Protein mishandling and impaired lysosomal proteolysis generated through calcium dysregulation in Alzheimer’s disease

Abstract Impairments in neural lysosomal- and autophagic-mediated degradation of cellular debris contribute to neuritic dystrophy and synaptic loss. While these are well-characterized features of neurodegenerative disorders such as Alzheimer’s disease (AD), the upstream cellular processes driving deficits in pathogenic protein mishandling are less understood. Using a series of fluorescent biosensors and optical imaging in model cells, AD mouse models and human neurons derived from AD patients, we reveal a novel cellular signaling cascade underlying protein mishandling mediated by intracellular calcium dysregulation, an early component of AD pathogenesis. Increased Ca 2+ release via the endoplasmic reticulum (ER) resident ryanodine receptor (RyR) is associated with reduced expression of the lysosome proton pump vATPase subunits (V1B2 and V0a1), resulting in lysosome deacidification and disrupted proteolytic activity in AD mouse models and human induced neurons (HiN). As a result of impaired lysosome digestive capacity, mature autophagosomes with hyperphosphorylated tau accumulated in AD murine neurons and AD HiN, exacerbating proteinopathy. Normalizing AD-associated aberrant RyR-Ca 2+ signaling with the negative allosteric modulator, dantrolene (Ryanodex), restored vATPase levels, lysosomal acidification and proteolytic activity, and autophagic clearance of intracellular protein aggregates in AD neurons. These results highlight that prior to overt AD histopathology or cognitive deficits, aberrant upstream Ca 2+ signaling disrupts lysosomal acidification and contributes to pathological accumulation of intracellular protein aggregates. Importantly, this is demonstrated in animal models of AD, and in human iPSC-derived neurons from AD patients. Furthermore, pharmacological suppression of RyR-Ca 2+ release rescued proteolytic function, revealing a target for therapeutic intervention that has demonstrated effects in clinically-relevant assays. Significance Statement We demonstrate in model cells, murine neuronal cultures, and iPSC-derived human neurons, that AD associated RyR-Ca 2+ dyshomeostasis impairs lysosomal acidification, lysosomal proteolytic activity and hinders autophagic-mediated protein aggregate clearance, which are processes vital to neuronal survival. These deficits were reversed by restoring intracellular Ca 2+ homeostasis. Notably, this provides a therapeutic target and emphasizes the pathogenic relationship between ER-Ca 2+ handling, that is known to be altered in AD, to pathogenic protein accumulation as a critical turning point in early stages of Alzheimer’s disease.