Investigating Tau Mediated Toxicity in Alzheimer's Disease and the Protein Complex Responsible for Tau Cleavage

Alzheimer’s disease (AD) is a debilitating neurodegenerative disease resulting in memory loss due to the accumulation of two pathologic hallmarks: extracellular amyloid beta plaques and intracellular neurofibrillary tau tangles in the brain. In a normal healthy individual, tau functions as a microtubule associated protein that binds and stabilizes axonal microtubules. However, in an AD brain, tau dissociates from the microtubule forming filaments that result in tangles which are correlated with neuronal degeneration. Tau undergoes many different post-translational modifications (PTM), however, how they lead to toxicity remains unknown. Tau cleavage by caspases is one PTM that is believed to be a far more toxic early modification that precedes later pathological modifications such as phosphorylation and drives neuronal toxicity in AD. The current model of tau cleavage suggests that active caspases cleave tau at the aspartic acid421 site in the C-terminus tail region. The larger 1-421 fragment can be recognized in pre-tangles, and believed to change its conformation, generating toxic disease-associated tau species. We developed a proteasome impairment (depicting early disease) model that activates caspase-3 resulting in robust tau cleavage. We found that the absence of caspase-3 resulted in a loss of cleaved tau production suggesting caspase-3 is the main mediator driving tau cleavage. To investigate where tau cleavage occurs in the neuron, we found cleaved tau and cleaved caspase-3 both enriched at the post-synaptic density (PSD) where it may be causing synaptotoxicity leading to synapse damage. By over-expressing cleaved tau in neurons, we were able to further characterize the downstream effects of cleavage in neurons. Measuring synapse activity led us to find changes in neuronal activity and synchronization due to cleaved tau compared to controls suggesting the cleaved tau is toxic leading to synaptic dysfunction. Ubiquitin ligase carboxy terminus of Hsp70-interacting protein (also known as CHIP) is an E3 ubiquitin ligase that ubiquitinates proteins for degradation. Not only has CHIP been identified with insoluble tau suggesting its correlation with tau pathology but has also been found to interact with cleaved tau at the exposed 421 C-terminal region. Using our early disease proteasome impairment model, we found that overexpressing CHIP in neurons resulted in a significantly higher amount of cleavage production while CHIP knock-out alleviated levels although some tau was still able to undergo cleavage. To investigate whether one of CHIP’s functions (chaperone binding domain or ubiquitination domain) is specifically needed for cleavage to occur, we utilized two CHIP mutants, K30A (unable to bind chaperones) and ΔUBox (unable to ubiquitinate proteins). Tau cleavage levels were ameliorated even under impairment however, there was no difference in the reduction between the two mutants suggesting that both domains are needed for tau cleavage to occur. Along with cleaved tau and cleaved caspase-3 enriched at the PSD, we found that CHIP not only exacerbates cleaved tau production at the PSD, but CHIP also interacts with the PSD specifically under impairment conditions. Although much work still needs to be performed on how cleaved tau is causing toxicity to the neuron leading to neurodegeneration, the work of this dissertation identifies a mechanism in which how tau is cleaved leading to toxicity, and the protein complex involved leading to cleavage.