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Cardiovascular Research 2004 62(1):43-52; doi:10.1016/j.cardiores.2004.01.013
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Copyright © 2004, European Society of Cardiology

S-nitrosylation: a potential new paradigm in signal transduction

Antonio Martínez-Ruiza,b and Santiago Lamas*,a,b

aCentro de Investigaciones Biológicas, Consejo Superior de Investigaciones Científicas and Instituto "Reina Sofía" de Investigaciones Nefrológicas, Ramiro de Maeztu, 9, Madrid E-28040, Spain
bFundación Centro Nacional de Investigaciones Cardiovasculares Carlos III (CNIC), Ronda de Poniente, 5, Tres Cantos E-28760, Madrid, Spain

* Corresponding author. Tel.: +34-91-8373112x4302; fax: +34-91-5360432. Email address: slamas{at}cib.csic.es

Received 24 November 2003; revised 26 December 2003; accepted 8 January 2004


   Abstract
Top
 Abstract
1. Introduction
 2. The chemical basis...
 3. Specificity and spatio...
 4. Current methodologies for...
 5. Regulation of protein...
 6. Conclusions and perspectives
 Acknowledgments
 References
 
Much 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.

KEYWORDS Signal transduction; Nitric oxide


   1. Introduction
Top
 Abstract
 1. Introduction
2. The chemical basis...
 3. Specificity and spatio...
 4. Current methodologies for...
 5. Regulation of protein...
 6. Conclusions and perspectives
 Acknowledgments
 References
 
Since the discovery of nitric oxide (NO) as a physiological mediator in the early 1980s, research devoted to the biological role of this molecule has evolved spectacularly, both in its intensity and extent. While the roles of NO as a modulator of vascular tone, neurotransmission and an effector in some immune responses are widely recognized, newer and attractive roles have emerged over the past 10 years. These relate to the capacity of NO to interact with and modify a wide variety of other molecules: free radicals such as the superoxide anion, key redox regulators such as glutathione, and macromolecules (DNA and proteins). This lends a basis for NO to regulate crucial processes within the cell such as the response to redox perturbations, protein function, and gene expression through non-enzymatic modifications. S-nitrosylation of proteins is one potential mechanism underlying these events that has attracted increasing interest in recent years. In the first part of this review, we briefly describe the chemical basis of the interaction of NO with heme groups, thiol groups and other radicals before discussing the specificity of these processes and describe the spatio-temporal conditions which might lead to S-nitrosylation within cells. In the subsequent sections, the currently available methodologies for the study of S-nitrosylation are reviewed, and examples of S-nitrosylation are examined in detail, with special focus on characterized in vivo examples and on proteins related to the cardiovascular system.


   2. The chemical basis of NO–protein interactions
Top
 Abstract
 1. Introduction
 2. The chemical basis...
3. Specificity and spatio...
 4. Current methodologies for...
 5. Regulation of protein...
 6. Conclusions and perspectives
 Acknowledgments
 References
 
Nitric oxide can modify proteins via different chemical processes. These either involve direct interaction with the protein by NO itself, or the prior production of "reactive nitrogen species" (RNS), formed as a result of the interaction of NO with other radicals and oxygen. The most well-known direct modification is the formation of complex (or coordinating) bonds between NO and transition metal ions within heme groups, this being the basis for the activation of soluble guanylate cyclase [1]. Other examples of NO interactions with heme groups include cytochrome P450 [2] and haemoglobin [3]. NO can also bind to non-heme iron proteins [4] and other transition metals. In the case of cytochrome c oxidase (complex IV of the mitochondrial respiratory chain), which contains two hemes and two copper centres, NO and oxygen compete for the binding to the active centres, but the exact mechanism of enzyme inhibition by NO is still under discussion [5,6].

Incorporation of NO moieties by covalent bonding to protein groups is chemically possible in the case of cysteine thiols, tryptophan indols and amines (lysine and N-terminal) [7–9], although S-nitrosylation, the formation of the S–NO bond by cysteine thiol nitrosylation, can be considered the most important, because of its higher reactivity, occurrence in physiological conditions and its influence on many protein functions.

The usage of the terms "nitrosation" and "nitrosylation" needs some clarification. From the chemical point of view, "nitrosation" means the addition of a nitroso group, i.e., the NO diatomic group; "nitrosylation" means the addition of a nitrosyl group, NO, stressing the concept of the addition of a chemical group that, if it were free, it would be a radical (in analogy to other chemical additions containing the -yl-particle: acetylation, phosphorylation, etc.). The fact is that in the nitric oxide species, both atomic groups are the same, as the radical is in itself relatively stable. Even so, some authors prefer to distinguish the incorporation of the ·NO radical to a metal by a complex (or coordinating) bond as "nitrosylation", and the covalent incorporation of a NO diatomic group to another chemical group (regardless of the reaction mechanism) as "nitrosation" (in accordance to the nomenclature, which uses the term "nitroso" when defining the names of the resulting compounds). However, the increasing recognition of the functional importance of this post-translational modification, and the widespread inclusion of the particle "-yl-" in terms describing other post-translational modifications (glycosylation, phosphorylation) has led pioneer investigators in the field to make a case for the use of nitrosylation both for thiols and metals [10,11]. In any case, incorporation to a thiol can be clearly distinguished because of the prefix "S-", referring to the incorporation of the NO moiety to a sulfur atom to form the S–NO bond: "S-nitrosation" or "S-nitrosylation".

It is also important to distinguish nitrosylation/nitrosation from nitration, a term which describes incorporation of a nitro triatomic group (–NO2) and which, in protein chemistry, is generally used to describe the incorporation of that group at position 3 of the phenolic ring of tyrosine residues. This modification has also been thoroughly studied in recent times, an is clearly related to the formation of peroxynitrite (for reviews, see Refs. [12,13]).

It is generally accepted that direct reaction of the ·NO radical with a thiol does not yield the nitrosothiol [14,15], although a mechanism with additional participation of oxygen species has been proposed [16]. Previous reaction of NO with O2 is considered to be necessary for S-nitrosylation, via formation of higher nitrogen oxides, among which N2O3 is thought to be the quintessential S-nitrosylating species [17,18]. This reaction of NO and O2 can be favoured in membrane environments, where NO and O2 may reach higher concentrations [19], or even in hydrophobic protein microenvironments [20]. N2O3 can be partially dissociated into [+ON··NO2–], which favours the reaction of the nitrosonium (NO+) moiety with the nucleophile sulfur atom. Thus, S-nitrosylation can be better understood considering the transfer of nitrosonium and not of ·NO. This happens also in the process of transnitrosation, or transfer of the S-nitrosylation between a nitrosothiol and another thiol [21–23]. This mechanism of protein S-nitrosylation is of biological relevance, especially due to the high concentrations of intracellular low molecular mass thiols like glutathione. Nitrite is also a nitrosylating agent in acidic environments, where it forms nitrous acid, HNO2 [11,24], although its occurrence in physiological systems would be restricted to highly acidic environments.

Formation of nitrosothiols can be favoured in more ionisable cysteines, such as those in which the thiolate anion can be stabilized by acid–base interactions with neighbouring groups, either belonging to adjacent residues in the primary sequence (which is the basis of the consensus sequence proposed by Lipton and Stamler [25]) or just in the proximity in the three-dimensional structure [26–28]. Thus, in the context of such an "acid–base motif", the pKa of different cysteines in a protein would be an important factor in determining the occurrence of S-nitrosylation. It is important to note that in many instances where S-nitrosylation has been described, the protein cysteines involved are also oxidized, and tend to form disulfide bonds. Also, nitrosylation of thiols can chemically induce the formation of a disulfide bond [21]. Given that the most abundant thiol in the cytoplasm is glutathione, S-nitrosylation is likely to promote S-glutathionylation: the incorporation of glutathione into proteins via mixed disulfide bonds. S-glutathionylation is increasingly recognized as an important post-translational modification capable of regulating protein function [29–31]. This kind of reactive environment for cysteines is commonly encountered in the active centres of enzymes, and might favour the alteration of protein function by S-nitrosylation (or by other thiol modifications).


   3. Specificity and spatio-temporal conditions for S-nitrosylation
Top
 Abstract
 1. Introduction
 2. The chemical basis...
 3. Specificity and spatio...
4. Current methodologies for...
 5. Regulation of protein...
 6. Conclusions and perspectives
 Acknowledgments
 References
 
Many comparisons have been made between S-nitrosylation and phosphorylation as signalling mechanisms in the cell [32,33]. The important issue here is not whether S-nitrosylation is used by the cell as a signalling mechanism, but rather to understand the particular ways in which the cell can use it, and try to delimitate its specificity and cellular functionality.

Although both S-nitrosylation and phosphorylation are covalent modifications of protein residues, the crucial difference is that phosphorylation is enzyme driven, whereas S-nitrosylation is achieved through the non-catalysed chemical modification of a protein residue. Thus, in S-nitrosylation, the reaction specificity does not rely on the recognition of a target structure by an enzyme. Instead it depends solely on the chemical reactivity between the nitrosylating agent and the target. Several factors can be identified as involved in this kind of specificity.

3.1. Reactivity of the target protein residue
Analysis of S-nitrosylation in several proteins shows that not all protein cysteines that remain in the free-thiol state become nitrosylated [27,34,35]. Finding a common structural (sequence or three-dimensional) motif that might determine this reactivity has not been straightforward. As we have already seen, more ionisable cysteines are among the more nitrosylable, but cysteines in hydrophobic environments may also be candidates for S-nitrosylation. Thus, as the residue environment is what determines its reactivity, chemical specificity does not occur at the molecular (protein) level, but at the "submolecular" (atomic) level. Also, as the modification can affect similar groups in different proteins, the concept of "plurimolecular" functional specifity has been postulated [36].

3.2. Concentration
While chemical reactivity can be expressed in terms of thermodynamic and kinetic constants, the occurrence of a reaction is determined also by the concentrations of the species involved: in this case, the nitrosylating agent and the protein. This basic chemical concept is usually forgotten when biochemical pathways are described because the enzymatic catalysis diminishes its influence. In the case of S-nitrosylation, where the rate of the chemical reaction is not enzymatically determined, this point can be crucial. It depends on the following facts:

(1) Production of NO and related RNS. NO production is a key step in S-nitrosylation, and originates by a tightly regulated group of enzymes, the nitric oxide synthases (NOS). As NO is not the main nitrosylating agent, formation of other RNS, for example by reaction with O2 or O2–, has to be considered, taking into account that the NOS enzymes can also be responsible for the production of different RNS and ROS, including O2– [37,38] (Fig. 1).
(2) Localization. Subcellular compartmentalization can be extremely important in determining local concentrations of biochemical reagents. There is growing evidence that at least some S-nitrosylable proteins are physically associated to eNOS or nNOS (reviewed in Refs. [33,39–41]), and that the subcellular compartmentalization of these enzymes is tightly regulated. Using a modified yeast two-hybrid system, a recent report screened protein interactions of procaspase-3 (a known target of S-nitrosylation) and confirmed that this protein interacts with eNOS, nNOS and acid sphingomyelinase inside cells [42]. Localization specificity may not apply to iNOS, as the amounts of NO generated by iNOS are of a higher magnitude, and form part of a non-constitutive response programme related to infection and inflammation [43].


Figure 1
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  		Fig. 1 Chemical relationship among different reactive oxygen species (ROS) and reactive nitrogen species (RNS), and their impact on some post-translational modification of proteins. GSH: glutathione; GSSG: glutathione disulfide; GSNO: S-nitrosoglutathione. ROS chemistry is simplified in the scheme by O2– formation from O2. Arrowheads which join mean reaction among species.

 
3.3. Stability of the bond
S-nitrosylation is a very labile covalent modification under physiological conditions. As with its synthesis, cleavage of this bond can occur without the help of specific enzymes, although several enzymes have been described which can break at least low mass S-nitrosothiols at a relatively high KM (reviewed in Ref. [39]). The cleavage reaction might be accelerated by reaction with transition metals [44], especially copper(I) [24], or by transnitrosation, among other mechanisms. Some researchers have stressed the potential of denitrosylation as an important intracellular signalling mechanism in its own right, and this area requires further investigation [45,46].


   4. Current methodologies for the detection of S-nitrosylation
Top
 Abstract
 1. Introduction
 2. The chemical basis...
 3. Specificity and spatio...
 4. Current methodologies for...
5. Regulation of protein...
 6. Conclusions and perspectives
 Acknowledgments
 References
 
The major methodological problems in the study of protein S-nitrosylation are the lability of the S–NO bond and the fact that the level of endogenously nitrosylated proteins in the cell can be very low, hampering in vivo detection of individual S-nitrosylated proteins (Fig. 2). Thus, although over 100 proteins have been reported to be S-nitrosylated in vitro, very few have been described in their cellular environment [32]. Moreover, artifactual nitrosylation can be produced during manipulation, so S-nitrosylation of a protein should be confirmed by several methods.


Figure 2
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  		Fig. 2 Overview of techniques for detection and quantification of S-nitrosylated proteins. See text for details and references. ECD: electrochemical detection; NMR: nuclear magnetic resonance.

 
There are a number of methods for the "direct" detection of the S–NO bond. These are measurement of its characteristic absorbance band at 340 nm (although the sensitivity of this method is low); electrospray ionization mass spectrometry (ESI-MS) [47–49] (the widely used laser ionization, MALDI, breaks the S–NO bond [50]); electrochemical detection after liquid chromatography [51]; and NMR with 15N [52]. "Indirect" chemical methods require cleavage of the S–NO bond and detection of the species formed, NO or nitrite. Ozone chemiluminescence can measure NO released from nitrosothiols by photolytic cleavage [51], specific reduction with Cu+/cysteine at pH 6 [53], or reduction with I2/I– [54]. Heterolytic cleavage with HgCl2 produces nitrite, which can be measured by colorimetric or fluorometric methods [55,56].

The methods outlined above all provide a measure of the amount of nitrosothiols in a sample. To detect individual S-nitrosylated proteins in cells, prior immunoprecipitation with specific antibodies followed by reduction with Cu+/cysteine or I2/I– and chemiluminescence has been used [57,58]. The sensitivity of this approach is, however, limited by the amount of protein immunoprecipitated. A rabbit antiserum raised against S-nitroso-BSA is commercially available (Calbiochem) and can be used in some immunological applications. Its specifity can be verified by comparison with a sample treated with HgCl2 in which the nitrosothiols are broken [46]. Successful immunoprecipitation with this antibody followed by detection of individual proteins by immunoblot has been described only recently [59]. The "biotin switch" method (see below) has also been used in combination with immunoblot to confirm modification of individual proteins [60].

The rapid development of proteomics in recent years has made possible the study of the proteomes, the pools of proteins present in cells, under different conditions. A major goal of specialized proteomics is the characterization of subproteomes that undergo specific post-translational modifications. Regarding S-nitrosylation, the study of the "nitrosylome" (that pool of proteins which are S-nitrosylated in cells) would identify new targets for this modification. More specifically, this approach could be used to evaluate the feasibility of the signal transduction mechanisms postulated on the basis of in vitro studies. Again, technical difficulties have hampered study in this area, and the only approach which has so far been fruitful is an indirect one, called the "biotin switch" method, in which the cysteine-bound NO is replaced by a biotin label [60]. Several chemical steps are required for this conversion, and the method is designed to be sufficiently specific [60], although it lacks sensitivity [33]. Proteomic identification of S-nitrosylated proteins with the biotin switch method has been achieved in extracts from brain [60] and mesangial cells [61], and we have applied the method to intact endothelial cells [62]. In all cases, extracts or intact cells were first treated with nitrosylating agents, thus leading to elevated levels of S-nitrosylated proteins. Further efforts will be needed to increase the sensitivity of proteomic methodologies so that they can be applied in more physiological conditions, where S-nitrosylation might be more subtly regulated.


   5. Regulation of protein function by S-nitrosylation
Top
 Abstract
 1. Introduction
 2. The chemical basis...
 3. Specificity and spatio...
 4. Current methodologies for...
 5. Regulation of protein...
6. Conclusions and perspectives
 Acknowledgments
 References
 
S-nitrosylation has been shown to modify the function of many proteins in vitro (for a list, see Ref. [63]). In the following section, we describe examples where the phenomenon has been well characterized, and which are all related to the common themes of gene expression regulation or relevance for the cardiovascular system.

5.1 P21Ras
The oncoprotein GTPase p21Ras is one of the best-characterized examples of S-nitrosylation of a protein, and of how this might affect its signalling properties. First, it was described that NO and higher nitrogen oxides (NOx), activated p21Ras signalling in cells, and that this protein was implicated in the activation of NF-{kappa}B by NO. This study with recombinant p21Ras showed that this effect was associated with the formation of 1 nitrosothiol per molecule [34]. The modification was confirmed by ESI-MS [47], and the affected residue mapped to cysteine 118 [48], which is located in a highly conserved motif (NKXD) implicated in nucleotide binding. Detailed structural studies using NMR have shown that the protein forms that are S-nitrosylated or mutated in the cysteine do not differ from unmodified p21Ras in their overall structures. The activity of the mutant protein is normal, although it is not influenced by nitric oxide [28,64]. Recent work has established that stably nitrosylated p21Ras does not differ in activity from the unmodified form, and the activity is altered only during the process of the nitrosylating reaction itself [28]. Thus, further information on the actual role of S-nitrosylated Ras is needed.

5.2. NF-{kappa}B
This transcription factor is involved in the response to many different stimuli, including oxidative stress, and regulates the expression of a wide variety of genes. These include interleukin 2, NOS2 (iNOS gene), and tumour necrosis factor {alpha}, all of which participate in crucial biological pathways such as inflammation and the protection from apoptosis. It has a complex regulation, its release from the inhibitory molecule I{kappa}B being a key step. This protein is degraded in a highly regulated way, which allows NF-{kappa}B to translocate to the nucleus where it can bind to DNA and activate transcription [65]. Although it is activated in situations of oxidative stress, the binding of NF-{kappa}B to cis regulatory elements depends on a specific cysteine in the p50 subunit, cysteine 62, being in the reduced state [66,67]. This cysteine is located in the DNA-binding region, and is surrounded by basic residues which confer a high reactivity to it. It is susceptible to oxidation and formation of disulfide bonds, both with another p50 molecule and with glutathione [66,68].

A role for p50 cysteine 62 has been established in the inhibition by S-nitrosothiols of recombinant NF-{kappa}B binding to DNA [69]. De la Torre et al. [70,71] confirmed S-nitrosylation of recombinant p50, and calculated its DNA binding kinetics. However, NO can influence the intracellular activity of NF-{kappa}B in many ways (reviewed in Refs. [72]), and against this background the importance of p50 S-nitrosylation remains unproven. Only two reports exist confirming the presence of this modification inside cells. Both studies relied on the immunoprecipitation of overexpressed p50 from cells, with detection of the nitrosothiol by photolysis-chemiluminescence in the case of kidney cells [73], and absorbance at 340 nm in macrophages activated by LPS [74]. There are also reports of inactivation of DNA-binding activity consistent with S-nitrosylation, but the link has not been demonstrated definitively [75,76]. Interestingly, S-nitrosylation of p50 may be dependent on the cell type [76].

5.3. Hypoxia-inducible factors (HIF)
Hypoxia-inducible factors (HIF) are a family of transcription factors that are activated in response to low oxygen concentrations, and they form part of the PAS superfamily of conditional transcription factors [77]. They are heterodimers of two PAS proteins: a constitutive subunit, called ARNT ("aryl receptor nuclear translocator") or HIF-β; and a regulatory one called HIF-{alpha}. In humans, two {alpha} subunits have been described: HIF-1{alpha} and HIF-2{alpha}, also known as EPAS ("endothelial PAS protein"). Under normoxic conditions, these subunits are specifically hydroxylated at proline and arginine residues, and these modifications promote ubiquitination and degradation, as well as inhibiting the transactivation capacity. Under hypoxic conditions, these hydroxylations are impaired, leading to accumulation of HIF, translocation to the nucleus and restoration of the transactivation capacity. When activated, HIFs induce a number of genes, such as erythropoietin or VEGF, involved in responses to hypoxia.

The effect of NO-related compounds on HIF activation is complex, and is currently an area of active research. A dual role for NO, depending on the cellular O2 concentration, is now clear. This explains initial observations that NO (as well as CO) inhibited hypoxic activation of HIF [78,79] while, under normoxic conditions, NO activates HIF activity in different cell lines [80,81]. The latter mechanism has been explained by NO-mediated inhibition of the Fe2+-dependent activity of the prolyl hydroxylases, which leads to accumulation of the HIF-{alpha} subunits [82,83]. In this scenario, NO is acting as an autocrine or paracrine mimicker of hypoxia. The dual effect considered in this way has been thoroughly reviewed recently by Brüne [84]. More recently, it has been found that, during 1% hypoxia, the previously observed competitive inhibition of mitochondrial respiration by added NO increases the availability of O2 in the cell. Moreover, the effect of NO in HIF-1{alpha} stabilization could be mimicked by mitochondrial respiration inhibitors in a ROS-independent manner. This suggests that the availability of O2 for other enzymatic systems in the cells would be increased by NO's repression of mitochondrial respiration, increasing the activity of the prolyl hydroxylases [85].

Recent observations suggest that the effect of endogenously produced NO depends on its concentration, and published data in the literature should be interpreted in light of this. When NO is produced at high concentration (>1 µM), it promotes stabilization of HIF-1{alpha}, both in normoxia and relative hypoxia (3%), probably by inhibiting the prolyl hydroxylases. However, when NO is produced at low concentrations (<400 nM), its effect is only seen in hypoxia, when it impairs HIF-1{alpha} in a mitochondria-dependent manner [86]. This is thus an example of how the availability of molecules like O2 and NO, which can be thought of as chemical mediators or effectors, can be critical for the perception of cellular signals that they affect.

HIF is also subject to regulation by more subtle mechanisms, which can might explain some of the observed effects of NO. Transactivation activity is enhanced by binding to CBP/p300. This binding is also controlled by hydroxylation, in this case of an asparagine residue [87], and recent reports revealed that the S-nitrosylation of cysteine 800 of HIF-1{alpha} favours this interaction [88,89]. However, these studies rely on the indirect "biotin switch" method of detecting S-nitrosylation, which is not reliable for the quantification described. Finally, there might be a divergence in the regulation of the two known human HIF-{alpha} subunits. EPAS/HIF-2{alpha} bears a cysteine in the DNA binding domain which is substituted by serine in HIF-1{alpha}, and which is subject to redox regulation [90], probably including S-nitrosylation.

5.4. Zinc finger transcription factors
The majority of transcription factors include zinc fingers as DNA binding motifs. These are folding motifs that contain a zinc atom bound to four complexing residues, cysteines and histidines, with the cysteines being in the thiolate form. Research published during the last years suggests that these structural motifs that bind DNA could also act as sensors of oxidative or nitrosative stress, triggering integrated programmes of gene regulation by structurally related transcription factors (reviewed in Ref. [91]). S-nitrosothiols can inhibit the DNA binding activity of the transcription factors Sp1 and EGR-1, and of nuclear receptor superfamily members such as the vitamin D3 receptor (VDR), the retinoid X receptor (RXR) and the glucocorticoid receptor [92–94]. This inactivation might result from S-nitrosylation of a thiolate group involved in zinc binding. This would disrupt the structure of the zinc finger, and perhaps lead to other cysteine modifications, such as disulfide formation. It is interesting that these zinc fingers are protected from modification when they are bound to DNA, and the functionality of zinc fingers was restored by treatment with dithiothreitol [92,93], which could control the signal response.

One possibility is that an increment of nitrosylating reagents inside cells would reduce transcription driven by zinc finger transcription factors. This has been demonstrated in the case of IL-2 expression in response to IL-1β, which is dependent on Sp1 and EGR-1, and which can be inhibited by S-nitroso-L-cysteine [92]. This phenomenon is seen in reverse in the case of TNF-{alpha} expression, where Sp1 acts as a repressor. In this case, its inactivation by NO leads to an enhanced expression of TNF-{alpha} [95].

It is important to consider the general availability of Zn2+, as this is also affected by nitrosylating agents. The main Zn2+ reservoir, metallothionein, accumulates Zn2+ by complexing it to cysteine centres, and releases it in response to Zn2+ requirements [96]. S-nitrosylation of metallothionein thiolates releases Zn2+ [97–99]. Thus, importance should be given to the relative sensitivities of different protein thiolates to S-nitrosylation and Zn2+ binding, because the function of Zn2+ binding domains could be altered in different ways in situations where the nitrosylating reagents are limiting.

5.5. Matrix metalloproteinases (MMP)
Matrix metalloproteinases are endopeptidases that are able to degrade extracellular matrix components. In the vascular wall, they are implicated in many physiological and pathological processes that require matrix remodelling, such as arteriosclerosis, angiogenesis, formation of aneurysms, wound healing, and tumour invasion. Acting through cGMP-dependent activation of signalling cascades, NO may have opposite effects on the expression of different MMPs. For example, induction has been reported for MMP-13 in endothelial cells [100,101], whereas abrogation of MMP-9 was reported in smooth muscle cells [102–104].

Direct activation of MMP-9 activity by S-nitrosylation of the so-called "cysteine switch" has been reported [105]. The catalytic activity of MMPs relies on a Zn2+ atom coordinated to three histidine residues in the MMP catalytic domain. MMPs are secreted in an inactive form as proproteins. These contain a sequence, common to many family members, which includes a cysteine that coordinates to the catalytic Zn2+(the "cysteine switch"), rendering it inactive. Generally, activation of the protein is achieved by proteolytic cleavage of the propeptide by another molecule of the same or a different MMP. S-nitrosylation of the cysteine switch in MMP-9 allows it to dissociate from the metal ion and be active in the uncleaved form [105]. Since other MMPs share the same inhibitory motif, the possibility exists that this might be a general mechanism among MMPs.

5.6. Ryanodine receptor
The ryanodine receptor is a Ca2+ channel that is highly expressed in the sarcoplasmic reticulum of skeletal and cardiac muscle. Initial investigations into the influence of NO were contradictory, with reports showing that NO could both inhibit or activate the channel in both muscle types [106–108]. The first description of S-nitrosylation of the channel reported that many cysteines are S-nitrosylated (there are about 100 cysteines in each monomer of the functional tetramer, with half of them remaining in the free thiol state), and that this affects the channel activity [109]. Other researchers found that the effect of NO varies depending on several parameters, including the type of NO donor used and even when different S-nitrosothiols were employed [110,111]. This was supported by subsequent findings that different thiols in the protein react in different ways, but that specific nitrosylation occurs at just one cysteine, under the influence of the O2 concentration [35,112–114]. Recently, the different reactivity of the thiols has been also highlighted when it was shown that S-nitrosoglutathione can exert different functions by promoting S-glutathionylation of cysteine residues different from the one that is S-nitrosylated [115].


   6. Conclusions and perspectives
Top
 Abstract
 1. Introduction
 2. The chemical basis...
 3. Specificity and spatio...
 4. Current methodologies for...
 5. Regulation of protein...
 6. Conclusions and perspectives
Acknowledgments
 References
 
S-nitrosylation is a biochemical modification that potentially plays an important, and conceptually novel, regulatory role in signal transduction. The increasing evidence of functional changes resulting from this modification, and the growing number of proteins shown to be S-nitrosylated in vitro support this contention, and confirm this as an attractive area of research. Detailed studies addressing S-nitrosylation in vivo by endogenous NO and RNS are not abundant, but those that are available pave the way for future developments. Complementary approaches combining proteomic studies with the physiological generation of NO, together with studies to unravel the routes of enzymatic regulation will become increasingly feasible as improved methodologies enter the field. Coming years are likely to see the establishment of S-nitrosylation as a new paradigm in the regulation of protein function.


   Acknowledgments
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 Abstract
 1. Introduction
 2. The chemical basis...
 3. Specificity and spatio...
 4. Current methodologies for...
 5. Regulation of protein...
 6. Conclusions and perspectives
 Acknowledgments
References
 
Research in our laboratory is supported by grants from the Plan Nacional de I+D+I, Ministerio de Ciencia y Tecnología (SAF 2000-0149 and SAF 2003-01039), Comunidad Autónoma de Madrid (08.4/0030.1/2003) and Ministerio de Sanidad (RECAVA network), all in Spain. A.M.-R. has received fellowships from the Fundación Ferrer Investigación and the Comunidad Autónoma de Madrid (co-financed by the European Social Fund).


   Notes
 
Time for primary review 23 days


   References
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 Abstract
 1. Introduction
 2. The chemical basis...
 3. Specificity and spatio...
 4. Current methodologies for...
 5. Regulation of protein...
 6. Conclusions and perspectives
 Acknowledgments
 References
 

  1. Stone J.R., Marletta M.A. Soluble guanylate cyclase from bovine lung: activation with nitric oxide and carbon monoxide and spectral characterization of the ferrous and ferric states. Biochemistry (1994) 33:5636–5640.[CrossRef][ISI][Medline]
  2. Wink D.A., Osawa Y., Darbyshire J.F., et al. Inhibition of cytochromes P450 by nitric oxide and a nitric oxide-releasing agent. Arch. Biochem. Biophys. (1993) 300:115–123.[CrossRef][ISI][Medline]
  3. Gow A.J., Stamler J.S. Reactions between nitric oxide and haemoglobin under physiological conditions. Nature (1998) 391:169–173.[CrossRef][Medline]
  4. Cooper C.E. Nitric oxide and iron proteins. Biochim. Biophys. Acta (1999) 1411:290–309.[Medline]
  5. Brown G.C. Regulation of mitochondrial respiration by nitric oxide inhibition of cytochrome c oxidase. Biochim. Biophys. Acta (2001) 1504:46–57.[Medline]
  6. Moncada S., Erusalimsky J.D. Does nitric oxide modulate mitochondrial energy generation and apoptosis? Nat. Rev. Mol. Cell Biol. (2002) 3:214–220.[CrossRef][ISI][Medline]
  7. Simon D.I., Mullins M.E., Jia L., et al. Polynitrosylated proteins: characterization, bioactivity, and functional consequences. Proc. Natl. Acad. Sci. U. S. A. (1996) 93:4736–4741.[Abstract/Free Full Text]
  8. Zhang Y.-Y., Xu A.-M., Nomen M., et al. Nitrosation of tryptophan residue(s) in serum albumin and model dipeptides. Biochemical characterization and bioactivity. J. Biol. Chem. (1996) 271:14271–14279.[Abstract/Free Full Text]
  9. Espey M.G., Miranda K.M., Thomas D.D., et al. A chemical perspective on the interplay between NO, reactive oxygen species, and reactive nitrogen oxide species. Ann. N.Y. Acad. Sci. (2002) 962:195–206.[Abstract/Free Full Text]
  10. Myers P.R., Minor R.L. Jr., Guerra R. Jr., Bates J.N., Harrison D.G. Vasorelaxant properties of the endothelium-derived relaxing factor more closely resemble S-nitrosocysteine than nitric oxide. Nature (1990) 345:161–163.[CrossRef][Medline]
  11. Stamler J.S., Simon D.I., Osborne J.A., et al. S-nitrosylation of proteins with nitric oxide: synthesis and characterization of biologically active compounds. Proc. Natl. Acad. Sci. U. S. A. (1992) 89:444–448.[Abstract/Free Full Text]
  12. Turko I.V., Murad F. Protein nitration in cardiovascular diseases. Pharmacol. Rev. (2002) 54:619–634.[Abstract/Free Full Text]
  13. Ischiropoulos H. Biological selectivity and functional aspects of protein tyrosine nitration. Biochem. Biophys. Res. Commun. (2003) 305:776–783.[CrossRef][ISI][Medline]
  14. Pryor W.A., Church D.F., Govindan C.K., Crank G. Oxidation of thiols by nitric oxide and nitrogen dioxide: synthetic utility and toxicological implications. J. Org. Chem. (1982) 47:156–159.[CrossRef][ISI]
  15. Wink D.A., Nims R.W., Darbyshire J.F., et al. Reaction kinetics for nitrosation of cysteine and glutathione in aerobic nitric oxide solutions at neutral pH. Insights into the fate and physiological effects of intermediates generated in the NO/O2 reaction. Chem. Res. Toxicol. (1994) 7:519–525.[CrossRef][ISI][Medline]
  16. Gow A.J., Buerk D.G., Ischiropoulos H. A novel reaction mechanism for the formation of S-nitrosothiol in vivo. J. Biol. Chem. (1997) 272:2841–2845.[Abstract/Free Full Text]
  17. Wink D.A., Darbyshire J.F., Nims R.W., Saavedra J.E., Ford P.C. Reactions of the bioregulatory agent nitric oxide in oxygenated aqueous media: determination of the kinetics for oxidation and nitrosation by intermediates generated in the NO/O2 reaction. Chem. Res. Toxicol. (1993) 6:23–27.[CrossRef][ISI][Medline]
  18. Hogg N. The biochemistry and physiology of S-nitrosothiols. Annu. Rev. Pharmacol. Toxicol. (2002) 42:585–600.[CrossRef][ISI][Medline]
  19. Liu X., Miller M.J., Joshi M.S., Thomas D.D., Lancaster J.R. Jr. Accelerated reaction of nitric oxide with O2 within the hydrophobic interior of biological membranes. Proc. Natl. Acad. Sci. U. S. A. (1998) 95:2175–2179.[Abstract/Free Full Text]
  20. Nedospasov A., Rafikov R., Beda N., Nudler E. An autocatalytic mechanism of protein nitrosylation. Proc. Natl. Acad. Sci. U. S. A. (2000) 97:13543–13548.[Abstract/Free Full Text]
  21. Arnelle D.R., Stamler J.S. NO+, NO, and NO– donation by S-nitrosothiols: implications for regulation of physiological functions by S-nitrosylation and acceleration of disulfide formation. Arch. Biochem. Biophys. (1995) 318:279–285.[CrossRef][ISI][Medline]
  22. Hogg N. The kinetics of S-transnitrosation—a reversible second-order reaction. Anal. Biochem. (1999) 272:257–262.[CrossRef][ISI][Medline]
  23. Liu Z., Rudd M.A., Freedman J.E., Loscalzo J. S-transnitrosation reactions are involved in the metabolic fate and biological actions of nitric oxide. J. Pharmacol. Exp. Ther. (1998) 284:526–534.[Abstract/Free Full Text]
  24. Singh R.J., Hogg N., Joseph J., Kalyanaraman B. Mechanism of nitric oxide release from S-nitrosothiols. J. Biol. Chem. (1996) 271:18596–18603.[Abstract/Free Full Text]
  25. Stamler J.S., Toone E.J., Lipton S.A., Sucher N.J. (S)NO signals: translocation, regulation, and a consensus motif. Neuron (1997) 18:691–696.[CrossRef][ISI][Medline]
  26. Ascenzi P., Colasanti M., Persichini T., et al. Re-evaluation of amino acid sequence and structural consensus rules for cysteine-nitric oxide reactivity. Biol. Chem. (2000) 381:623–627.[CrossRef][ISI][Medline]
  27. Pérez-Mato I., Castro C., Ruiz F.A., Corrales F.J., Mato J.M. Methionine adenosyltransferase S-nitrosylation is regulated by the basic and acidic amino acids surrounding the target thiol. J. Biol. Chem. (1999) 274:17075–17079.[Abstract/Free Full Text]
  28. Williams J.G., Pappu K., Campbell S.L. Structural and biochemical studies of p21Ras S-nitrosylation and nitric oxide-mediated guanine nucleotide exchange. Proc. Natl. Acad. Sci. U. S. A. (2003) 100:6376–6381.[Abstract/Free Full Text]
  29. Klatt P., Pineda Molina E., Lamas S. Nitric oxide inhibits c-Jun DNA binding by specifically targeted S-glutathionylation. J. Biol. Chem. (1999) 274:15857–15864.[Abstract/Free Full Text]
  30. Klatt P., Pineda Molina E., Pérez-Sala D., Lamas S. Novel application of S-nitrosoglutathione–Sepharose to identify proteins that are potential targets for S-nitrosoglutathione-induced mixed-disulphide formation. Biochem. J. (2000) 349:567–578.[CrossRef][ISI][Medline]
  31. Klatt P., Lamas S. Regulation of protein function by S-glutathiolation in response to oxidative and nitrosative stress. Eur. J. Biochem. (2000) 267:4928–4944.[ISI][Medline]
  32. Stamler J.S., Lamas S., Fang F.C. Nitrosylation: the prototypic redox-based signaling mechanism. Cell (2001) 106:675–683.[CrossRef][ISI][Medline]
  33. Mannick J.B., Schonhoff C.M. Nitrosylation: the next phosphorylation? Arch. Biochem. Biophys. (2002) 408:1–6.[CrossRef][ISI][Medline]
  34. Lander H.M., Ogiste J.S., Pearce S.F.A., Levi R., Novogrodsky A. Nitric oxide-stimulated guanine nucleotide exchange on p21ras. J. Biol. Chem. (1995) 270:7017–7020.[Abstract/Free Full Text]
  35. Sun J., Xin C., Eu J.P., Stamler J.S., Meissner G. Cysteine-3635 is responsible for skeletal muscle ryanodine receptor modulation by NO. Proc. Natl. Acad. Sci. U. S. A. (2001) 98:11158–11162.[Abstract/Free Full Text]
  36. Nathan C. Specificity of a third kind: reactive oxygen and nitrogen intermediates in cell signaling. J. Clin. Invest. (2003) 111:769–778.[CrossRef][ISI][Medline]
  37. Schmidt H.H.H.W., Hofmann H., Schindler U., et al. No NO from NO synthase. Proc. Natl. Acad. Sci. U. S. A. (1996) 93:14492–14497.[Abstract/Free Full Text]
  38. Stuehr D., Pou S., Rosen G.M. Oxygen reduction by nitric-oxide synthases. J. Biol. Chem. (2001) 276:14533–14536.[Free Full Text]
  39. Gaston B. Nitric oxide and thiol groups. Biochim. Biophys. Acta (1999) 1411:323–333.[Medline]
  40. Bredt D.S. Nitric oxide signaling specificity—the heart of the problem. J. Cell Sci. (2003) 116:9–15.[Abstract/Free Full Text]
  41. Ziolo M.T., Bers D.M. The real estate of NOS signaling: location, location, location. Circ. Res. (2003) 92:1279–1281.[Free Full Text]
  42. Matsumoto A., Comatas K.E., Liu L., Stamler J.S. Screening for nitric oxide-dependent protein–protein interactions. Science (2003) 301:657–661.[Abstract/Free Full Text]
  43. Nathan C., Shiloh M.U. Reactive oxygen and nitrogen intermediates in the relationship between mammalian hosts and microbial pathogens. Proc. Natl. Acad. Sci. U. S. A. (2000) 97:8841–8848.[Abstract/Free Full Text]
  44. Vanin A.F., Malenkova I.V., Serezhenkov V.A. Iron catalyzes both decomposition and synthesis of S-nitrosothiols: optical and electron paramagnetic resonance studies. Nitric Oxide (1997) 1:191–203.[CrossRef][ISI][Medline]
  45. Mannick J.B., Hausladen A., Liu L., et al. Fas-induced caspase denitrosylation. Science (1999) 284:651–654.[Abstract/Free Full Text]
  46. Hoffmann J., Haendeler J., Zeiher A.M., Dimmeler S. TNF{alpha} and oxLDL reduce protein S-nitrosylation in endothelial cells. J. Biol. Chem. (2001) 276:41383–41387.[Abstract/Free Full Text]
  47. Mirza U.A., Chait B.T., Lander H.M. Monitoring reactions of nitric oxide with peptides and proteins by electrospray ionization–mass spectrometry. J. Biol. Chem. (1995) 270:17185–17188.[Abstract/Free Full Text]
  48. Lander H.M., Hajjar D.P., Hempstead B.L., et al. A molecular redox switch on p21ras. Structural basis for the nitric oxide-p21ras interaction. J. Biol. Chem. (1997) 272:4323–4326.[Abstract/Free Full Text]
  49. Zech B., Wilm M., van Eldik R., Brüne B. Mass spectrometric analysis of nitric oxide-modified Caspase-3. J. Biol. Chem. (1999) 274:20931–20936.[Abstract/Free Full Text]
  50. Kaneko R., Wada Y. Decomposition of protein nitrosothiolsin matrix-assisted laser desorption/ionization and electrospray ionization mass spectrometry. J. Mass Spectrom. (2003) 38:526–530.[CrossRef][ISI][Medline]
  51. Welch G.N., Upchurch G.R. Jr., Loscalzo J. S-nitrosothiol detection. Methods Enzymol. (1996) 268:293–298.[ISI][Medline]
  52. Wang K., Hou Y., Zhang W., et al. 15N NMR and electronic properties of S-nitrosothiols. Bioorg. Med. Chem. Lett. (1999) 9:2897–2902.[CrossRef][Medline]
  53. Fang K., Ragsdale N.V., Carey R.M., MacDonald T., Gaston B. Reductive assays for S-nitrosothiols: implications for measurements in biological systems. Biochem. Biophys. Res. Commun. (1998) 252:535–540.[CrossRef][ISI][Medline]
  54. Yang B.K., Vivas E.X., Reiter C.D., Gladwin M.T. Methodologies for the sensitive and specific measurement of S-nitrosothiols, iron-nitrosyls, and nitrite in biological samples. Free Radic. Res. (2003) 37:1–10.[ISI][Medline]
  55. Wink D.A., Kim S., Coffin D., et al. Detection of S-nitrosothiols by fluorometric and colorimetric methods. Methods Enzymol. (1999) 301:201–211.[CrossRef][ISI][Medline]
  56. Kostka P., Park J.K. Fluorometric detection of S-nitrosothiols. Methods Enzymol. (1999) 301:227–235.[ISI][Medline]
  57. Mannick J.B., Schonhoff C., Papeta N., et al. S-nitrosylation of mitochondrial caspases. J. Cell Biol. (2001) 154:1111–1116.[Abstract/Free Full Text]
  58. Schonhoff C.M., Gaston B., Mannick J.B. Nitrosylation of cytochrome c during apoptosis. J. Biol. Chem. (2003) 278:18265–18270.[Abstract/Free Full Text]
  59. Matsushita K., Morrell C.N., Cambien B., et al. Nitric oxide regulates exocytosis by S-nitrosylation of N-ethylmaleimide-sensitive factor. Cell (2003) 115:139–150.[CrossRef][ISI][Medline]
  60. Jaffrey S.R., Erdjument-Bromage H., Ferris C.D., Tempst P., Snyder S.H. Protein S-nitrosylation: a physiological signal for neuronal nitric oxide. Nat. Cell Biol. (2001) 3:193–197.[CrossRef][ISI][Medline]
  61. Kuncewicz T., Sheta E.A., Goldknopf I.L., Kone B.C. Proteomic analysis of S-nitrosylated proteins in mesangial cells. Mol. Cell Proteomics (2003) 2:156–163.[Abstract/Free Full Text]
  62. Martínez-Ruiz A., Lamas S. Detection and proteomic identification of S-nitrosylated proteins in endothelial cells. Arch. Biochem. Biophys. (2004) 423:193–200.
  63. Stamler JS, Lamas S, Fang FC. Cell 2001: http://www.cell.com/cgi/content/full/106/106/675/DC101/.
  64. Mott H.R., Carpenter J.W., Campbell S.L. Structural and functional analysis of a mutant Ras protein that is insensitive to nitric oxide activation. Biochemistry (1997) 36:3640–3644.[CrossRef][ISI][Medline]
  65. Janssen-Heininger Y.M., Poynter M.E., Baeuerle P.A. Recent advances towards understanding redox mechanisms in the activation of nuclear factor kappaB. Free Radic. Biol. Med. (2000) 28:1317–1327.[CrossRef][ISI][Medline]
  66. Matthews J.R., Wakasugi N., Virelizier J.L., Yodoi J., Hay R.T. Thioredoxin regulates the DNA binding activity of NF-kappa B by reduction of a disulphide bond involving cysteine 62. Nucleic Acids Res. (1992) 20:3821–3830.[Abstract/Free Full Text]
  67. Hayashi T., Ueno Y., Okamoto T. Oxidoreductive regulation of nuclear factor kappa B. Involvement of a cellular reducing catalyst thioredoxin. J. Biol. Chem. (1993) 268:11380–11388.[Abstract/Free Full Text]
  68. Pineda-Molina E., Klatt P., Vázquez J., et al. Glutathionylation of the p50 subunit of NF-kappaB: a mechanism for redox-induced inhibition of DNA binding. Biochemistry (2001) 40:14134–14142.[CrossRef][ISI][Medline]
  69. Matthews J.R., Botting C.H., Panico M., Morris H.R., Hay R.T. Inhibition of NF-kappaB DNA binding by nitric oxide. Nucleic Acids Res. (1996) 24:2236–2242.[Abstract/Free Full Text]
  70. DelaTorre A., Schroeder R.A., Kuo P.C. Alteration of NF-kappa B p50 DNA binding kinetics by S-nitrosylation. Biochem. Biophys. Res. Commun. (1997) 238:703–706.[CrossRef][ISI][Medline]
  71. DelaTorre A., Schroeder R.A., Bartlett S.T., Kuo P.C. Differential effects of nitric oxide-mediated S-nitrosylation on p50 and c-jun DNA binding. Surgery (1998) 124:137–142.[ISI][Medline]
  72. Bogdan C. Nitric oxide and the regulation of gene expression. Trends Cell Biol. (2001) 11:66–75.[CrossRef][ISI][Medline]
  73. Marshall H.E., Stamler J.S. Inhibition of NF-kappa B by S-nitrosylation. Biochemistry (2001) 40:1688–1693.[CrossRef][ISI][Medline]
  74. DelaTorre A., Schroeder R.A., Punzalan C., Kuo P.C. Endotoxin-mediated S-nitrosylation of p50 alters NF-kappaB-dependent gene transcription in ANA-1 murine macrophages. J. Immunol. (1999) 162:4101–4108.[Abstract/Free Full Text]
  75. Park S.K., Lin H.L., Murphy S. Nitric oxide regulates nitric oxide synthase-2 gene expression by inhibiting NF-kappaB binding to DNA. Biochem. J. (1997) 322:609–613.[ISI][Medline]
  76. Marshall H.E., Stamler J.S. Nitrosative stress-induced apoptosis through inhibition of NF-kappaB. J. Biol. Chem. (2002) 277:34223–34228.[Abstract/Free Full Text]
  77. Semenza G.L. HIF-1: mediator of physiological and pathophysiological responses to hypoxia. J. Appl. Physiol. (2000) 88:1474–1480.[Abstract/Free Full Text]
  78. Liu Y., Christou H., Morita T., et al. Carbon monoxide and nitric oxide suppress the hypoxic induction of vascular endothelial growth factor gene via the 5' enhancer. J. Biol. Chem. (1998) 273:15257–15262.[Abstract/Free Full Text]
  79. Sogawa K., Numayama-Tsuruta K., Ema M., et al. Inhibition of hypoxia-inducible factor 1 activity by nitric oxide donors in hypoxia. Proc. Natl. Acad. Sci. U. S. A. (1998) 95:7368–7373.[Abstract/Free Full Text]
  80. Kimura H., Weisz A., Kurashima Y., et al. Hypoxia response element of the human vascular endothelial growth factor gene mediates transcriptional regulation by nitric oxide: control of hypoxia-inducible factor-1 activity by nitric oxide. Blood (2000) 95:189–197.[Abstract/Free Full Text]
  81. Palmer L.A., Gaston B., Johns R.A. Normoxic stabilization of hypoxia-inducible factor-1 expression and activity: redox-dependent effect of nitrogen oxides. Mol. Pharmacol. (2000) 58:1197–1203.[ISI][Medline]
  82. Wang F., Sekine H., Kikuchi Y., et al. HIF-1{alpha}-prolyl hydroxylase: molecular target of nitric oxide in the hypoxic signal transduction pathway. Biochem. Biophys. Res. Commun. (2002) 295:657–662.[CrossRef][ISI][Medline]
  83. Metzen E., Zhou J., Jelkmann W., Fandrey J., Brüne B. Nitric oxide impairs normoxic degradation of HIF-1{alpha} by Inhibition of prolyl hydroxylases. Mol. Biol. Cell (2003) 14:3470–3481.[Abstract/Free Full Text]
  84. Brüne B., Zhou J. The role of nitric oxide (NO) in stability regulation of hypoxia inducible factor-1a (HIF-1{alpha}). Curr. Med. Chem. (2003) 10:845–855.[CrossRef][ISI][Medline]
  85. Hagen T., Taylor C.T., Lam F., Moncada S. Redistribution of intracellular oxygen in hypoxia by nitric oxide: effect on HIF1{alpha}. Science (2003) 302:1975–1978.[Abstract/Free Full Text]
  86. Mateo J., García-Lecea M., Cadenas S., Hernández C., Moncada S. Regulation of hypoxia-inducible factor-1alpha by nitric oxide through mitochondria-dependent and -independent pathways. Biochem. J. (2003) 376:537–544.[CrossRef][ISI][Medline]
  87. Lando D., Peet D.J., Whelan D.A., Gorman J.J., Whitelaw M.L. Asparagine hydroxylation of the HIF transactivation domain: a hypoxic switch. Science (2002) 295:858–861.[Abstract/Free Full Text]
  88. Sumbayev V.V., Budde A., Zhou J., Brüne B. HIF-1{alpha} protein as a target for S-nitrosation. FEBS Lett. (2003) 535:106–112.[CrossRef][ISI][Medline]
  89. Yasinska I.M., Sumbayev V.V. S-nitrosation of Cys-800 of HIF-1{alpha} protein activates its interaction with p300 and stimulates its transcriptional activity. FEBS Lett. (2003) 549:105–109.[CrossRef][ISI][Medline]
  90. Lando D., Pongratz I., Poellinger L., Whitelaw M.L. A redox mechanism controls differential DNA binding activities of hypoxia-inducible factor (HIF) 1{alpha} and the HIF-like factor. J. Biol. Chem. (2000) 275:4618–4627.[Abstract/Free Full Text]
  91. Kröncke K.-D. Zinc finger proteins as molecular targets for nitric oxide-mediated gene regulation. Antioxid. Redox Signal (2001) 3:565–575.[CrossRef][ISI][Medline]
  92. Berendji D., Kolb-Bachofen V., Zipfel P.F., et al. Zinc finger transcription factors as molecular targets for nitric oxide-mediated immunosuppression: inhibition of IL-2 gene expression in murine lymphocytes. Mol. Med. (1999) 5:721–730.[ISI][Medline]
  93. Kröncke K.D., Carlberg C. Inactivation of zinc finger transcription factors provides a mechanism for a gene regulatory role of nitric oxide. FASEB J. (2000) 14:166–173.[Abstract/Free Full Text]
  94. Galigniana M.D., Piwien-Pilipuk G., Assreuy J. Inhibition of glucocorticoid receptor binding by nitric oxide. Mol. Pharmacol. (1999) 55:317–323.[Abstract/Free Full Text]
  95. Wang S., Wang W., Wesley R.A., Danner R.L. A Sp1 binding site of the tumor necrosis factor alpha promoter functions as a nitric oxide response element. J. Biol. Chem. (1999) 274:33190–33193.[Abstract/Free Full Text]
  96. Maret W. The function of zinc metallothionein: a link between cellular zinc and redox state. J. Nutr. (2000) 130:1455S–1458S.[Abstract/Free Full Text]
  97. Berendji D., Kolb-Bachofen V., Meyer K.L., et al. Nitric oxide mediates intracytoplasmic and intranuclear zinc release. FEBS Lett. (1997) 405:37–41.[CrossRef][ISI][Medline]
  98. Pearce L.L., Gandley R.E., Han W., et al. Role of metallothionein in nitric oxide signaling as revealed by a green fluorescent fusion protein. Proc. Natl. Acad. Sci. U. S. A. (2000) 97:477–482.[Abstract/Free Full Text]
  99. Chen Y., Irie Y., Keung W.M., Maret W. S-nitrosothiols react preferentially with zinc thiolate clusters of metallothionein III through transnitrosation. Biochemistry (2002) 41:8360–8367.[CrossRef][ISI][Medline]
  100. Zaragoza C., Soria E., López E., et al. Activation of the mitogen activated protein kinase extracellular signal-regulated kinase 1 and 2 by the nitric oxide-cGMP-cGMP-dependent protein kinase axis regulates the expression of matrix metalloproteinase 13 in vascular endothelial cells. Mol. Pharmacol. (2002) 62:927–935.[Abstract/Free Full Text]
  101. Zaragoza C., Balbín M., López-Otín C., Lamas S. Nitric oxide regulates matrix metalloprotease-13 expression and activity in endothelium. Kidney Int. (2002) 61:804–808.[CrossRef]