Nitration of Hsp90 induces cell death

Abstract

Oxidative stress is a widely recognized cause of cell death associated with neurodegeneration, inflammation, and aging. Tyrosine nitration in these conditions has been reported extensively, but whether tyrosine nitration is a marker or plays a role in the cell-death processes was unknown. Here, we show that nitration of a single tyrosine residue on a small proportion of 90-kDa heat-shock protein (Hsp90), is sufficient to induce motor neuron death by the P2X7 receptor-dependent activation of the Fas pathway. Nitrotyrosine at position 33 or 56 stimulates a toxic gain of function that turns Hsp90 into a toxic protein. Using an antibody that recognizes the nitrated Hsp90, we found immunoreactivity in motor neurons of patients with amyotrophic lateral sclerosis, in an animal model of amyotrophic lateral sclerosis, and after experimental spinal cord injury. Our findings reveal that cell death can be triggered by nitration of a single protein and highlight nitrated Hsp90 as a potential target for the development of effective therapies for a large number of pathologies.apoptosis | peroxynitrite | PC12 cells

Nitric Oxide and Heat Shock Protein 90 Activate Soluble Guanylate Cyclase by Driving Rapid Change in Its Subunit Interactions and Heme Content

“…We saw that the heme-independent sGC activator BAY 60-2770 could mimic the effect of NO in promoting hsp90 dissociation, whereas the heme-dependent sGC activator BAY 41-2272 could not. The ability of BAY 60-2770 to do so is perhaps the best indicator that the mechanism of NO action does not necessarily require any NO-based protein modifications such as protein S-nitrosation or tyrosine nitration, which can otherwise occur in hsp90 and sGC proteins when cells are exposed to NO (23)(24)(25). Rather, our results suggest a mechanism of action that involves fundamental changes in the apo-sGC-␤1 subunit.…”

“…3, B and E). In particular, there emerged a lower M r subpopulation of sGC-␤1 that clearly had no hsp90 associated with it (lanes [23][24][25][26]. This subpopulation existed only transiently in the cells because it was no longer present in cells that underwent a longer SNAP exposure for 30 min (Fig.…”

“…Gel filtration analysis of the heme-deficient RFL-6 cell supernatant (Fig. 6E) revealed that the BAY 60-2770 treatment shifted the apparent M r distribution of sGC-␤1 to cause significant buildup of the low M r , hsp90-free sGC-␤1 subpopulation in the cells (fractions [22][23][24][25]. Similar results were obtained with RFL-6 cells that had not been made heme-deficient prior to the BAY 60-2770 treatment (data not shown), consistent with their containing some apo-sGC-␤1 under normal culture conditions.…”

Site-specific Nitration of Apolipoprotein A-I at Tyrosine 166 Is Both Abundant within Human Atherosclerotic Plaque and Dysfunctional

“…NO 2 -Tyr 166 -apoA-I protein was expressed in an E. coli TOP10 strain harboring the evolved orthogonal Methanococcus jannaschii (Mj) nitro-Tyr-tRNA synthetase/tRNA CUA pair that specifically incorporates 3-nitrotyrosine into amber codons (16).…”

“…This capability has the potential to allow one to study the functional consequences of individual PTM to residues that have previously been implicated as playing a functional role based on their detection by proteomics studies in the material recovered from diseased tissue (16,17).…”