Insulin-Degrading Enzyme Sorting in Exosomes: A Secretory Pathway for a Key Brain Amyloid-β Degrading Protease

Abstract

The accumulation of Aβ peptides in the senile plaques is one of the hallmarks of Alzheimer disease (AD) progression. The endocytic pathway has been proposed as a major subcellular site for Aβ generation while the compartments in which Aβ-degrading proteases interact with Aβ are still elusive. It was suggested that extracellular Aβ degradation may take place by plasma-membrane associated proteases or by extracellular proteases, among which insulin-degrading enzyme (IDE) is the most relevant. However, the mechanisms of IDE secretion are poorly understood. In the present study we used N2a cells to explore if IDE is indeed released through exosomes and the effect of exosomes release on extracellular levels of Aβ. We demonstrated that proteolitically active plasma membrane associated-IDE is routed in living N2a cells to multivesicular bodies and subsequently, a major fraction is sorted to exosomes. We described that extracellular IDE levels decrease if the MVBs generation is interfered and may be positively modulated by exosomes release under stress-induced conditions. Our results reinforce the relevance of functional IDE in the catabolism of extracellular Aβ.

Inositol phosphates and phosphoinositides activate insulin-degrading enzyme, while phosphoinositides also mediate binding to endosomes

“…For example, insulin is metabolized primarily in early to late endosomes after internalization of the peptide bound to its receptor (23,52,53) and thus is inaccessible to cytosolic IDE. However, small pools of IDE have been found in endosomes (23)(24)(25) as confirmed here, as well as in peroxisomes (26,54) and mitochondria (55). In addition, low levels of IDE are reportedly secreted from some cell types through an unconventional pathway (56) that may involve routing through multivesicular bodies and release in association with exosomes (25).…”

“…Given that IDE has been reported to be present in endosomes (23)(24)(25), we hypothesized that the enzyme initially might be recruited to endosomes by binding to PtdIns(3)P known to be present in the outer leaflet of the endosomal membrane (21,22,31). Discontinuous sucrose density centrifugation was used to enrich endosomes from COS-1 cells as described by Jang et al (32).…”

“…IDE recruitment to endosomes also may be relevant to the mechanism underlying the reported secretion of IDE from certain cell types (25,56,69). Secreted IDE has been proposed to be involved in extracellular Aβ degradation (6,70,71) and potentially some insulin hydrolysis as well (72,73).…”

“…Secreted IDE has been proposed to be involved in extracellular Aβ degradation (6,70,71) and potentially some insulin hydrolysis as well (72,73). IDE does not contain any standard secretion signaling sequences, reportedly being secreted by unconventional pathways (25,56,69,71). Enzyme localized to the endosomal system could contribute to this secreted pool, given that some multivesicular bodies fuse with the cell plasma membrane, releasing their contents, including the intraluminal vesicles (exosomes), into the extracellular medium (25,74).…”

“…Found mainly in the cytosol, IDE also has been reported to be associated with various subcellular compartments, including endosomes (23)(24)(25) and peroxisomes (26). It also is reported to be secreted from cells.…”