The inter-cellular prion-like propagation of α-synuclein aggregation is emerging as an important mechanism driving the progression of neurodegenerative diseases including Parkinson’s disease and multiple system atrophy (MSA). To discover therapeutic strategies reducing the spread of α-synuclein aggregation, we performed a genome-wide CRISPR interference screen in a human cell-based model. We discovered that inhibiting PIKfyve dramatically reduced α-synuclein aggregation induced with both recombinant α-synuclein fibrils and fibrils isolated from MSA patient brain. While PIKfyve inhibition did not affect fibril uptake or α-synuclein clearance or secretion, it reduced α-synuclein trafficking from the early endosome to the lysosome, thereby limiting fibril escape from the lysosome and reducing the amount of fibrils that reach cytosolic α-synuclein to induce aggregation. These findings point to the endolysosomal transport of fibrils as a critical step in the propagation of α-synuclein aggregation and a potential therapeutic target.

Rejuvenation, long a quixotic dream, recently became a possibility through exciting new approaches to counteract aging. For example, parabiosis and partial reprogramming through overexpressing four stem cell transcription factors (Yamanaka factors) both rejuvenate organisms and cells. We hypothesize there are many other genetic solutions to human cell rejuvenation, and some solutions may be safer and more potent than current gene targets. We set out to develop a systematic approach to identify novel genes that, when overexpressed or repressed, reprogram the global gene expression of a cell back to a younger state. Using the Hayflick model of human cell replicative aging, we performed a Perturb-seq screen of 200 transcription factors (TFs) selected through a combination of bioinformatic analysis and literature search. We identified dozens of potentially rejuvenating TFs, those that when overexpressed or repressed in late passage cells reprogrammed global gene expression patterns back to an earlier passage state. We further validated four top TF perturbations through molecular phenotyping of various aging hallmarks. Late passage cells either overexpressing EZH2 or E2F3 or repressing STAT3 or ZFX had more cell division, less senescence, improved proteostasis, and enhanced mitochondrial function. These TF perturbations led to similar downstream gene expression programs. In addition, the rejuvenating effects of these TFs were independent of telomeres. We believe our general approach for identifying rejuvenating factors can be applied to other model systems, and some of the top TF perturbations we discovered will lead to future research in novel, safer rejuvenation therapies.