Thioredoxin-interacting Protein (Txnip) Is a Feedback Regulator of S-Nitrosylation

Abstract

Nitric oxide exerts a plethora of biological effects via protein S-nitrosylation, a redox-based reaction that converts a protein Cys thiol to a S-nitrosothiol. However, although the regulation of protein S-nitrosylation has been the subject of extensive study, much less is known about the systems governing protein denitrosylation. Most recently, thioredoxin/thioredoxin reductases were shown to mediate both basal and stimulus-coupled protein denitrosylation. We now demonstrate that protein denitrosylation by thioredoxin is regulated dynamically by thioredoxin-interacting protein (Txnip), a thioredoxin inhibitor. Endogenously synthesized nitric oxide represses Txnip, thereby facilitating thioredoxin-mediated denitrosylation. Autoregulation of denitrosylation thus allows cells to survive nitrosative stress. Our findings reveal that denitrosylation of proteins is dynamically regulated, establish a physiological role for thioredoxin in protection from nitrosative stress, and suggest new approaches to manipulate cellular S-nitrosylation.It has become increasingly appreciated that protein S-nitrosylation, the covalent attachment of a nitroso group to a cysteine thiol side chain, is a principle mechanism by which nitric oxide modulates numerous cellular functions and phenotypes (1, 2). These include G-protein-coupled receptor signaling (3-5), death receptor-mediated apoptosis (6 -9), vesicular trafficking (10 -12), stimulation of prostaglandin synthesis (13-15), hypoxia-dependent control of blood flow (16 -18), and the unfolded protein response (19). In addition, aberrant S-nitrosylation is implicated in pathologies such as tumor initiation and growth (20, 21), neurodegeneration (19,22,23), and pulmonary hypertension (24).The three isoforms of nitric-oxide synthase (neuronal NOS/ NOS1, iNOS/NOS2, 2 and endothelial NOS/NOS3) are well established mediators of S-nitrosylation, and numerous studies have demonstrated that their localization is critical for S-nitrosylation of target proteins. For example, binding of iNOS to COX2 is required for S-nitrosylation and activation of prostaglandin synthesis (13), whereas the subcellular localization of endothelial NOS is a major determinant of S-nitrosylation-mediated protein trafficking (25). Moreover, although it is generally assumed that S-nitrosylation reactions are nonenzymatic, there is precedence for hemoglobin-dependent S-nitrosylation of the anion exchanger protein AE1 (26) and ceruloplasmin-dependent S-nitrosylation of glypican-1 (27) and GSH (28), consistent with a recent report that metalloproteins may play a general role in SNO synthesis (29).By contrast with progress in elucidating the enzymatic determinants of S-nitrosylation, the intracellular mediators of denitrosylation and their possible contributions to overall Snitrosylation status have only recently gained attention. By analogy to phosphorylation (where kinases and phosphatases together regulate phosphorylation), the steady-state level of S-nitrosylation is the net difference between nitrosylation and denitros...

Endogenously produced nitric oxide mitigates sensitivity of melanoma cells to cisplatin

“…S-nitrosation and denitrosation of cellular proteins are regulated through an enzymatic complex, in which thioredoxins, thioredoxin reductase, thioredoxin-interacting protein, and Snitrosoglutathione reductase orchestrate the cellular pool of SNOs and, therefore, protein function (24,41). In addition, the fact that S-nitrosation of a given protein depends on its subcellular localization, rather than solely on the amount of available NO, further emphasizes that S-nitrosation is a finely controlled mechanism (17,29,41), an imbalance of which is linked to genesis of several pathologic states (13, 16-18, 41). We conjecture that, during the response to cisplatin, a process is activated in which NO concentration increases and SNO levels are modulated by the aforementioned enzymes (together with other mechanisms yet to be identified), enabling cancer cells to mitigate apoptosis.…”

Specificity in S-Nitrosylation: A Short-Range Mechanism for NO Signaling?

“…On the other hand, Trx2 was identified as a denitrosylase of mitochondrial caspase-3 upon Fas ligand stimulation, a process required for its activation and, therefore, for apoptotic cell death (16). Furthermore, the Trx-interacting protein (Txnip) has been shown to inhibit denitrosylate activity of Trx, since it increases SNOprotein levels (43).…”

Nitric oxide vs insulin secretion, action and clearance

“…In addition, NO cGMP independently represses the transcription of thioredoxin-interacting protein (TXNIP), which not only enables antioxidative actions of thioredoxins, but also stimulates thioredoxin-dependent protein denitrosylation, thus providing a negative feedback mechanism protecting against excessive protein S -nitrosylation and nitrosative stress [10]. Shaked et al [11] demonstrated that the NO-dependent TXNIP inhibition contributed to the protective effect of insulin against glucose-induced beta cell apoptosis.…”