Neurodegenerative diseases afflict millions of individuals ranging from children to the elderly. Without known causes or effective therapies, this class of diseases outstrips the brain's capacity for compensatory neuronal plasticity with debilitating results. Two pathways involved with neurodegeneration are mitochondrial dysfunction and macroautophagy. This research proposal focuses on the interplay between these two pathways as regulators of axonal and dendritic degeneration (neurite degeneration). Macroautophagy (hereafter, autophagy) is the bulk catabolism of long-lived proteins and organelles, including mitochondria. While basal levels of autophagy are required for health, dysregulation has been implicated as a cause for neurite degeneration. Mitochondria are ubiquitous organelles that play key roles in the proper function and plasticity of neurons. Dysregulated macroautophagy and mitochondrial dysfunction/loss have been implicated in the pathogenesis of Parkinson's disease (PD). Recent studies have illustrated the neuroprotective activity of the PTEN-induced kinase 1 (PINK1). Additionally, PINK1 has been linked to autosomal recessive PD, presumably through loss of function mutations. I hypothesize that PINK1 signaling regulates neurite degeneration through its effects on autophagy/mitophagy. The role of PINK1 in regulating neurite degeneration, autophagy, and mitochondrial loss will be measured in vitro by biochemical and microscopy studies.
Aim 1 will determine whether levels of PINK1 expression regulate autophagy and protect against neuronal injuries that cause neurite degeneration.
Aim 2 will determine whether PINK1 knockdown or disease-associated mutations induce autophagy and neurite degeneration. I will also determine whether PINK1 knockdown or mutants require autophagy for neurite degeneration.
Aim 3 will use a non-biased proteomies approach to identify downstream mitochondrial targets of PINK1. Phosphorylated protein profiles will be compared between PINK1 deficient mice or cells and wild type mice or cells, respectively. Potential downstream PINK1 targets will be identified as phosphoproteins in wild-type samples that are reduced in PINK1 deficient samples. Using RNAi, I will determine if the putative downstream PINK1 mediators are required for the PINK1 neuroprotective effects. By identifying potential pathways that regulate PINK1-mediated neuroprotection and autophagic """"""""selfdigestion"""""""", the results of this study will provide insight into the mechanisms of neurite degeneration in Parkinson's and related diseases. In addition to providing predoctoral training to the applicant, this research proposal is designed to uncover promising drug targets for the treatment of neurodegenerative diseases.
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