Since the large proportion of soluble synuclein makes it difficult to detect a membrane-bound fraction by morphological techniques in most cells other than neurons, digitonin was used to permeabilize selectively the plasma membrane of HeLa cells expressing human α-synuclein, releasing the unbound cytosolic protein (Fortin et al., 2004). The remaining synuclein appeared punctate but failed to colocalize with markers for many organelles. Rather, it colocalized with components of lipid rafts, a membrane microdomain with reduced fluidity
that is enriched in cholesterol and saturated acyl chains (Fortin et al., 2004). The PD-associated A30P mutation abolished this localization, supporting the specificity of the interaction, and the biochemical analysis of detergent-resistant EPZ-6438 order membranes by flotation gradient confirmed the localization to rafts. Importantly, the disruption of lipid rafts also prevents the accumulation of synuclein in presynaptic boutons (Fortin et al., 2004), supporting selleck screening library the relevance of
this interaction for neurons. In addition to the requirement for acidic phospholipid, biochemical studies in vitro have indicated that synuclein requires a combination of phospholipid with oleoyl as well as polyunsaturated acyl chains (Kubo et al., 2005), suggesting that it may specifically recognize the phase boundary that arises between membranes that differ in fluidity. Remarkably, there was an apparent requirement for the acidic headgroup on the polyunsaturated acyl rather than oleoyl chain (Kubo et al., 2005), raising the possibility of a distinct and previously unknown microdomain in neurons. Further, recent work has found that synuclein can influence lipid packing within raft-like domains containing cholesterol (Leftin et al., 2013), suggesting that synuclein may not simply be recruited by these structures but also contributes to their formation, very similar to other peripheral membrane proteins such as caveolin (Parton and del Pozo, 2013). It
has also been suggested that synuclein might act as a fatty acid binding protein (Sharon et al., 2001). Synuclein however promotes the uptake of polyunsaturated fatty acids into cells, and polyunsaturated fatty acids promote the oligomerization of synuclein (Assayag et al., 2007, Perrin et al., 2001, Sharon et al., 2003a and Sharon et al., 2003b). Supporting a role for this activity in vivo, the analysis of α-synuclein knockout mice has shown remarkable changes in brain cardiolipin, including acyl chain composition (Ellis et al., 2005). Fatty acid uptake and metabolism also appear affected (Golovko et al., 2005, Golovko et al., 2006 and Golovko et al., 2007), although with only modest changes in other brain phospholipids (Barceló-Coblijn et al., 2007 and Rappley et al., 2009b).