Apoptotic neuronal cell death may play a role in many acute and chronic neurologic disorders. These disorders range from acute stroke, head trauma and epilepsy to more chronic states, such as Huntington's disease, Alzheimer's disease, amyotrophic lateral sclerosis, HIV-associated dementia, and glaucoma. Moreover, a contributing factor to such damage is excessive excitation of glutamate receptors, particularly (but not exclusively) the N-methyI-D-aspartate (NMDA) subtype of glutamate receptor because of its high permeability to Ca 2+ and subsequent free radical generation.
The aim of this proposed research project is to uncover the role of myocyte enhancer factor-2 (MEF2) transcription factors in this excitotoxic/apoptotic process in neurons during ischemic stroke in vivo. MEF2 transcription factors are activated by p38 mitogen-activated protein kinase during neuronal and myogenic differentiation. Recent work has shown that stimulation of this pathway is anti-apoptotic in stem cells but pro-apoptotic in mature neurons exposed to mild excitotoxic or other stresses. Here, preliminary data in vitro show that mild excitotoxic (NMDA) insults to mature cerebrocortical neurons activate caspases-3, -7, in turn cleaving MEF2A, C and D isoforms. Endogenous MEF2 cleavage fragments containing a truncated transactivation domain but preserved DNA binding domain are shown to block MEF2 transcriptional activity via dominant interference. In vitro transfection of constitutively-active/uncleavable MEF2 (MEF2-CA) rescues MEF2 transcriptional activity following NMDA insult and prevents neuronal apoptosis. Conversely, dominant-interfering MEF2 (MEF2-DN) abrogates neuroprotection by MEF2C-CA. Our underlying hypothesis is that these results obtained in vitro can now be applied in vivo using tetracycline (or doxycycline, """"""""dox"""""""")-controlled transgenic mice expressing these MEF2-CA and MEF2-DN transgenes. This grant will define a novel pathway to neuronal apoptosis in ischemia via caspase-catalyzed cleavage of MEF2.
The Specific Aims are as follows: 1. To characterize anti-apoptotic effects of MEF2-CA in stroke using dox-controlled transgenic mice. 2. To characterize the effect of caspase cleavage fragments of MEF2 as dominant interfering forms that contribute to stroke damage using dox-controlled transgenic mice that express doxycycline-controlled, MEF2 cleavage products. 3. To characterize MEF2 transcriptional activity in vivo after an hypoxic/ischemic (stroke) insult but prior to cell loss using a MEF2-indicator mouse that has been engineered to activate the LacZ gene in accord with the degree of MEF2 transcriptional activity (designated des-mef2-LacZ).

Agency
National Institute of Health (NIH)
Institute
National Institute of Neurological Disorders and Stroke (NINDS)
Type
Research Project (R01)
Project #
5R01NS044326-02
Application #
6747654
Study Section
Molecular, Cellular and Developmental Neurosciences 2 (MDCN)
Program Officer
Nunn, Michael
Project Start
2003-08-01
Project End
2007-07-31
Budget Start
2004-08-01
Budget End
2005-07-31
Support Year
2
Fiscal Year
2004
Total Cost
$456,000
Indirect Cost
Name
Sanford-Burnham Medical Research Institute
Department
Type
DUNS #
020520466
City
La Jolla
State
CA
Country
United States
Zip Code
92037
Cho, Eun-Gyung; Zaremba, Jeffrey D; McKercher, Scott R et al. (2011) MEF2C enhances dopaminergic neuron differentiation of human embryonic stem cells in a parkinsonian rat model. PLoS One 6:e24027
Li, Zhen; McKercher, Scott R; Cui, Jiankun et al. (2008) Myocyte enhancer factor 2C as a neurogenic and antiapoptotic transcription factor in murine embryonic stem cells. J Neurosci 28:6557-68
Li, Hao; Radford, Jonathan C; Ragusa, Michael J et al. (2008) Transcription factor MEF2C influences neural stem/progenitor cell differentiation and maturation in vivo. Proc Natl Acad Sci U S A 105:9397-402
Nakagomi, Saya; Barsoum, Mark J; Bossy-Wetzel, Ella et al. (2008) A Golgi fragmentation pathway in neurodegeneration. Neurobiol Dis 29:221-31
Okamoto, Shu-ichi; Kang, Yeon-Joo; Brechtel, Christopher W et al. (2007) HIV/gp120 decreases adult neural progenitor cell proliferation via checkpoint kinase-mediated cell-cycle withdrawal and G1 arrest. Cell Stem Cell 1:230-6
Yuan, H; Gerencser, A A; Liot, G et al. (2007) Mitochondrial fission is an upstream and required event for bax foci formation in response to nitric oxide in cortical neurons. Cell Death Differ 14:462-71
Barsoum, Mark J; Yuan, Hua; Gerencser, Akos A et al. (2006) Nitric oxide-induced mitochondrial fission is regulated by dynamin-related GTPases in neurons. EMBO J 25:3900-11
Kaul, M; Lipton, S A (2005) Experimental and potential future therapeutic approaches for HIV-1 associated dementia targeting receptors for chemokines, glutamate and erythropoietin. Neurotox Res 8:167-86
Gu, Zezong; Cui, Jiankun; Brown, Stephen et al. (2005) A highly specific inhibitor of matrix metalloproteinase-9 rescues laminin from proteolysis and neurons from apoptosis in transient focal cerebral ischemia. J Neurosci 25:6401-8
Manabe, Shin-Ichi; Gu, Zezong; Lipton, Stuart A (2005) Activation of matrix metalloproteinase-9 via neuronal nitric oxide synthase contributes to NMDA-induced retinal ganglion cell death. Invest Ophthalmol Vis Sci 46:4747-53

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