The liberation of zinc from intracellular stores during pathophysiological conditions, especially those in which reactive oxygen and nitrogen species have been implicated, is central to the progression of neuronal injury. In addition, cellular efflux of a second ionic species, potassium, is similarly responsible for facilitating apoptotic cell death downstream of the initial oxidant-induced zinc liberation. Potassium efflux is mediated by Kv2.1-encoded potassium channels in apoptotic neurons. Here, I have characterized several critical molecular components that link zinc liberation to the loss of cytoplasmic potassium following oxidative injury. First, electrophysiological and viability studies using Kv2.1 channel mutants identified a p38 phosphorylation site at serine 800 (S800) that is required for Kv2.1 membrane insertion, potassium current enhancement, and cell death. In addition, a phospho-specific antibody for S800 detected a p38-dependent increase in Kv2.1 phosphorylation in apoptotic neurons, and reveals phosphorylation of S800 in immunopurified channels incubated with active p38. Next, I present data indicating that an N-terminal tyrosine of Kv2.1 (Y124), which is targeted by src, is also critical for the apoptotic current surge. These latter studies suggest that Y124 works in concert with the C-terminal serine (S800) target of p38 MAPK to regulate Kv2.1-mediated current enhancement. While zinc was previously shown to activate p38 and src, I demonstrate here that this metal inhibits cytoplasmic protein tyrosine phosphatase å (Cyt-PTPå), which targets Y124 and antagonizes the actions of src. Therefore, I have identified two requisite phosphorylation sites on Kv2.1, at Y124 and S800, that cooperatively mediate apoptotic potassium current enhancement. Importantly, disruption of either phosphorylation event is neuroprotective. The work presented here provides a more complete understanding of neuronal apoptotic processes by revealing the intracellular signaling events linking intracellular zinc liberation and cytoplasmic potassium efflux.
| Identifer | oai:union.ndltd.org:PITT/oai:PITTETD:etd-11112008-162403 |
| Date | 21 November 2008 |
| Creators | Redman, Patrick Timothy |
| Contributors | Elias Aizenman, Ph.D., Donald B. DeFranco, Ph.D., Stephen D. Meriney, Ph.D., Edwin S. Levitan, Ph.D., Michael M. Tamkun, Ph.D, Teresa G. Hastings, Ph.D. |
| Publisher | University of Pittsburgh |
| Source Sets | University of Pittsburgh |
| Language | English |
| Detected Language | English |
| Type | text |
| Format | application/pdf |
| Source | http://etd.library.pitt.edu/ETD/available/etd-11112008-162403/ |
| Rights | restricted, I hereby certify that, if appropriate, I have obtained and attached hereto a written permission statement from the owner(s) of each third party copyrighted matter to be included in my thesis, dissertation, or project report, allowing distribution as specified below. I certify that the version I submitted is the same as that approved by my advisory committee. I hereby grant to University of Pittsburgh or its agents the non-exclusive license to archive and make accessible, under the conditions specified below, my thesis, dissertation, or project report in whole or in part in all forms of media, now or hereafter known. I retain all other ownership rights to the copyright of the thesis, dissertation or project report. I also retain the right to use in future works (such as articles or books) all or part of this thesis, dissertation, or project report. |
Page generated in 0.002 seconds