Our yeast model allowed us to conduct an unbiased screen of an entire genome for modifiers of Aβ toxicity. The emergence of three different genes involved in the process of clathrin-mediated endocytosis from nearly 6,000 tested ORFs confirms that the Aβ peptide in our model is trafficking through the secretory compartments as expected. More importantly, the ability of endocytic genes to rescue Aβ toxicity, together with the effects of Aβ on clathrin localization and the trafficking of a G protein-coupled receptor, establish that within these highly diverse organisms clathrin-mediated endocytosis is a critical point of vulnerability to Aβ.
Aβ oligomers have been reported to increase endocytosis in cultured cells (39
), and human-induced neuronal cells derived from the fibroblasts of AD patients exhibit defects in endocytosis (40
). Mechanistically, in our Aβ-expressing cells, the increased number of clathrin foci, the internalized foci of Ste3, and the effects of genetic modifiers on vacuolar localization all suggest that Aβ affects this pathway by interfering with the ability of endocytosed transmembrane receptors to reach their proper destinations.
PICALM, as well as two genes whose protein products (BIN1 and CD2AP) interact with hits from our screen are AD risk factors. Given the diversity of pathologies however, their connection to Aβ toxicity was unknown. Our work in yeast, nematodes, and rat cortical neurons clearly places these factors within the Aβ cascade, linking Aβ to the genetics of sporadic AD.
Neurons are particularly vulnerable to perturbations in the homeostasis of endocytosis, because they must constantly recycle both neurotransmitters and their receptors (41
). Aβ interacts with, and alters signaling by, a variety of neuronal receptors (42
). We propose that the conformational flexibility of these oligomers allows them to interact rather promiscuously with conformationally flexible unliganded receptors, which, in turn, disrupts endocytic homeostasis.
Our yeast screen also identified seven conserved genes functionally associated with the cytoskeleton. Because yeasts do not express tau, our findings may indicate that the connection between Aβ toxicity and the cytoskeleton is more deeply rooted than tau alone, probably involving clathrin-mediated endocytosis. In analyzing human GWAS data we also uncovered suggestive associations between AD and three other genes, XPO1, ADSSL1, and RABGEF1, and confirmed their Aβ relationships in yeast and nematode.
The treatments available for AD are few and their efficacy limited. Determining how best to rescue neuronal function in the context of the whole brain is a problem of staggering proportions. Our yeast model provides a tool for identifying genetic leads, investigating their mechanisms of action, and screening for genetic and small molecule modifiers of this devastating and etiologically complex disease.