Abstract
Oxidative stress induced by glutathione depletion in the mouse HT22 neuroblastoma cell line and embryonic rat immature cortical neurons causes a delayed, sustained activation of extracellular signal-regulated kinase (ERK) 1/2, which is required for cell death. This sustained activation of ERK1/2 is mediated primarily by a selective inhibition of distinct ERK1/2-directed phosphatases either by enhanced degradation (i.e., for mitogen-activated protein kinase phosphatase-1) or as shown here by reductions in enzymatic activity (i.e., for protein phosphatase type 2A). The inhibition of ERK1/2 phosphatases in HT22 cells and immature neurons subjected to glutathione depletion results from oxidative stress because phosphatase activity is restored in cells treated with the antioxidant butylated hydroxyanisole. This leads to reduced ERK1/2 activation and neuroprotection. Furthermore, an increase in free intracellular zinc that accompanies glutathione-induced oxidative stress in HT22 cells and immature neurons contributes to selective inhibition of ERK1/2 phosphatase activity and cell death. Finally, ERK1/2 also functions to maintain elevated levels of zinc. Thus, the elevation of intracellular zinc within neurons subjected to oxidative stress can trigger a robust positive feedback loop operating through activated ERK1/2 that rapidly sets into motion a zinc-dependent pathway of cell death.
Footnotes
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This work was supported by National Institutes of Health grants NS38319 (to D.B.D.) and NS43277 (to E.A.)
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Y.H. and R.S. contributed equally to this work and are considered co-first authors.
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ABBREVIATIONS: ROS, reactive oxygen species; MAPK, mitogen-activated protein kinase; ERK, extracellular signal-regulated kinase; JNK, Jun NH2-terminal kinase; PI, propidium iodine; HCA, homocysteate; MEK, mitogen-activated protein kinase kinase; U0126, 1,4-diamino-2,3-dicyano-1,4-bis(methylthio)butadiene; PP2A, protein phosphatase-type 2A; IP, immunoprecipitation; PAGE, polyacrylamide gel electrophoresis; PBST, phosphate-buffered saline with Tween 20; BSA, bovine serum albumin; TPEN, N,N,N′,N′-tetrakis-(2-pyridylmethyl)-ethylenediamine; AM, acetoxymethyl ester; MRE, metal regulatory element; DSP, dual-specificity protein phosphatase; BHA, butylated hydroxyanisole; TNF, tumor necrosis factor; MKP, mitogen-activated protein kinase phosphatase; DTT, dithiothreitol; p, phosphorylated.
- Received May 21, 2008.
- Accepted July 16, 2008.
- The American Society for Pharmacology and Experimental Therapeutics
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