CRISPRi-based screens in iAssembloids to elucidate neuron-glia interactions.

The complexity of the human brain makes it challenging to understand the molecular mechanisms underlying brain function. Genome-wide association studies have uncovered variants associated with neurological phenotypes. Single-cell transcriptomics have provided descriptions of changes brain cells undergo during disease. However, these approaches do not establish molecular mechanism. To facilitate the scalable interrogation of causal molecular mechanisms in brain cell types, we developed a 3D co-culture system of induced pluripotent stem cell (iPSC)-derived neurons and glia, termed iAssembloids. Using iAssembloids, we ask how glial and neuronal cells interact to control neuronal death and survival. Our CRISPRi-based screens identified that GSK3β inhibits the protective NRF2-mediated oxidative stress response elicited by high neuronal activity. We then investigate the role of APOE-ε4, a risk variant for Alzheimer's disease, on neuronal survival. We find that APOE-ε4-expressing astrocytes may promote neuronal hyperactivity as compared with APOE-ε3-expressing astrocytes. This platform allows for the unbiased identification of mechanisms of neuron-glia cell interactions.