APP Is Cleaved by Bace1 in Pre-Synaptic Vesicles and Establishes a Pre-Synaptic Interactome, via Its Intracellular Domain, with Molecular Complexes that Regulate Pre-Synaptic Vesicles Functions

Abstract

Amyloid Precursor Protein (APP) is a type I membrane protein that undergoes extensive processing by secretases, including BACE1. Although mutations in APP and genes that regulate processing of APP, such as PSENs and BRI2/ITM2B, cause dementias, the normal function of APP in synaptic transmission, synaptic plasticity and memory formation is poorly understood. To grasp the biochemical mechanisms underlying the function of APP in the central nervous system, it is important to first define the sub-cellular localization of APP in synapses and the synaptic interactome of APP. Using biochemical and electron microscopy approaches, we have found that APP is localized in pre-synaptic vesicles, where it is processed by Bace1. By means of a proteomic approach, we have characterized the synaptic interactome of the APP intracellular domain. We focused on this region of APP because in vivo data underline the central funtional and pathological role of the intracellular domain of APP. Consistent with the expression of APP in pre-synaptic vesicles, the synaptic APP intracellular domain interactome is predominantly constituted by pre-synaptic, rather than post-synaptic, proteins. This pre-synaptic interactome of the APP intracellular domain includes proteins expressed on pre-synaptic vesicles such as the vesicular SNARE Vamp2/Vamp1 and the Ca2+ sensors Synaptotagmin-1/Synaptotagmin-2, and non-vesicular pre-synaptic proteins that regulate exocytosis, endocytosis and recycling of pre-synaptic vesicles, such as target-membrane-SNAREs (Syntaxin-1b, Syntaxin-1a, Snap25 and Snap47), Munc-18, Nsf, α/β/γ-Snaps and complexin. These data are consistent with a functional role for APP, via its carboxyl-terminal domain, in exocytosis, endocytosis and/or recycling of pre-synaptic vesicles.

Novel Insights into the Physiological Function of the APP (Gene) Family and Its Proteolytic Fragments in Synaptic Plasticity

“…It is further hypothesized that homodimerized APP acts as a G-Protein coupled receptor which is activated by Aß and might be involved in neurotransmitter release following enhanced Ca 2+ influx. Especially the intracellular domains of APP and APLP2 seem to be associated with proteins of the synaptic vesicle release machinery regulating the molecular composition of synaptic vesicles at the presynaptic active zone (Del Prete et al, 2014; Fanutza et al, 2015; Laßek et al, 2015). At the postsynaptic compartment (Figure 2C) patterns of synaptic activity modulate APP family protein processing.…”

“…APP and APLP2 show a variety of possible interactions with the synaptic vesicle release machinery: Biochemical approaches showed that APP is associated with synaptic vesicle proteins (Del Prete et al, 2014; Laßek et al, 2014) and that it can be cleaved within vesicles by BACE-1 (Del Prete et al, 2014). Especially the intracellular regions of APP, APLP2, and CTF-ß have been shown to interact with presynaptic vesicle proteins like Rab, AP-2 subunits, the Ca 2+ sensors synaptotagmins, clathrin, and complexin (Del Prete et al, 2014; Fanutza et al, 2015). Results from APP-KO animals point toward a role of APP in controlling synaptic vesicle protein content in the presynaptic active zone as synaptophysin, synaptotagmin-1, and SV2A protein levels are reduced in APP KO mice.…”

“…Subsequent processing of the remaining membrane anchored CTF η by the α- or ß-secretase generates two new peptides, Aη-α and Aη-ß (Willem et al, 2015). Importantly, APP processing is not restricted to the plasma membrane, but was also shown to occur within synaptic vesicles (Del Prete et al, 2014). …”

Amyloid Precursor Protein (APP) May Act as a Substrate and a Recognition Unit for CRL4CRBN and Stub1 E3 Ligases Facilitating Ubiquitination of Proteins Involved in Presynaptic Functions and Neurodegeneration

“…This short sequence lacks enzymatic activity and may function as a docking domain for cytosolic as well as membrane-bound proteins (46,(55)(56)(57)(58)(59)(60)(61)(62)(63)(64)(65)(66)(67)(68)(69)(70)(71)(72). To explore the potential role of the ACR, we used a proteomic approach (28,48 ), and St-ACR with phosphorylation on both Thr 668 and Tyr 682 (St-ACR Thr(P)Tyr(P) ), were immobilized on StrepTactin resin. The numbering of phosphorylated residues is based on the APP isoform of 695 amino acids.…”

“…Using an unbiased proteomic approach, we have characterized the ACR brain interactome (28,48). Here, we show that the ACR interacts with several proteins that regulate the UPS.…”