A genome-wide CRISPR/Cas9 screen identifies genes that regulate the cellular uptake of α-synuclein fibrils by modulating heparan sulfate proteoglycans

作者：Benoît Vanderperre, Amitha Muraleedharan, Marie‐France Dorion, Frédérique Larroquette, Esther del Cid‐Pellitero, Nishani Rajakulendran, Carol X Chen, Roxanne Larivière, Charlotte Michaud-Tardif, Rony Chidiac, Damien Lipuma, Graham MacLeod, Rhalena A. Thomas, Zejing Wang, Wolfgang Reintsch, Wen Luo, Irina Shlaifer, Z. Fuming, Ke Xia, Lufeng Yan, Zachary Steinhart, Robert J. Linhardt, Jean‐François Trempe, Jian Liu, Thomas M. Durcan, Stéphane Angers, Edward A. Fon · 发表于：bioRxiv (Cold Spring Harbor Laboratory) · 年份：2023 · DOI：10.1101/2023.09.29.560170 · 被引用次数：9 · 研究领域：Parkinson's Disease Mechanisms and Treatments、Cellular transport and secretion、Lysosomal Storage Disorders Research

ABSTRACT Synucleinopathies are characterized by the accumulation and propagation of α-synuclein (α-syn) aggregates throughout the brain, leading to neuronal dysfunction and death. Understanding how these aggregates propagate from cell to cell in a prion-like fashion thus holds great therapeutic promises. Here, we focused on understanding the cellular processes involved in the entry and accumulation of pathological α-syn aggregates. We used an unbiased FACS-based genome-wide CRISPR/Cas9 knockout (KO) screening to identify genes that regulate the accumulation of α-syn preformed fibrils (PFFs) in cells. We identified key genes and pathways specifically implicated in α-syn PFFs intracellular accumulation, including heparan sulfate proteoglycans (HSPG) biosynthesis and Golgi trafficking. We show that all confirmed hits affect heparan sulfate (HS), a post-translational modification known to act as a receptor for proteinaceous aggregates including of α-syn and tau. Intriguingly, KO of SLC39A9 and C3orf58 genes, encoding respectively a Golgi-localized exporter of Zn 2+ , and the Golgi-localized putative kinase DIPK2A, specifically impaired the uptake of α-syn PFFs uptake but not of tau oligomers, by preventing the binding of PFFs to the cell surface. Mass spectrometry-based analysis of HS chains indicated major defects in HS maturation in SLC39A9 and C3orf58 KO cells, explaining the cell surface binding deficit. Our findings now clearly establish these two genes as HSPG-modulating fa...