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Chem. 2017; 398(12): 1267–1293 Review Eugenia Belcastro, Caroline Gaucher, Alessandro Corti, Pierre Leroy, Isabelle Lartaud and Alfonso Pompella* Regulation of protein function by S-nitrosation and S-glutathionylation: processes and targets in cardiovascular pathophysiology https://doi.org/10.1515/hsz-2017-0150 aims to provide an update of available knowledge in the Received April 26, 2017; accepted August 7, 2017; previously field, with a special focus on the respective (sometimes published online August 19, 2017 competing and antagonistic) roles played by protein S-nitrosations and S-thionylations in biochemical and cel- Abstract: Decades of chemical, biochemical and patho- lular processes specifically pertaining to pathogenesis of physiological research have established the relevance cardiovascular diseases. of post-translational protein modifications induced by processes related to oxidative stress, with critical reflec- Keywords: cardiovascular diseases; glutathione; mixed tions on cellular signal transduction pathways. A great disulfides; nitric oxide; RNS; ROS; S-glutathionylation; deal of the so-called ‘redox regulation’ of cell function is S-nitrosation. in fact mediated through reactions promoted by reactive oxygen and nitrogen species on more or less specific ami- noacid residues in proteins, at various levels within the cell machinery. Modifications involving cysteine residues Introduction have received most attention, due to the critical roles they The role of nitric oxide (NO) in several signaling routes play in determining the structure/function correlates in has been clearly established (Grisham et al., 1999; Pacher proteins. The peculiar reactivity of these residues results et al., 2007). NO, one of the most important substances in two major classes of modifications, with incorpora- produced by the endothelium, plays a key role in home- tion of NO moieties (S-nitrosation, leading to formation ostasis maintenance and has gained recognition as a of protein S-nitrosothiols) or binding of low molecular crucial modulator in vascular pathophysiology. NO has weight thiols (S-thionylation, i.e. in particular S-glutath- a number of intracellular effects, e.g. vasorelaxation, ionylation, S-cysteinylglycinylation and S-cysteinylation). endothelial regeneration, inhibition of leukocyte chemo- A wide array of proteins have been thus analyzed in detail taxis, and platelet adhesion. The physiological chemistry as far as their susceptibility to either modification or both, of NO is complex, for it encompasses numerous potential and the resulting functional changes have been described reactions. Increasing attention has been paid to its ability in a number of experimental settings. The present review to produce covalent post-translational protein modifi- cations, and among these, the modification of cysteine *Corresponding author: Alfonso Pompella, Department of residues has received more attention due to the functional Translational Research NTMS, University of Pisa Medical School, Via relevance of many of them. The cysteine thiol may be modi- Roma 55, I-56126 Pisa, Italy, e-mail: [email protected] fied by incorporation of a NO moiety (S-nitrosation) or of a Eugenia Belcastro: CITHEFOR EA3452 “Drug targets, formulation glutathione moiety (S-glutathionylation). Both modifica- and preclinical assessment”, Faculté de Pharmacie, Université de Lorraine, 5 rue Albert Lebrun, BP 80403, F-54001 Nancy Cedex, tions may ensue from different reactions induced by nitric France; and Department of Translational Research NTMS, University oxide-related species, especially S-nitrosoglutathione of Pisa Medical School, Via Roma 55, I-56126 Pisa, Italy (GSNO). GSNO is one of the most important intracellular Caroline Gaucher, Pierre Leroy and Isabelle Lartaud: CITHEFOR S-nitrosothiols (Martínez-Ruiz and Lamas, 2007), and as EA3452 “Drug targets, formulation and preclinical assessment”, will be detailed below, a number of studies have investi- Faculté de Pharmacie, Université de Lorraine, 5 rue Albert Lebrun, BP 80403, F-54001 Nancy Cedex, France gated its potential use as a therapeutic agent in selected Alessandro Corti: Department of Translational Research NTMS, conditions, including cardiovascular dideases (Hornyák University of Pisa Medical School, Via Roma 55, I-56126 Pisa, Italy et al., 2011). Brought to you by | Universita di Pisa Authenticated Download Date | 3/27/19 1:01 PM 1268 E. Belcastro et al.: S-nitrosation and S-glutathionylation in cardiovascular pathophysiology In the present review, the first part will focus on the soluble guanylyl cyclase (sGC), producing increased con- role of NO as a signaling molecule, by analyzing the main centrations of cyclic guanosine monophosphate (cGMP). post-translational modifications (PTMs) it can induce in Cyclic GMP interacts with three types of intracellular pro- cellular proteins. The second part will explore the simi- teins: cGMP-dependent protein kinases (PKGs), cGMP-reg- larities, differences and commonalities between the pro- ulated ion channels, and cGMP-regulated cyclic nucleotide cesses of protein S-nitrosation and S-glutathionylation. phosphodiesterases (PDEs). Thus, cGMP can alter cell In fact, both modifications share many mechanistic and function through mechanisms dependent or independent functional features, making them solid candidates as of protein phosphorylation. Depending on the sites of NO general mechanisms for intracellular signal transduc- release and cGMP activation, different biological effects tion. Finally, the third section will focus on the identifi- can be observed. In vascular smooth muscle cells (VSMCs), cation of proteins individually modified by S-nitrosation increased cyclic GMP concentrations will activate cGMP- and S-gluthathionylation. The most abundant protein dependent kinases capable of decreasing intracellular thiol is the free cysteine of albumin, which makes of this calcium, thus producing relaxation (Moncada et al., 1991), abundant protein an inevitable target of both processes. whereas increased cGMP in platelets will decrease platelet S-nitrosylation of albumin has been recognized since the activation and adhesion to the surface of the endothelium beginning as a mechanism involved in the bioactivity of (Radomski and Moncada, 1993). endothelium-derived NO, at that time denoted as EDRF Conversely, the activation of iNOS, induced during (Stamler et al., 1992; Keaney et al., 1993). Our review will pathophysiological processes such as inflammation, pro- be focused on the resulting effects of these PTMs on other duces much higher concentrations of NO (>1 μm). iNOS protein targets critical in cardiovascular pathophysiology. is expressed physiologically, but is induced by certain The competition of the two processes on the same protein inflammatory cytokines (IL-1, INFγ, TNF-α), LPS and oxi- targets will also be discussed, taking into account the dual dizing agents. The effects of NO will depend on the site of ability of GSNO to both S-nitrosate as well as S-glutathio- its formation, concentrations achieved, as well as the type nylate proteins. of targeted tissue. Generally, modulation of protein function by NO can occur through three main reactions: 1. Nitrosylation, a reversible coordination of NO to tran- NO signaling in the cardiovascular sition metal ions in enzymes, such as ferrous (Fe2+) system heme prosthetic groups within the sGC enzyme, lead- ing to enzyme activation and increased formation of NO is a gaseous radical with a short half-life, continuously c G M P. synthesized from L-arginine by the nitric oxide synthase 2. Protein nitrosation, with formation of a covalent bond (NOS) (Palmer et al., 1988). There are three distinct iso- between NO and cysteine (S-nitrosation) or tryptophan forms of NOS that differ in structure and function (Stuehr, (N-nitrosation) residues. Modifications of free cysteine 1997). Endothelial NOS (eNOS) and neuronal NOS (nNOS) residues present at active sites of effector proteins and are constitutively expressed and are referred as Ca2+- peptides will change the activity or function of these dependent enzymes (Ayajiki et al., 1996) and generate proteins. This configures a mode of post-translational small amounts of NO for signaling. The third type is the protein modification as important as phosphorylation inducible isoform (iNOS), Ca2+-independent and inducible (Lima, 2010; Heikal, 2011). A number of studies have by immunological stimuli (Schulz et al., 1992). This latter focused on the mechanistic aspects of protein nitro- is activated in response to inflammation and generates sation (see e.g. Wolhuter and Eaton, 2017), however high amounts of NO (Knowles and Moncada, 1994). it has been shown that de-nitrosation reactions also From a functional point of view, NO acts as a chemical play critical roles in the control of nitrosated proteins messenger particularly in vascular and immune systems, levels as well as of NO release. In the specific case of where it participates in the regulation of a wide range of S-nitrosation, the formed S-nitrosothiols (RSNOs) can physiological processes. Its production within the cell is undergo spontaneous or assisted trans-nitrosations, finely adjusted to ensure appropriate effects. Indeed, in resulting in the transfer of the NO moiety from high physiological conditions, the low concentrations of NO molecular weight (protein) thiols