Beyond the SNARE: Munc18-1 chaperones α-synuclein

641 The Rockefeller University Press $30.00 J. Cell Biol. Vol. 214 No. 6 641–643 www.jcb.org/cgi/doi/10.1083/jcb.201608060 Early infantile epileptic encephalopathy (EIEE), also known as Ohtahara syndrome, is a debilitating neurological disorder that results in early-onset tonic seizures and severe intellectual disability. Heterozygous mutations in MUNC18-1 (also known as syntaxin binding protein 1 [STX BP1]) are linked to EIEE (Saitsu et al., 2008; Stamberger et al., 2016). Munc18-1 was first identified as an essential component of the synaptic vesicle fusion machinery and binds to the SNA RE receptor Syntaxin-1A (Hata et al., 1993). Munc18-1 plays an important role in regulating neuronal exocytosis by acting as a molecular chaperone for Syntaxin-1A and regulating its levels at the plasma membrane and has additional functions in vesicle fusion (Han et al., 2011; Ma et al., 2015; Shen et al., 2015). Although the genetic link between Munc18-1 and EIEE has been established through multiple studies, it remains unclear how loss of Munc18-1 leads to EIEE. One EIEE-associated Munc18-1 mutation (C180Y) is located in the hydrophobic core of the protein and results in decreased thermostability (Saitsu et al., 2008; Martin et al., 2014). Although Munc18-1C180Y retains the ability to bind Syntaxin-1A, it has a tendency to form intracellular aggregates, which are targeted for proteosomal degradation (Martin et al., 2014). Therefore, one model to explain EIEEassociated haploinsufficiency is that Munc18-1C180Y mutants draw wild-type Munc18-1 proteins into aggregates, thereby lowering the levels of available functional molecules. In this issue, Chai et al. directly tested this model by analyzing aggregate formation in the presence of Munc18-1C180Y mutants. Using single-molecule fluorescence spectroscopy in a cell-free system, Chai et al. (2016) found that mutant Munc18-1 coaggregated with wild-type Munc18-1 in vitro. The researchers tested the ability of Munc18-1C180Y to recruit new monomers, and thus seed larger aggregates, by using Munc18-1C180Y tagged with two different fluorophores. These experiments showed that small aggregates of the mutant protein could seed formation of larger fibrils composed not only of the mutant Munc18-1 but also of the wild-type protein. Coaggregation with wild-type protein also occurred in vivo, when the authors expressed Munc18-1C180Y in PC12 pheochromocytoma cells or in rat hippocampal neurons. Several other EIEE-associated Munc18-1 mutants exhibited a similar ability to cause wild-type Munc18-1 aggregation in cells. Thus, the authors conclude that EIEE mutants are likely to act dominantly by sequestering wildtype Munc18-1 into aggregates (Fig. 1 A). A surprising twist came when Chai et al. (2016) noticed Lewy body–like structures in Munc18-1C180Y–expressing cells. Lewy bodies are fibrillar intracellular inclusions that are hallmarks of Parkinson’s disease and Lewy body dementia. The primary component of Lewy bodies is α-synuclein, a presynaptic protein highly expressed in the brain (Burré, 2015). The researchers followed up on this observation by showing that α-synuclein coaggregated with Munc18-1C180Y oligomers by single-molecule fluorescence spectroscopy in the cell-free system. In PC12 cells and in hippocampal neurons, expression of Munc18-1C180Y and other EIEE-linked mutants resulted in recruitment of α-synuclein into Lewy body–like aggregates. Conversely, Parkinson’s disease–associated aggregation-prone mutations in α-synuclein drew wild-type Munc18-1 into aggregates (Fig. 1 B). Finally, Chai et al. (2016) showed that endogenous α-synuclein coimmunoprecipitated with endogenous Munc18-1. Together, these results point to the interesting new hypothesis that Munc18-1 serves as a chaperone for α-synuclein, in addition to its role in SNA RE regulation. A molecular chaperone facilitates proper folding, assembly, and disassembly of its target proteins and prevents aggregation. In support of a role for endogenous Munc18-1 in chaperoning α-synuclein, Chai et al. (2016) showed that loss of Munc18-1 leads to increased α-synuclein aggregation, in both neurosecretory cells and in hippocampal neurons (Fig. 1 C). This α-synuclein aggregation phenotype could be rescued by reintroducing Munc18-1 in a dose-dependent manner. Thus, the ability of endogenous Munc18-1 to bind α-synuclein and to control its capacity to aggregate suggests that it has a physiological chaperone-like function for α-synuclein. What role does the Munc18-1–α-synuclein interaction play in a cell, and how might this impinge on the pathological roles of α-synuclein in neurological disease? α-Synuclein binds directly to membranes and has several proposed functions in normal membrane trafficking, including serving as a chaperone for SNA RE assembly via an interaction with the SNA RE synaptobrevin-2 (Burré et al., 2010), as well as additional functions in diverse other processes, including membrane remodeling, lipid metabolism, neurotransmitter synthesis and transport, and synaptic vesicle mobilization (Burré, 2015). In pathological states Early infantile epileptic encephalopathy (EIEE)–associated mutations in MUNC18-1 cause Munc18-1 misfolding and cellular aggregation. In this issue, Chai et al. (2016. J. Cell Biol. http ://dx .doi .org /10 .1083 /jcb .201512016) find that Munc18-1 is a molecular chaperone for α-synuclein and that aggregated Munc18-1 EIEE-causing mutants promote α-synuclein aggregation. Beyond the SNA RE: Munc18-1 chaperones α-synuclein