Arc In AMPA Receptor Endocytosis: Direct Evidence Indicating Arc As A Scaffolding Protein

Abstract

Long-term potentiation (LTP) and long-term depression (LTD) are processes thought to underlie learning and memory, which require proper regulation of activity-regulated cytoskeletonassociated protein (Arc; also known as Arg3.1). Abnormal expression levels of Arc have been associated with Alzheimer’s disease, schizophrenia, Fragile-X syndrome, Angelmann’s syndrome and substance abuse. Arc is an immediate early gene product that is transcribed in dendritic spines and is a positive regulator of AMPA receptor (AMPAR) endocytosis during LTD. Verified protein-protein interactions between Arc and proteins involved in endocytosis, including dynamin and endophilin, support the role of Arc as a regulator of AMPAR endocytosis. Nevertheless, the mechanism by which Arc specifically targets AMPARs for endocytosis is currently unknown. This is key to understanding the mechanisms of learning and memory and how they are affected by Arc associated human conditions. Here we show evidence of a novel interaction between Arc and protein interacting with C kinase 1 (PICK1), a protein known to bind to the GluR2 subunit of AMPARs and associated with AMPAR trafficking. Cross-correlation raster image correlation spectroscopy and Förster resonance energy transfer (FRET) with Arc-mCherry and EGFP-PICK1 demonstrate this interaction. FRET is more apparent in the projections of transfected SH-SY5Y cells and is enhanced by depolarization. Interestingly, TIRF imaging shows PICK1 aggregates that are only present when cotransfected with Arc that colocalize with Arc aggregates. These findings show an interaction of Arc and PICK1, which may answer how Arc directs endocytic machinery to AMPARs during LTD.