The Amyloid Precursor Protein (APP) of Alzheimer Disease and Its Paralog, APLP2, Modulate the Cu/Zn-Nitric Oxide-catalyzed Degradation of Glypican-1 Heparan Sulfate in Vivo

Abstract

Processing of the recycling proteoglycan glypican-1 involves the release of its heparan sulfate chains by copper ion-and nitric oxide-catalyzed ascorbate-triggered autodegradation. The Alzheimer disease amyloid precursor protein (APP) and its paralogue, the amyloid precursor-like protein 2 (APLP2), contain copper ion-, zinc ion-, and heparan sulfate-binding domains. We have investigated the possibility that APP and APLP2 regulate glypican-1 processing during endocytosis and recycling. By using cell-free biochemical experiments, confocal laser immunofluorescence microscopy, and flow cytometry of tissues and cells from wild-type and knock-out mice, we find that (a) APP and glypican-1 colocalize in perinuclear compartments of neuroblastoma cells, (b) ascorbate-triggered nitric oxidecatalyzed glypican-1 autodegradation is zinc ion-dependent in the same cells, (c) in cell-free experiments, APP but not APLP2 stimulates glypican-1 autodegradation in the presence of both Cu(II) and Zn(II) ions, whereas the Cu(I) form of APP and the Cu(II) and Cu(I) forms of APLP2 inhibit autodegradation, (d) in primary cortical neurons from APP or APLP2 knock-out mice, there is an increased nitric oxide-catalyzed degradation of heparan sulfate compared with brain tissue and neurons from wild-type mice, and (e) in growth-quiescent fibroblasts from APLP2 knock-out mice, but not from APP knock-out mice, there is also an increased heparan sulfate degradation. We propose that the rate of autoprocessing of glypican-1 is modulated by APP and APLP2 in neurons and by APLP2 in fibroblasts. These observation identify a functional relationship between the heparan sulfate and copper ion binding activities of APP/APLP2 in their modulation of the nitroxyl anion-catalyzed heparan sulfate degradation in glypican-1. Processing of the amyloid precursor protein (APP)1 of Alzheimer disease (AD) involves several proteases and regulatory proteins, collectively designated ␣-, ␤-, and ␥-secretases (Fig. 1). ␤-and ␥-Cleavages lead to the generation of amyloid-␤ (A␤) peptides A␤1-40 and A␤1-42 (1, 2). The A␤ peptides are believed to cause the neurotoxicity associated with AD via an increase in A␤ levels (3). Eventually, A␤ accumulates into amyloid fibrils and finally senile plaques, the key pathological hallmark of AD. However, the APP paralogue amyloid precursor-like protein 2 (APLP2) does not contain the A␤ sequence and hence does not contribute to amyloid formation. Both APP and APLP2 bind metal ions and heparin, but the physiological significance of this property is unclear. Nevertheless, several in vitro studies have shown that APP and its ␣-secretory form are involved in neuronal growth and survival and in neurite outgrowth (for reviews, see Refs. 4 -7).The primary transcript of the APP gene undergoes extensive alternative splicing. The isoform APP695 is expressed at high levels in central nervous system neurons (8). APP-deficient mice (APP Ϫ/Ϫ ) show reactive gliosis, especially in the cerebral cortex and the hippocampus, and decreased locomotor activity ...

Suppression of Amyloid β A11 Antibody Immunoreactivity by Vitamin C

“…Brain tissue from non-demented controls and AD patients was obtained from the Victorian Brain Bank Network. Embryonic fibroblasts from wild-type and Tg2576 mice were prepared and maintained as described elsewhere (38). Synthetic A␤42 was purchased from Millipore.…”

“…Free copper ions are scarce in vivo, but Cu(II)-loaded cuproproteins, such as the glycosylphosphatidylinositol-anchored ceruloplasmin (36) and prion proteins (34,37) as well as APP, can support S-nitrosylation of Gpc-1. Moreover, APP and Gpc-1 colocalize in subcellular compartments of neuroblastoma cells (38).…”

“…Because APP interacts with Gpc-1 and modulates the copper-and NO-dependent release of HS from Gpc-1 both in vitro and in vivo (38), we decided to investigate whether anMancontaining HS degradation products generated by Gpc-1 autoprocessing interact with APP degradation products and whether such HS is ultimately deposited in amyloid plaques. For this purpose, we examined normal human and AD brains as well as brains and/or fibroblasts from wild-type, Tg2576, and 3xTg-AD mice for anMan-and A␤-immunoreactive components.…”

Novel aspects of vitamin C: how important is glypican-1 recycling?

“…Free Zn(II) ions, which are particularly abundant in neural cells, can block Cu(II) binding to glypican-1 and thereby attenuate S-nitrosylation [33][34][35][36]. We have identified three copperbinding proteins that can accomodate the Cu(II)-to-Cu(I) reduction that is required to support formation of glypican-1-SNO in the presence of free Zn(II) ions.…”

“…We have identified three copperbinding proteins that can accomodate the Cu(II)-to-Cu(I) reduction that is required to support formation of glypican-1-SNO in the presence of free Zn(II) ions. These are the GPI-linked cellular prion protein [34,37], the GPI-linked, brain-specific splice variant of ceruloplasmin 8 [35] and the amyloid precursor protein of Alzheimer´s disease [36], all of which may thus be regarded as catalysts of S-nitrosylation [26].…”