Rapid iPSC inclusionopathy models shed light on formation, consequence and molecular subtype of α-synuclein inclusions

ABSTRACT

In neurodegenerative proteinopathies, intracellular inclusions are histopathologically and ultrastructurally heterogeneous but the significance of this heterogeneity is unclear. Patient-derived iPSC models, while promising for disease modeling, do not form analogous inclusions in a reasonable timeframe and suffer from limited tractability and scalability. Here, we developed an iPSC toolbox that utilizes piggyBac-based or targeted transgenes to rapidly induce CNS cells with concomitant expression of misfolding-prone proteins. The system is scalable and amenable to screening and longitudinal tracking at single-cell and single-inclusion resolution. For proof-of-principle, cortical neuron α-synuclein “inclusionopathy” models were engineered to form inclusions spontaneously or through exogenous seeding by α-synuclein fibrils. These models recapitulated known fibril- and lipid-rich inclusion subtypes in human brain, shedding light on their formation and consequences. Genetic-modifier and protein-interaction screens identified sequestered proteins in these inclusions, including RhoA, that were deleterious to cells when lost. This new iPSC platform should facilitate biological and drug discovery for neurodegenerative proteinopathies.