Introduction to serial reviews: reactive nitrogen species, tyrosine nitration and cell signaling 1,2 1This article is part of a series of reviews on “Reactive Nitrogen Species, Tyrosine Nitration and Cell Signaling.” The full list of papers may be found on the homepage of the journal. 2Guest Editor: Harry Ischiropoulos

Peroxynitrite is a reactive nitrogen species produced when nitric oxide and superoxide react. In vivo studies suggest that reactive oxygen species and, perhaps, peroxynitrite can influence Na-K-ATPase function. However, the direct effects of peroxynitrite on Na-K-ATPase function remain unknown. We show that a single bolus addition of peroxynitrite inhibited purified renal Na-K-ATPase activity, with IC50 of 107 ± 9 μM. To mimic cellular/physiological production of peroxynitrite, a syringe pump was used to slowly release (∼0.85 μM/s) peroxynitrite. The inhibition of Na-K-ATPase activity induced by this treatment was similar to that induced by a single bolus addition of equal cumulative concentration. Peroxynitrite produced 3-nitrotyrosine residues on the α, β, and FXYD subunits of the Na pump. Interestingly, the flavonoid epicatechin, which prevented tyrosine nitration, was unable to blunt peroxynitrite-induced ATPase inhibition, suggesting that tyrosine nitration is not required for inhibition. Peroxynitrite led to a decrease in iodoacetamidofluorescein labeling, implying that cysteine modifications were induced. Glutathione was unable to reverse ATPase inhibition. The presence of Na+ and low MgATP during peroxynitrite treatment increased the IC50 to 145 ± 10 μM, while the presence of K+ and low MgATP increased the IC50 to 255 ± 13 μM. This result suggests that the EPNa conformation of the pump is slightly more sensitive to peroxynitrite than the E(K) conformation. Taken together, these results show that peroxynitrite is a potent inhibitor of Na-K-ATPase activity and that peroxynitrite can induce amino acid modifications to the pump.

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Inhibition of glutathione-s-transferases via tyrosine nitration by reactive nitrogen species

Free Radical Biology and Medicine ◽

10.1016/s0891-5849(98)90219-2 ◽

1998 ◽

Vol 25 ◽

pp. S69

Keyword(s):

Reactive Nitrogen Species ◽

Reactive Nitrogen ◽

Tyrosine Nitration ◽

Nitrogen Species ◽

Glutathione S Transferases

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Evidence for peroxynitrite as a signaling molecule in flow-dependent activation of c-Jun NH2-terminal kinase

AJP Heart and Circulatory Physiology ◽

10.1152/ajpheart.1999.277.4.h1647 ◽

1999 ◽

Vol 277 (4) ◽

pp. H1647-H1653 ◽

Cited By ~ 30

Author(s):

Young-Mi Go ◽

Rakesh P. Patel ◽

Matthew C. Maland ◽

Heonyong Park ◽

Joseph S. Beckman ◽

...

Keyword(s):

Endothelial Cells ◽

Shear Stress ◽

Protein Kinase ◽

Reactive Nitrogen Species ◽

Mitogen Activated Protein Kinase ◽

Reactive Nitrogen ◽

Tyrosine Nitration ◽

Nitrogen Species ◽

Laminar Shear Stress ◽

As Stress

The c-Jun NH2-terminal kinase (JNK), also known as stress-activated protein kinase, is a mitogen-activated protein kinase that determines cell survival in response to environmental stress. Activation of JNK involves redox-sensitive mechanisms and physiological stimuli such as shear stress, the dragging force generated by blood flow over the endothelium. Laminar shear stress has antiatherogenic properties and controls structure and function of endothelial cells by mechanisms including production of nitric oxide (NO) and superoxide ([Formula: see text]). Here we show that both NO and [Formula: see text] are required for activation of JNK by shear stress in endothelial cells. The present study also demonstrates that exposure of endothelial cells to shear stress increases tyrosine nitration, a marker of reactive nitrogen species formation. Furthermore, inhibitors or scavengers of NO, [Formula: see text], or reactive nitrogen species prevented shear-dependent increase in tyrosine nitration and activation of JNK. Peroxynitrite alone, added to cells as a bolus or generated over 60 min by 3-morpholinosydnonimine, also activates JNK. These results suggest that reactive nitrogen species, in this case most likely peroxynitrite, act as signaling molecules in the mechanoactivation of JNK.

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