Comparing Nitrosative Versus Oxidative Stress toward Zinc Finger-dependent Transcription

Abstract

Effects of nitric oxide (NO ⅐ ) on cellular functions are complex and even appear to be contradictory, janus-headed. NO ⅐ may act cytotoxically but may also protect cells from toxic insults (1), it may compromise the cellular redox state but may also act as an antioxidant (2, 3), and it may activate or inhibit signal transduction pathways (4, 5) and gene transcription (6 -9), respectively. Many of these effects can, at least in part, be explained by different NO ⅐ concentrations achieved in the respective microenvironment. Nanomolar concentrations of NO ⅐ , as typically synthesized in a tightly regulated fashion by constitutively expressed nitric oxide synthase (cNOS), 1 serve as a signal molecule activating the soluble guanylate cyclase to produce cGMP, which acts as a second messenger. However, in addition to two cNOS there is also an inducible NOS (iNOS) expressed in a variety of acute or chronic disease states (for reviews see Refs. 10, 11). Originally described as a cytotoxic activated macrophage effector molecule (12) it is now evident that iNOS-derived NO ⅐ exerts a multitude of biological functions. In an apparently unregulated fashion iNOS synthesizes NO ⅐ for hours or even days resulting in micromolar concentrations of NO ⅐ . Under these conditions NO ⅐ may react with oxygen in a reaction mainly depending on the NO ⅐ concentration to yield higher nitrogen oxides (NO x such as N 2 O 3 , etc.), which display a much broader chemical reaction spectrum than NO ⅐ itself (13). A growing body of evidence suggests that NO ⅐ after reaction to NO x helps to orchestrate gene expression, e.g. via posttranslational modifications of transcription factors. A prevalent DNA binding motif of transcription factors is the zinc finger structure with Zn 2ϩ tetrahedrally coordinated between a ␤-hairpin and a short ␣-helix, creating a small, functional and independently folded domain (14). In these zinc fingers cysteine thiols and histidine imidazole nitrogens serve as direct ligands for the zinc ion.We previously found that NO ⅐ S-nitrosates cysteines in metallothionein, mediating the release of Zn 2ϩ from this zincstoring protein (15), induces Zn 2ϩ release within cells (16) and is able to inhibit zinc finger-dependent transcription (17-19). However, zinc fingers can easily be disrupted by cysteine oxidation, e.g. by reactive oxygen species, or by electrophilic attack, e.g. by alkylating compounds (for reviews see Refs. 20,21). The question therefore arises, whether the reaction of NO ⅐ with zinc fingers may have a special role, different from the reaction with other reactive species generated during inflammatory reactions. Such species are superoxide, its dismutation product hydrogen peroxide, the product of its reaction with NO ⅐ , peroxynitrite, or singlet oxygen, as well as peroxyl radicals.To investigate the impact of these species on the zinc finger integrity, we used the transcription factors VDR and RXR as a model system both containing two Cys 4 -type zinc fingers, which bind to specific promoter sequences as the he...

Regulation of zinc homeostasis by inducible NO synthase-derived NO: Nuclear metallothionein translocation and intranuclear Zn2+release

“…NO induces disulfide formation in MT (7), which can be reduced by cells, whereas H 2 O 2 oxidizes MT cysteines beyond the levels of disulfides (48). This result is in line with previous findings, showing that cells exposed to nitrosative in contrast to oxidative stress are able to repair disrupted zinc fingers of transcription factors after the stress has declined (49).…”

Mammalian Zinc Transport, Trafficking, and Signals

“…In vitro studies have shown that the metal thiolate clusters of MT possess the unique ability to function as a redox unit; therefore, the protein has the potential to be involved in redox-sensitive signaling pathways (56). Comparative studies with nitric oxide (NO), H 2 O 2 , singlet oxygen, peroxyl radicals, and peroxynitrite suggest only stress from NO causing S-nitrosylation of MT cysteines and Zn 2ϩ release is sufficiently mild to allow reconstitution of MT through Zn 2ϩ rebinding (57). Fluorescence resonance energy transfer experiments have confirmed that in cultured endothelial cells MT undergoes conformational changes in the presence of NO donors, e.g.…”

Biological chemistry of reactive oxygen and nitrogen and radiation-induced signal transduction mechanisms

“…However, there appears to be an important difference between the ROS and RNS mechanisms. Using the vitamin D receptor (VDR) and retinoid X receptor (RXR) as model zinc-finger transcription factors, Kroncke et al (2002) showed that the zinc-fingers of VDR and RXR could be repaired after nitrosative but not oxidative stresses. This suggests that RNSs are not just inhibitors but regulators of zinc-finger transcription factor activities.…”