Peroxynitrite induces F-actin depolymerization and blockade of myosin ATPase stimulation

Abstract

Treatment of F-actin with the peroxynitrite-releasing agent 3-morpholinosydnonimine (SIN-1) produced a dose-dependent F-actin depolymerization. This is due to released peroxynitrite because it is not produced by 'decomposed SIN-1', and it is prevented by superoxide dismutase concentrations efficiently preventing peroxynitrite formation. F-actin depolymerization has been found to be very sensitive to peroxynitrite, as exposure to fluxes as low as 50-100 nM peroxynitrite leads to nearly 50% depolymerization in about 1 h. G-actin polymerization is also impaired by peroxynitrite although with nearly 2-fold lower sensitivity. Exposure of F-actin to submicromolar fluxes of peroxynitrite produced cysteine oxidation and also a blockade of the ability of actin to stimulate myosin ATPase activity. Our results suggest that an imbalance of the F-actin/G-actin equilibrium can account for the observed structural and functional impairment of myofibrils under the peroxynitrite-mediated oxidative stress reported for some pathophysiological conditions. Ó 2006 Elsevier Inc. All rights reserved.Keywords: F-actin; Peroxynitrite; Myosin ATPase; Actin polymerization/depolymerization; Oxidative stress; SIN-1; Cysteine oxidation Exposure of muscle cells to chronic oxidative stress conditions results in impairment of muscle contraction, which has been proposed to be, at least in part, the result of oxidative modification of myofibril proteins [1,2]. On the other hand, it has been shown that Cys 374 of actin monomer (G-actin) is particularly sensitive to oxidation by hydrogen peroxide [3] and nitric oxide donors [4], which are stress conditions leading to a reduction of the actin polymer (F-actin), that plays a major role in myofibril contraction. Peroxynitrite is an important oxidative stress agent in ischemia/reperfusion insults, such as heart infarct or atrial fibrillation [1,5,6], and inflammation [7], and it has been reported that peroxynitrite can inhibit actin polymerization in neutrophils [8]. Furthermore, in aged muscle, actin is one of the proteins showing higher content of the commonly used fingerprint marker of peroxynitrite-reactive proteins 3-nitrotyrosine [9]. During inflammation and ischemia/reperfusion episodes tissue cells are exposed to fluxes of peroxynitrite ranging from submicromolar to micromolar concentrations [7,10]. SIN-1 is being used to mimic the effects of chronic exposure of cells in culture to a peroxynitrite oxidative stress [11,12], because it slowly decomposes in neutral and weakly alkaline aqueous solutions releasing nitric oxide and superoxide anion [13], which react with each other to produce peroxynitrite with a second order rate constant of (4-7) · 10 9 M À1 s À1 close to the diffusion limit for chemical reactions [10,14]. Moreover, the kinetics of peroxynitrite generation and the peroxynitrite concentration attained in the solution during SIN-1 decomposition can be reliably monitored in the buffered solutions commonly used for biochemical studies with isolated subcellular components [15]. 0 ...

Peroxynitrite diminishes myogenic tone in cerebral arteries: role of nitrotyrosine and F-actin

“…This is in agreement with other studies that have shown that ONOO Ϫ can induce both depolymerization of F-actin and inhibition of G-actin polymerization (16,51). Since a dynamic actin cytoskeleton is a significant contributor to pressure-induced myogenic activity (13,15), production of ONOO Ϫ may have a deleterious effect on cerebrovascular resistance through disruption of VSM actin following conditions of increased oxidative stress.…”

“…ONOO Ϫ has been shown to cause depolymerization of F-actin and inhibit polymerization in vitro (4,16,51). In addition, studies in neutrophils and red blood cells showed a loss of F-actin and actin-dependent functions following exposure to ONOO Ϫ that was associated with increased NT (16,22).…”

Actin as a potential target for decavanadate

“…As described above, an explanation can emerge by the fact that actin polymerization/depolymerization is involved in vesicles trafficking, such as Glut-4, as mentioned earlier, and decavanadate interferes with this equilibrium [6]. Additionally, decavanadate is known to induce changes in reactive oxygen species production [17], and these can by themselves affect both calcium homeostasis and actin cytoskeleton dynamics, processes that are linked to each other [45]. However, besides the impact of the interaction of decavanadate with actin, perhaps the most potent cellular target of decavanadate is mitochondria, being affected by decameric vanadate concentrations as low as 0.1 μM [18,46].…”

Characterization of myosin light chain in shrimp hemocytic phagocytosis☆

“…Myosin is a hexameric protein, consisting of two heavy chains (MHC) of molecular weight about 200,000 and four light chains (MLC) of about 20,000 [18]. They are cell skeleton proteins and play a role in the interaction of actin thin filament [19]. Some conventional myosins, have been implicated to play vital roles in cell adhesion and phagocytosis, producing the driving force for phagosome formation and transport during phagocytosis [20,21].…”