Genome-wide CRISPR screen identifies neddylation as a regulator of neuronal aging and AD neurodegeneration.

Aging is the biggest risk factor for the development of Alzheimer's disease (AD). Here, we performed a whole-genome CRISPR screen to identify regulators of neuronal age and show that the neddylation pathway regulates both cellular age and AD neurodegeneration in a human stem cell model. Specifically, we demonstrate that blocking neddylation increased cellular hallmarks of aging and led to an increase in Tau aggregation and phosphorylation in neurons carrying the APPswe/swe mutation. Aged APPswe/swe but not isogenic control neurons also showed a progressive decrease in viability. Selective neuronal loss upon neddylation inhibition was similarly observed in other isogenic AD and in Parkinson's disease (PD) models, including PSENM146V/M146V cortical and LRRK2G2019S/G2019S midbrain dopamine neurons, respectively. This study indicates that cellular aging can reveal late-onset disease phenotypes, identifies new potential targets to modulate AD progression, and describes a strategy to program age-associated phenotypes into stem cell models of disease.

TNF-NF-κB-p53 axis restricts in vivo survival of hPSC-derived dopamine neurons.

Ongoing, early-stage clinical trials illustrate the translational potential of human pluripotent stem cell (hPSC)-based cell therapies in Parkinson's disease (PD). However, an unresolved challenge is the extensive cell death following transplantation. Here, we performed a pooled CRISPR-Cas9 screen to enhance postmitotic dopamine neuron survival in vivo. We identified p53-mediated apoptotic cell death as a major contributor to dopamine neuron loss and uncovered a causal link of tumor necrosis factor alpha (TNF-α)-nuclear factor κB (NF-κB) signaling in limiting cell survival. As a translationally relevant strategy to purify postmitotic dopamine neurons, we identified cell surface markers that enable purification without the need for genetic reporters. Combining cell sorting and treatment with adalimumab, a clinically approved TNF-α inhibitor, enabled efficient engraftment of postmitotic dopamine neurons with extensive reinnervation and functional recovery in a preclinical PD mouse model. Thus, transient TNF-α inhibition presents a clinically relevant strategy to enhance survival and enable engraftment of postmitotic hPSC-derived dopamine neurons in PD.

Deep sequencing of proteotoxicity modifier genes uncovers a Presenilin-2/beta-amyloid-actin genetic risk module shared among alpha-synucleinopathies.

Whether neurodegenerative diseases linked to misfolding of the same protein share genetic risk drivers or whether different protein-aggregation pathologies in neurodegeneration are mechanistically related remains uncertain. Conventional genetic analyses are underpowered to address these questions. Through careful selection of patients based on protein aggregation phenotype (rather than clinical diagnosis) we can increase statistical power to detect associated variants in a targeted set of genes that modify proteotoxicities. Genetic modifiers of alpha-synuclein (ɑS) and beta-amyloid (Aß) cytotoxicity in yeast are enriched in risk factors for Parkinson's disease (PD) and Alzheimer's disease (AD), respectively. Here, along with known AD/PD risk genes, we deeply sequenced exomes of 430 ɑS/Aß modifier genes in patients across alpha-synucleinopathies (PD, Lewy body dementia and multiple system atrophy). Beyond known PD genes GBA1 and LRRK2, rare variants AD genes (CD33, CR1 and PSEN2) and Aß toxicity modifiers involved in RhoA/actin cytoskeleton regulation (ARGHEF1, ARHGEF28, MICAL3, PASK, PKN2, PSEN2) were shared risk factors across synucleinopathies. Actin pathology occurred in iPSC synucleinopathy models and RhoA downregulation exacerbated ɑS pathology. Even in sporadic PD, the expression of these genes was altered across CNS cell types. Genome-wide CRISPR screens revealed the essentiality of PSEN2 in both human cortical and dopaminergic neurons, and PSEN2 mutation carriers exhibited diffuse brainstem and cortical synucleinopathy independent of AD pathology. PSEN2 contributes to a common-risk signal in PD GWAS and regulates ɑS expression in neurons. Our results identify convergent mechanisms across synucleinopathies, some shared with AD.