Disruption of Endosomal Microautophagy by Pathological Tau Protein - PROJECT SUMMARY/ABSTRACT One of the pathological features of Alzheimer’s disease (AD) is the dysfunction of protein degradation pathways including macro- and chaperone mediated autophagy. Interestingly, while tau is a substrate of these pathways, it can also contribute to their impairment. Our data and that from other groups suggest that tau also causes disruption of endosomal microautophagy (eMi) as well. Increased numbers of multivesicular bodies and higher levels of Hsc70, a chaperone involved in eMi, are found in AD patients and tau models. Further evidence of perturbation in the endosomal system can be seen in the association of tau and eMi pathway markers with granulovacuolar degeneration. In AD, disruption of eMi is especially important as can result in further buildup of tau aggregates, impaired neurotransmission, and spreading of pathology between cells. Tau potentially impacts eMi at multiple points in the pathway including: 1) decreased mitochondrial ATP production, complex expression, ROS production, and depolarization; 2) altering Hsc70 activity or reducing Hsc70 availability through aberrant binding or sequestration; 3) changes in the localization or levels of eMi proteins, and; 4) disruption of endosomal membrane integrity. We hypothesize that tau plays a causal role in destabilization of eMi through mitochondrial disruption as well as lowered availability of key pathway components via aberrant binding, changes in activity, or sequestration. The proposed experiments will answer these questions by elucidating the role of pathological tau in the disruption of eMi, clarifying which eMi components are vulnerable to tau pathology, and whether eMi normalization is sufficient to correct tau-induced deficits. In Aim 1 we will manipulate components of the microautophagy (eMi) pathway to determine their vulnerability to pathological tau. We will culture neurons from iPSCs derived from human patients, which express wild-type or mutant tau. Cells will be exposed to pathological tau isolated from AD brain, allowing us to measure the impact of wild-type, mutant, and aggregated tau on eMi. Further, we will determine the impact of tau on flux and which autophagy pathways are active, using live cell imaging. We will also directly alter the activity and/or expression of key eMi components, allowing us to discover which are most affected by tau pathology. In Aim 2 we will express mutant Hsc70, which is eMi competent but lacks ATPase function, in the brain of tauopathy mice. Published work and our preliminary data indicate that Hsc70 activity is altered in the presence of tau pathology. This approach will allow us to specifically target eMi, without interfering with the native Hsc70, to test whether normalizing its activity will rescue tau-induced dysfunction in eMi and neuronal signaling. An understanding of how tau pathology leads to disruption of eMi is of great relevance to public health. eMi is crucial for maintaining protein quality control and neuronal function, and impacts the spreading of tau aggregates. At the conclusion of the proposed experiments, we will have identified which aspects of eMi are most vulnerable to tau pathology and homed in on potential targets for future intervention.
