role of myogenin as a regulatory protein that determines the level of multiple muscle genes expressed during late myogenesis, our results might represent a novel mode of regulation of muscle development under conditions of nitric oxide-mediated toxicity.Item Functional interplay between endothelial nitric oxide synthase and membrane type 1–matrix metalloproteinase in migrating endothelial cells(Blood, 2007) Genís, Laura; Gonzalo, Pilar; Tutor, Antonio S.; Gálvez, Beatriz G.; Martínez Ruiz, Antonio; Zaragoza, Carlos; Lamas, Santiago; Tryggvason, Karl; Apte, Suneel S.; Arroyo, Alicia G.Nitric oxide (NO) is essential for vascular homeostasis and is also a critical modulator of angiogenesis; however, the molecular mechanisms of NO action during angiogenesis remain elusive. We have investigated the potential relationship between NO and membrane type 1–matrix metalloproteinase (MT1-MMP) during endothelial migration and capillary tube formation. Endothelial NO synthase (eNOS) colocalizes with MT1-MMP at motilityassociated structures in migratory human endothelial cells (ECs); moreover, NO is produced at these structures and is released into the medium during EC migration. We have therefore addressed 2 questions: (1) the putative regulation of MT1-MMP by NO in migratory ECs; and (2) the requirement for MT1-MMP in NOinduced EC migration and tube formation. NO upregulates MT1-MMP membrane clustering on migratory human ECs, and this is accompanied by increased degradation of type I collagen substrate. MT1-MMP membrane expression and localization are impaired in lung ECs from eNOS-deficient mice, and these cells also show impaired migration and tube formation in vitro. Inhibition of MT1-MMP with a neutralizing antibody impairs NOinduced tube formation by human ECs, and NO-induced endothelial migration and tube formation are impaired in lung ECs from mice deficient in MT1-MMP. MT1-MMP thus appears to be a key molecular effector of NO during the EC migration and angiogenic processes, and is a potential therapeutic target for NO-associated vascular disorders.Item S-nitrosylation: a potential new paradigm in signal transduction(Cardiovascular Research, 2004) Martínez Ruiz, Antonio; Lamas, SantiagoMuch attention has been paid to nitric oxide (NO) research since its discovery as a physiological mediator in the cardiovascular system. In recent years, newer roles have been attributed to this molecule and its close relatives, termed collectively reactive nitrogen species (RNS). These roles relate to different mechanisms of protein modification, among which S-nitrosylation of cysteines has emerged as a potential new paradigm in signal transduction and regulation of protein function. We review here the chemical basis of this modification compared with other protein modifications related to nitric oxide, as well as the kind of specificity we can expect from it. We also review the current methodologies that can be applied to the study of S-nitrosylation and identification of S-nitrosylated proteins in cells, and detail the relevance of this modification in several proteins related to cardiovascular system.Item A “fluorescence switch” technique increases the sensitivity of proteomic detection and identification of S‐nitrosylated proteins(Proteomics, 2009) Tello, Daniel; Tarín, Carlos; Ahicart, Patricia; Bretón‐Romero, Rosa; Lamas, Santiago; Martínez Ruiz, AntonioProtein S-nitrosylation is a reversible post-translational modification of protein cysteines that is increasingly being considered as a signal transduction mechanism. The ‘‘biotin switch’’ technique marked the beginning of the study of the S-nitrosoproteome, based on the specific replacement of the labile S-nitrosylation by a more stable biotinylation that allowed further detection and purification. However, its application for proteomic studies is limited by its relatively low sensitivity. Thus, typical proteomic experiments require high quantities of protein extracts, which precludes the use of this method in a number of biological settings. We have developed a ‘‘fluorescence switch’’ technique that, when coupled to 2-DE proteomic methodologies, allows the detection and identification of S-nitrosylated proteins by using limited amounts of starting material, thus significantly improving the sensitivity. We have applied this methodology to detect proteins that become S-nitrosylated in endothelial cells when exposed to S-nitroso-L-cysteine, a physiological S-nitrosothiol, identifying already known S-nitrosylation targets, as well as proteins that are novel targets. This ‘‘fluorescence switch’’ approach also allowed us to identify several proteins that are denitrosylated by thioredoxin in cytokine-activated RAW264.7 (murine macrophage) cells. We believe that this method represents an improvement in order to approach the identification of S-nitrosylated proteins in physiological conditions.Item S-nitrosylation of Hsp90 promotes the inhibition of its ATPase and endothelial nitric oxide synthase regulatory activities(Proceedings of the National Academy of Science of the United States os America, 2005) Martínez Ruiz, Antonio; Villanueva, Laura; González de Orduña, Cecilia; López-Ferrer, Daniel; Higueras, María Ángeles; Tarín, Carlos; Rodríguez Crespo, José Ignacio; Vázquez, Jesús; Lamas, Santiago; Ignarro, Louis J.Nitric oxide is implicated in a variety of signaling pathways in different systems, notably in endothelial cells. Some of its effects can be exerted through covalent modifications of proteins and, among these modifications, increasing attention is being paid to S-nitrosylation as a signaling mechanism. In this work, we show by a variety of methods (ozone chemiluminescence, biotin switch, and mass spectrometry) that the molecular chaperone Hsp90 is a target of S-nitrosylation and identify a susceptible cysteine residue in the region of the C-terminal domain that interacts with endothelial nitric oxide synthase (eNOS). We also show that the modification occurs in endothelial cells when they are treated with S-nitrosoL-cysteine and when they are exposed to eNOS activators. Hsp90 ATPase activity and its positive effect on eNOS activity are both inhibited by S-nitrosylation. Together, these data suggest that S-nitrosylation may functionally regulate the general activities of Hsp90 and provide a feedback mechanism for limiting eNOS activation.