Endocytosis of synaptic ADAM10 in neuronal plasticity and Alzheimer’s disease
Elena Marcello, … , Fabrizio Gardoni, Monica Di Luca
Elena Marcello, … , Fabrizio Gardoni, Monica Di Luca
Published May 8, 2013
Citation Information: J Clin Invest. 2013;123(6):2523-2538. https://doi.org/10.1172/JCI65401.
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Research Article Neuroscience

Endocytosis of synaptic ADAM10 in neuronal plasticity and Alzheimer’s disease

Abstract

A disintegrin and metalloproteinase 10 (ADAM10), a disintegrin and metalloproteinase that resides in the postsynaptic densities (PSDs) of excitatory synapses, has previously been shown to limit β-amyloid peptide (Aβ) formation in Alzheimer’s disease (AD). ADAM10 also plays a critical role in regulating functional membrane proteins at the synapse. Using human hippocampal homogenates, we found that ADAM10 removal from the plasma membrane was mediated by clathrin-dependent endocytosis. Additionally, we identified the clathrin adaptor AP2 as an interacting partner of a previously uncharacterized atypical binding motif in the ADAM10 C-terminal domain. This domain was required for ADAM10 endocytosis and modulation of its plasma membrane levels. We found that the ADAM10/AP2 association was increased in the hippocampi of AD patients compared with healthy controls. Long-term potentiation (LTP) in hippocampal neuronal cultures induced ADAM10 endocytosis through AP2 association and decreased surface ADAM10 levels and activity. Conversely, long-term depression (LTD) promoted ADAM10 synaptic membrane insertion and stimulated its activity. ADAM10 interaction with the synapse-associated protein-97 (SAP97) was necessary for LTD-induced ADAM10 trafficking and required for LTD maintenance and LTD-induced changes in spine morphogenesis. These data identify and characterize a mechanism controlling ADAM10 localization and activity at excitatory synapses that is relevant to AD pathogenesis.

Authors

Elena Marcello, Claudia Saraceno, Stefano Musardo, Hugo Vara, Alerie Guzman de la Fuente, Silvia Pelucchi, Daniele Di Marino, Barbara Borroni, Anna Tramontano, Isabel Pérez-Otaño, Alessandro Padovani, Maurizio Giustetto, Fabrizio Gardoni, Monica Di Luca

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Figure 2

AP2 interacts with 735RQR737 motif of ADAM10 tail.

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AP2 interacts with 735RQR737 motif of ADAM10 tail. (A) Mouse brain hom...
(A) Mouse brain homogenate was IP with ADAM10 antibody. AP2 subunits coprecipitate with ADAM10. IP ADAM10 was detected by WB. No signal is detectable when the sample is precipitated without ADAM10 antibody. (B) Pulldown assays performed with GST-ADAM10 Ct and GST from mouse brain homogenate. α-adaptin and μ2, but not γ-adaptin and ε-adaptin, are precipitated by ADAM10 Ct. (C) aa sequences of ADAM10 Ct deletion mutants (747Δ, 741Δ, 737Δ, 734Δ, 721Δ). (D) Pulldown assays performed with GST-ADAM10 Ct, 734Δ, and 721Δ mutants. Deletion of the last 15 aa abolishes AP2 subunit binding, whereas SAP97 interacts with the membrane proximal 27 aa of ADAM10 tail. (E) Pulldown assays carried out with all deletion mutants. The 735RQR737 motif is necessary for α-adaptin precipitation. (F) Pulldown assays performed with GST-ADAM10 Ct bearing mutations of 735RQR737 motif. No change in α-adaptin binding was detected. (G) Pulldown assays performed with GST-ADAM10 Ct mutated 735RQR737 to AQA. The mutation of both arginine residues to alanine abolishes α-adaptin binding, but does not affect SAP97 association. (H) Representative images of ADAM10 membrane staining (red) and α-adaptin (green) of COS7 cells expressing ADAM10 WT or ADAM10 AQA. Merged images are shown on the right. Scale bar: 20 μm. (I) Quantification of ADAM10 WT/α-adaptin and ADAM10 AQA/α-adaptin colocalization of experiments in H (*P < 0.05, ADAM10 WT versus ADAM10 AQA, n = 2 experiments, 12 cells per condition). In A and E–G, lanes were run on the same gel but were not contiguous.