PT  - JOURNAL ARTICLE
AU  - Qi Zhang
AU  - Yue Xu
AU  - Juhyung Lee
AU  - Michal Jarnik
AU  - Xufeng Wu
AU  - Juan S. Bonifacino
AU  - Jingshi Shen
AU  - Yihong Ye
TI  - A Myosin-7B dependent endocytosis pathway mediates cellular entry of α-Synuclein fibrils and polycation-bearing cargos
AID  - 10.1101/2020.02.12.946137
DP  - 2020 Jan 01
TA  - bioRxiv
PG  - 2020.02.12.946137
4099  - http://biorxiv.org/content/early/2020/02/13/2020.02.12.946137.short
4100  - http://biorxiv.org/content/early/2020/02/13/2020.02.12.946137.full
AB  - Cell-to-cell transmission of misfolding-prone α-Synuclein (α-Syn) has emerged as a key pathological event in Parkinson’s disease. This process is initiated when α-Syn-bearing fibrils enter cells via clathrin-mediated endocytosis, but the underlying mechanisms are unclear. Using a CRISPR-mediated knockout screen, we identify SLC35B2 and Myosin-7B (MYO7B) as critical endocytosis regulators for α-Syn preformed fibrils (PFF). We show that SLC35B2, as a key regulator of heparan sulfate proteoglycan (HSPG) biosynthesis, is essential for recruiting α-Syn PFF to the cell surface because this process is mediated by interactions between negatively charged sugar moieties of HSPGs and clustered K-T-K motifs in α-Syn PFF. By contrast, MYO7B regulates α-Syn PFF entry by maintaining a plasma-membrane-associated actin network that controls membrane dynamics. Without MYO7B or actin filaments, many clathrin-coated pits fail to be severed from the membrane, causing accumulation of large clathrin-containing ‘scars’ on the cell surface. Intriguingly, the requirement for MYO7B in endocytosis is only restricted to α-Syn PFF and other cargos that enter cells via HSPGs. Thus, by identifying new regulatory factors for α-Syn PFF endocytosis, our study defines a mechanistically distinct clathrin-mediated endocytosis pathway that requires additional force generated by MYO7B and actin filaments.Significance The spreading of misfolded protein aggregates such as α-Synuclein preformed fibrils (α-Syn PFF) from cell to cell is a pathologic hallmark associated with the progression of many neurodegenerative diseases, but it is unclear how mammalian cells take up these large protein aggregates to initiate this prion-like protein transmission process. Here we define the mechanism of α-Syn PFF endocytosis using a combination of genetic, biochemical, and live-cell imaging techniques. Our study reveals how α-Syn PFF binds to the cell surface heparan sulfate proteoglycans using two lysine-bearing motifs and then enters cells following a Myosin-7B- and actin-dependent endocytosis mechanism that is specifically tailored for polycation-bearing cargos.