Cell Surface Amyloid β-Protein Precursor Colocalizes with β1 Integrins at Substrate Contact Sites in Neural Cells

Abstract

Amyloid ␤-protein (A␤), the principal constituent of the senile plaques seen in Alzheimer's disease (AD), is derived by proteolysis from the ␤-amyloid precursor protein (␤PP). The distribution and trafficking of cell surface ␤PP are of particular interest because some of these molecules are direct precursors of secreted A␤ and because the localization of ␤PP at the cell surface may be related directly to its physiological functions. Recently, we reported that, in cultured hippocampal neurons, cell surface ␤PP is preferentially expressed on axons in a striking discontinuous pattern. In this study, we describe the colocalization of cell surface ␤PP and integrins in primary cultured cells. In rat hippocampal neurons, cell surface ␤PP was colocalized selectively with ␣1␤1 and ␣5␤1 integrin heterodimers at these characteristic segmental locations. In rat cortical astrocytes, both cell surface ␤PP and ␤1 integrin were located at the cell periphery in the "spreading" stage shortly after plating. In "flattened" astrocytes cultured for several days, ␤PP was found in punctate deposits called point contacts. In these sites, ␤PP was colocalized with ␣1␤1, but not with ␣5␤1 integrin heterodimers, the latter of which were situated at focal contact sites. In both neurons and astrocytes examined after shearing, clathrin and ␣-adaptin were colocalized with ␤PP on the surface that directly contacts the substratum. These results are consistent with the putative role of ␤PP in cell adhesion and suggests that ␤PP either interacts with selected integrins or shares similar cellular machinery to promote cell adhesion. Key words: amyloid ␤-protein precursor; amyloid ␤-protein; integrins; cell adhesion; clathrin; substrate attachment; point contactsAlzheimer's disease (AD) is a progressive, neurodegenerative disorder characterized by the extracellular deposition of the 39 -43 amino acid amyloid ␤-protein (A␤) in the brain parenchyma (Selkoe, 1994). A␤ is derived by proteolytic cleavages from the ␤-amyloid precursor protein (␤PP) (Kang et al., 1987). Constitutive cleavage of ␤PP by "␣-secretase" occurring in vivo and in vitro results in the secretion of the N-terminal ectodomain ␤PP S . The ubiquitous ␣-secretase cleavage occurs within the A␤ region and therefore precludes the generation of an intact A␤ peptide . In contrast to most non-neural cells, neurons and glia secrete relatively small amounts of ␤PP S for unclear reasons (Haass et al., 1991). Some ␤PP molecules not cleaved on the cell surface are internalized and targeted to endosomes and lysosomes. A smaller population of ␤PP molecules seems to be recycled back to the surface for secretion or internalization . Both the secretory and endocytic pathways have been shown to contribute to A␤ released into medium by cultured cells, although endocytic processing seems to be the predominant source (Koo and Squazzo, 1994).The physiological role of ␤PP remains to be defined. Proposed functions for ␤PP S include neurite-promoting properties, wound healing, cell adhesion, cell growth and diff...

Amyloid beta1-42 peptide regulates microtubule stability independently of tau

“…Treatment of primary cortical neurons with Ab results in FAK activation (Williamson et al, 2002), and aberrant activation of focal adhesion proteins mediates Ab-induced neuronal dystrophy (Grace et al, 2002;Williamson et al, 2002). In addition, APP, a potential receptor for Ab (Lorenzo et al, 2000), is found at focal contacts (Sabo et al, 2001); it colocalizes with b1 integrins in neuronal cells (Yamazaki et al, 1997), and antibody-mediated clustering of APP induces significant loss of dendritic spines (Lefort et al, 2012) and neuronal injury through FAK activation (Xu et al, 2009). We found that clustering of APP was sufficient to induce Glu MTs and both APP and caspase-2 expression were required for the formation of stable Glu MTs by Ab, suggesting that induction of MT stability by Ab is initiated by the same APP/ caspase-2/RhoA pathway that leads to neurotoxicity by Ab in neurons (Lefort et al, 2012;Pozueta et al, 2013;Troy et al, 2000).…”

“…Given the direct involvement of APP with both cell adhesion and neuronal injury by Ab (Sabo et al, 2001;Xu et al, 2009;Yamazaki et al, 1997), we tested whether stimulation of APP signaling was also required to induce stable Glu MTs by Ab. Interestingly, downregulation of APP expression prior to Ab treatment completely abrogated the induction of stable Glu MTs by Ab (Fig.…”

Phosphorylation of Actin-Depolymerizing Factor/Cofilin by LIM-Kinase Mediates Amyloid -Induced Degeneration: A Potential Mechanism of Neuronal Dystrophy in Alzheimer's Disease

“…Several membrane proteins have been shown to participate in A␤-induced apoptosis ; in contrast, A␤PP and/or integrins could be the main cell-surface candidates for A␤ fibril-induced neuronal dystrophy. Both A␤PP and integrins participate in mechanisms of neuronal plasticity (Breen et al, 1991;Perez et al, 1997;Benson et al, 2000;Sabo et al 2001a), bind to A␤ fibrils (Sabo et al, 1995;Lorenzo et al, 2000;Van Nostrand et al, 2002), colocalize in adhesion sites (Storey et al, 1996;Yamazaki et al, 1997), and cluster around A␤ deposits in aberrant focal adhesion structures (Grace and Busciglio, 2003;Heredia et al, 2004). In fact, A␤ dystrophy requires aberrant activation of focal adhesion proteins, including Paxillin (Grace and Busciglio, 2003), activation of which could also depend on A␤PP or integrins, through their interaction with different cytoplasmic proteins (Sabo et al, 2001b;Zambrano et al, 2001;Martin et al, 2002).…”

Integrins as a distinct subtype of dependence receptors

“…4 However, integrin-mediated death is similar to apoptosis induced by the b-amyloid precursor protein (APP), a dependence receptor which initiates apoptosis via activation of caspase 8 69 and also happens to associate with integrins at sites of ECM contact. 70 Interestingly, substrate-ECM contact may also prevent apoptosis induced by APP.…”