Review articleThe role of hypernitrosylation in the pathogenesis and pathophysiology of neuroprogressive diseases
Introduction
Nitric oxide (NO), produced in vivo by nitric oxide synthase isoforms, is a gaseous free radical signal transducer which regulates a plethora of biological processes in the cardiovascular, immune and nervous systems, and indeed in virtually every other system in the body (Lundberg et al., 2015, Mattila and Thomas, 2014, Steinert et al., 2010). NO signalling has been traditionally described as canonical and non-canonical (Martinez-Ruiz et al., 2011). Canonical signalling involves the binding of NO with soluble guanylate leading to the production of cGMP and subsequent activation of a range of downstream cGMP-dependent kinases, which in turn transduce a virtually countless number of signaling events via protein phosphorylation (Moncada et al., 1997, Murad, 2006). Extending this, there is an accumulating data indicating that the bulk of nitric oxide signalling is effected via post-transcriptional modification of protein cysteine groups in processes such as glutathionylation and nitrosylation. These processes enable redox sensing and facilitate homeostatic regulation of redox dependent protein signalling, function, stability and trafficking (Banerjee, 2012, Hill and Bhatnagar, 2012, Paulsen and Carroll, 2010, Winterbourn and Hampton, 2008).
S-nitrosylation involves the reversible covalent addition of a NO group, usually derived from higher oxides such as N2O3, to specific regulatory protein cysteine thiolate anions. A consensus motif involving acid base or tyrosine groups some 8 Å from the target cysteine and/or a highly hydrophobic local environment facilitate this process (Gould et al., 2013, Lima et al., 2010, Nakamura and Lipton, 2013, Sun and Murphy, 2010). Accumulating evidence indicates that S-nitrosylation regulates the biological activity of many proteins in a similar manner to phosphorylation and other post-translational modifications, such as palmitoylation (Chung et al., 2004, Gould et al., 2013, Stamler et al., 2001, Uehara et al., 2006).
The process and cellular levels of protein nitrosylation are regulated and counterbalanced by denitrosylation via the activity of a range of NADH- or NADPH-dependent enzymes described as denitrosylases. Examples include the thioredoxin system (Benhar et al., 2009, Forrester et al., 2009), the nitrosogluthathione reductase (GSNOR) system (Liu et al., 2001, Liu et al., 2004) and lesser players such as protein disulphide isomerase (PDI), superoxide dismutase, glutathione peroxidase, xanthine oxidase and possibly lipoic acid (Anand and Stamler, 2012, Sengupta and Holmgren, 2013, Trujillo et al., 1998). The weight of evidence suggests that transnitrosylation between cell proteins is, in practice, the most important mechanism for nitrosylation and de-nitrosylation of proteins and hence the major vehicle for NO based redox regulation of cellular signalling pathways in vivo (Benhar et al., 2008, Kornberg et al., 2010, Mitchell et al., 2007, Nakamura and Lipton, 2011a, Qu et al., 2011, Wu et al., 2010, Wu et al., 2011).
Nitrosylation of key proteins plays a pivotal role in cellular adaptation to increased levels of oxidative and nitrosative stress (O&NS) (Gorelenkova Miller and Mieyal, 2015, Okamoto and Lipton, 2015). Initially, increases in levels of reactive oxygen species (ROS) and reactive nitrogen species (RNS) lead to the protective nitrosylation of a plethora of crucial functional and regulatory proteins as a conserved defence against the irreversible oxidation of cysteine and the ensuing permanent changes in their secondary and tertiary conformation, structure and loss of function (Kohr et al., 2014, Penna et al., 2014, Sun and Murphy, 2010, Sun et al., 2006). However, further increases in O&NS stress lead to the breakdown of mechanisms governing denitrosylation and transnitrosylation and protein thiols may be irreversibly oxidized to sulfenic acid, sulfinic acid or sulfonic acid (Halloran et al., 2013, Nakamura and Lipton, 2013).
Several studies have demonstrated that thioredoxin (Trx-1) can function as a transnitrosylase or denitrosylase dependent on the redox state of specific cysteine residues. In particular nitrosylation of Cys73 enables its transnitrosylating activity (Li et al., 2013, Wu et al., 2010, Wu et al., 2011). Significantly, nitrosylation of Cys73 can only be achieved following the formation of a disulphide bridge between Cys32 and Cys35, whereupon the denitrosylation capacity of the thioredoxin system is greatly diminished or even terminated (Li et al., 2013, Wu et al., 2010, Wu et al., 2011). This change in the thioredoxin system from facilitating denitrosylation to driving transnitrosylation during an environment of increasing O&NS is sometimes described as “a redox switch“ (Wu et al., 2010, Wu et al., 2011). Given the universal presence of chronic oxidative and nitrosative stress in neurological and neuro progressive illnesses this is likely a key factor in their pathogenesis (Morris and Berk, 2015).
The pathological consequences stemming from the loss of the denitrosylating capacity of thioredoxin in conditions of excessive O&NS is exacerbated by the impairment of the S-nitrosoglutathione reductase (GSNOR) system due to GSH depletion via oxidation to GSSG by ROS and RNS coupled with increased efflux of GSH and GSSH into the extracellular environment (Circu et al., 2009, Franco and Cidlowski, 2012, Hansen et al., 2006). The antioxidant activity of pyridoxin 2 is also suppressed by the S-nitrosylation of two pivotal cysteine residues (Cys51 and Cys172) which enable its antioxidant capacity (Fang et al., 2007, Romero-Puertas et al., 2007). In this scenario, the cell is left bereft of denitrosylating and reducing power. As a consequence, following engagement of this “redox switch”, the rates of nitrosylation cumulatively exceed the rate of denitrosylation leading to the development of a state of hypernitrosylation.
Such a state of inhibited denitrosylation and transnitrosylation leads to the development of pathology in several dimensions and plays a major role in the pathogenesis of many and diverse neurodegenerative diseases (Fig. 1). These dimensions include impairment of mitochondrial function and anti-inflammatory and antioxidant defenses, which in turn result in compromised energy production and the development of autoimmune responses (Nakamura and Lipton, 2011b, Nakamura et al., 2013). Other sources of pathology stemming from hypernitrosylation include loss of function of neuroprotective proteins, compromised transcription factor activity and dysregulated lysosomal and proteasomic protein degradation pathways, coupled with chronically misfolded proteins and activated apoptotic machinery (Nakamura and Lipton, 2011a). Furthermore, persistent nitrosylation in an environment of chronic neuro-inflammation leads to compromised neural function, impaired neurogenesis and frank neurodegeneration (Okamoto et al., 2014).
While these abnormalities flowing from a state of hypernitrosylation in neurodegenerative diseases have been detailed in a thorough review by (Nakamura et al., 2013) such a range of abnormalities are also documented in illnesses as schizophrenia, bipolar disorder and major depression, which are also characterized by chronic inflammation and O&NS (Morris and Berk, 2015) and are increasingly being described as neuroprogressive (Gama et al., 2013, Gildengers et al., 2014, Maes et al., 2011). Neuroprogression is characterised by diminished neurogenesis, changed neurotransmitter systems, enhanced neuronal apoptosis and several other markers of neurodegeneration, together with elevated autoimmune activity and impaired mitochondrial function (Bakunina et al., 2015, Berk et al., 2011, Maes et al., 2015). These abnormalities are underpinned by the biological foundation of chronic O&NS, activated immune and inflammatory pathways, reduced levels of neurotrophin activity and modulation of hypothalamic-pituitary-adrenal (HPA) axis function (Bakunina et al., 2015, Berk et al., 2011, Berk et al., 2013, Moylan et al., 2013).
Section snippets
Aims of the study
This paper will focus on the regulation of S-nitrosylation as a cellular signaling system to explain the role of chronic O&NS in the development of hypernitrosylation and the potential role of abnormal transnitrosylation and inactivated denitrosylation in the pathogenesis and pathophysiology of neuroprogressive illnesses.
Methods
The electronic databases PUBMED, Google Scholar and Scopus were utilized as sources for this narrative review focusing on the keywords: nitric oxide; nitrosylation; transnitrosylation; O&NS; oxidative stress; neurogenesis; neuroprogression; HPA axis; bipolar disorder; schizophrenia and depression. These sources were subsequently consulted for mechanisms leading to abnormal nitrosylation and evidence of pathological consequences stemming from such a state both in general terms and as a specific
Impaired transnitrosylation as a source of pathology
Glyceraldehyde-3-Phosphate Dehydrogenase (GAPDH) is a multifunctional protein with well documented roles in the regulation of glycolysis, endocytosis, gene expression, DNA repair and apoptosis (Sawa et al., 1997, Sirover, 2005, Tristan et al., 2011). In an environment of increasing O&NS, a catalytic cysteine in GAPDH becomes nitrosylated by NO provoking a conformational change promoting the binding of the molecule with the E3 ubiqitin ligase Siah1 (Kornberg et al., 2010). This enzyme possesses
Hypernitrosylation and p53 activity
The transcription factor p53 has been described as “the guardian of the genome” as a result of its role in suppressing tumour formation effected by inhibiting the expression of multiple genes involved in tumourigenesis and its capacity to affect DNA repair and induce cell cycle arrest, senesence and apoptosis (Efeyan and Serrano, 2007, Rivlin et al., 2011). However, p53 family proteins also exert a number of other functions in cells including stimulating oxidative phosphorylation while
Background information
PTEN acts as a negative regulator of PI3K/AKT/mTOR signalling (Numajiri et al., 2011), which is enabled by an essential cysteine residue at the signature active site motif CX5R which regulates its phosphatase activity and suppressive properties (Cho et al., 2004, Yu et al., 2005). Hence the fact that S-nitrosylation of these catalytic thiolate groups inhibits the activity of the enzyme leading to the activation of PI3K/AKT/mTOR signalling is unsurprising (Kwak et al., 2010, Numajiri et al., 2011
Nitrosylation, NDMA receptor complex and glutamate excitotoxicity
S-nitrosylation/denitrosylation of the NMDA receptor (NMDAR) is a an important regulator of NMDAR activity under physiological conditions and a crucial defense against the development of glutamate excitotoxicity in conditions of neuropathology (Choi et al., 2001, Lei et al., 1992, Lipton et al., 1993). Briefly, in physiological conditions, NO modulates several aspects of synaptic function and plasticity by S-nitrosylation of a broad range of target proteins, resulting in significant changes in
Conclusion
There appears to be a clear role of persistent hypernitrosylation as at least a partial explanation for the inhibition of mitochondrial function and ATP synthesis seen in many patients with neuroprogressive illnesses. The presence of nitrosylated amino acids and damage to lipids, proteins and DNA functioning as inflammogens and damage-associated molecular patterns (DAMPS) respectively would go some way to explaining the chronic oxidative stress and inflammation present in many patients, as well
Conflicts of interest
The authors have no conflicts of interest to declare.
Authorships
All authors contributed equally to the writing up of the paper.
Acknowledgement
MB is supported by a NHMRC Senior Principal Research Fellowship1059660.
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