Ischemic optic neuropathy is a type of central nervous system (CNS) white matter stroke that causes permanent blindness due to retinal ganglion cell (RGC) axon damage and cell death. Although such white matter ischemia comprises an important source of morbidity in humans, the molecular mechanisms contributing to neuronal dysfunction and death remain poorly understood, and there exist few effective therapies to prevent or restore loss of vision. A major goal for this research is to define the signaling pathways defective in neurons following ischemic injury, so that novel therapeutic regimens that confer neuroprotection and neuroregeneration may be rationally designed. The myocyte enhancer factor 2 (MEF2) transcription factor family is an important mediator of pro-survival signaling in neurons that is activated by both retrograde neurotrophin stimulation and neuronal depolarization. Transcriptional activity of MEF2A and MEF2D, the primary MEF2 isoforms expressed in RGCs, is regulated by post-translational modification of distinct amino acid residues by extracellular signal-regulated kinase 5 (ERK5) catalyzed phosphorylation and calcineurin (CaN) phosphatase catalyzed dephosphorylation. MEF2A/D, ERK5, and CaN are all binding partners for the scaffold protein muscle A-kinase anchoring protein ? (mAKAP?/AKAP6), a scaffold protein expressed within the retina primarily in RGCs. In this application we will test the novel hypothesis that mAKAP? signalosomes serve as critical nodes in the neuronal signal transduction network by orchestrating MEF2 activation in neuroprotection and axon growth.
In Aim 1 using primary cultures, we will determine the regulation and function of MEF2A/D post- translational modifications, including phosphorylation and sumoylation, by mAKAP? signalosomes in response to brain-derived neurotrophic factor (BDNF) and depolarizing stimuli.
In Aim 2 using conditional MEF2 knock-out mice and a photochemically-induced ischemic optic neuropathy (PCI-ION) model, we will test whether MEF2A and MEF2D transcriptional activity are required for RGC survival in stroke and for the pro-survival and regenerative effects of neurotrophic factors such as BDNF after intravitreal injection.
In Aim 3, we propose to enhance neuroprotection and regeneration following PCI-ION by increasing ERK5 signaling and MEF2 activity using intravitreal injection of adeno-associated virus (AAV) gene therapy vectors to express constitutively active MEK5 and MEF2 proteins in RGCs in vivo. Data from these experiments will advance our basic understanding of how MEF2 is regulated in neurons and how compartmentalized signaling regulates neuronal survival and regeneration, ultimately contributing to new therapeutic strategies for stroke and optic neuropathy.
Ischemic optic neuropathy (optic nerve stroke) directly impacts the vision of over 2/3 of patients, but there are no therapies that target retinal ganglion cell survival or regeneration in these diseases. A better understanding of the cellular mechanisms that impact neuroprotection and axonal regeneration may yield better therapeutic regimens, decreasing morbidity and potentially restoring vision.
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