Although the trafficking of apoE4 through the ER and Golgi apparatus was significantly impaired compared with apoE3 (Figure 5A), blocking domain interaction by site-directed mutagenesis

Selleckchem Saracatinib (i.e., mutation of arginine-61 to threonine) or by exposure to small-molecule structure correctors restored normal trafficking properties to apoE4 (Figure 5B and 5C) and led to decreased neurotoxic fragment formation. These domain interaction-blocking approaches will be discussed in more detail below. Thus, it is envisioned that (1) the impaired transit of apoE4 occurs because of its abnormal structure, because blocking domain interaction restores the transit, (2) the abnormal structure and trafficking likely target the protein for proteolysis, and (3) small-molecule structure correctors likely target apoE as it is synthesized or soon after entering the ER lumen. Such findings suggest that one way to resolve the negative effects of apoE4 expression is to convert apoE4’s structure to be more apoE3-like. The cellular mechanisms and organelles

that promote Enzalutamide the clearance of abnormally folded proteins are ubiquitous, and abnormal forms of apoE, especially apoE4, can indeed be targeted for proteolysis. In fact, neurotoxic fragments are generated only by neurons, and not by astrocytes or other apoE-synthesizing cells (Brecht et al., 2004; Harris et al., 2003; Huang et al., 2001). Why, then, are neurons less effective than other cell types at completely degrading and clearing misfolded apoE? It is possible that, because apoE is an avid lipid-binding protein, lipid-based interactions may protect some domains from proteolytic cleavage, thus resulting in the accumulation of a spectrum of neurotoxic fragments. While Dabigatran full-length apoE is 34 kDa, a fragment pattern of bands ranging from 29–30 kDa to 12–14 kDa is consistently seen in extracts from cultured neurons expressing

apoE4, apoE4 transgenic mice and in the brains and cerebrospinal fluid from humans with AD (Brecht et al., 2004; Harris et al., 2003; Huang et al., 2001; Jones et al., 2011). Furthermore, more of these fragments are observed in AD patients expressing the apoE4 allele compared with normal, nondemented apoE4-carrying humans (Figure 6; Harris et al., 2003; Jones et al., 2011). Although the unique protease that is responsible for apoE4 fragmentation remains to be identified, it is thought to be a chymotrypsin-like serine protease (Harris et al., 2003). This protease, most likely residing in the ER or Golgi apparatus, generates the unique series of fragments ranging from 29–30 kDa to 12 kDa (Huang, 2010; Huang and Mucke, 2012; Mahley et al., 2006). The 29–30 kDa fragments result from cleavage at methionine-272 and leucine-268, respectively, and subsequent cleavage results in the generation of smaller fragments, primarily in the 12–20 kDa range.