Expanding the electron cryo-tomography toolset to enable imaging of disease in the brain

Alzheimer’s disease involves a complex cascade of aberrant protein processing leading to aggregates of Aβ and Tau. Despite being hallmarks, aggregates do not necessarily result in cognitive decline. Approaches enabling the imaging of intermediates responsible for cytotoxicity within native brain tissue are needed. This will facilitate understanding of the molecular landscape underpinning protein dysfunction, cell death and disease. The cutting edge of structural biology is cellular. New instrumentation harnessing plasma ion sources allow tissues to be prepared for electron cryo-tomography (cryoET). However, new methodology is needed to robustly streamline tissue specimen preparation for this challenge. This is because a throughput of electron transparent specimens is needed to produce reliable insight. I will deliver a workflow combining reproducible vitrification of brain tissues, optimised sculpting of brain tissue by plasma ion beam milling and high-resolution TEM analysis. This will uncover the molecular changes in mouse disease models and human clinical samples. The organisation of macromolecules in healthy and disease mouse and human tissues and structures of disease-relevant intermediates of Aβ and Tau will be determined within their subcellular context. This will lead to a molecular understanding of mechanisms for cell disruption in native tissue, and a greater understanding of Alzheimer’s disease pathogenesis.