One of the main challenges in modern societies is the increase in average life span that is associated with an increase in age-related disorders such as Alzheimer's disease (AD) or Parkinson's disease (PD). Autophagy is increasingly recognized as a key factor in counteracting age and age-related disorders such as neurodegeneration. A major hallmark of many neurodegenerative disorders is the accumulation of misfolded proteins, and the presence of such characteristic protein aggregates in those late-onset neurodegenerative disorders indicates that protein homeostasis may be overwhelmed. Among the various components of the proteostasis machinery, only the autophagosome-lysosome system is capable to engulf large protein aggregates via a process called macroautophagy (herein after called autophagy). A direct experimental link between autophagy and neurodegeneration was demonstrated by the neuron- and macroglia-specific knockout of essential core autophagy genes such as ATG5 or ATG7 in the CNS of mice. Those studies demonstrated the progressive accumulation of ubiquitinated proteins forming inclusion bodies in neurons, followed by neuronal loss and premature death. Despite the strong evidence linking autophagy to neurodegeneration and aging in model organisms, there is very limited information about the role of autophagy in authentic, disease-relevant human neurons. Here we propose to establish an inducible and reversible human model of autophagy inhibition to test the hypothesis that autophagy inhibition in specific neuronal lineages will mimic the characteristic aggregation of proteins observed in the corresponding human neurodegenerative disorders providing a rationale for age-related sporadic manifestation of disease phenotypes. For this purpose, we will utilize human pluripotent stem cells (hPSCs) that enable routine access to disease-relevant neurons at high purity and scale including cortical, midbrain dopaminergic and spinal motoneurons, the key lineages affected in AD, PD and ALS, respectively. We will assess the impact of autophagy inhibition at sequential stages of differentiation for each neuronal subtype and determine reversibility. Importantly, we will use unbiased, state-of-the-art proteomics to determine whether manipulation of autophagy is sufficient to induce proteomic changes that mimic known and potentially novel neuron subtype-specific processes involved during neurodegeneration in AD, PD and ALS. Finally, we will address to what extent those changes are reversible and whether we can use this approach to define novel candidate therapeutic targets that may intervene with disease progression of the neurodegenerative process.
There is a close link between an age-related decline in autophagy function and the onset of pathogenesis of various neurodegenerative disorders. Here, we will use human pluripotent stem cell technology to manipulate autophagy in an inducible and reversible manner in specific neuronal lineages vulnerable to neurodegenerative disease. We will test the hypothesis that loss of autophagy triggers a cascade of neurodegenerative changes characteristic for each neuron type, that unbiased proteomics can be used to define the sequence of neurodegenerative changes at the level of protein homeostasis and that this approach can yield novel candidate therapeutic targets to counteract the neurodegenerative process.
