Abstract
Activity-regulated cytoskeleton-associated protein, Arc, is a major regulator of long-term synaptic plasticity and memory formation. Here we reveal a novel interaction partner of Arc, a resident endoplasmic reticulum transmembrane protein, calnexin. We show an interaction between recombinantly-expressed GST-tagged Arc and endogenous calnexin in HEK293, SH-SY5Y neuroblastoma and PC12 cells. The interaction was dependent on the central linker region of the Arc protein that is also required for endocytosis of AMPA-type glutamate receptors. High-resolution proximity-ligation assays (PLAs) demonstrate molecular proximity of endogenous Arc with the cytosolic C-terminus, but not the lumenal N-terminus of calnexin. In hippocampal neuronal cultures treated with brain-derived neurotrophic factor (BDNF), Arc interacted with calnexin in the perinuclear cytoplasm and dendritic shaft. Arc also interacted with C-terminal calnexin in the adult rat dentate gyrus (DG). After induction of long-term potentiation (LTP) in the perforant path projection to the DG of adult anesthetized rats, enhanced interaction between Arc and calnexin was obtained in the dentate granule cell layer (GCL). Although Arc and calnexin are both implicated in the regulation of receptor endocytosis, no modulation of endocytosis was detected in transferrin uptake assays. Previous work showed that Arc interacts with multiple protein partners to regulate synaptic transmission and nuclear signaling. The identification of calnexin as a binding partner further supports the role of Arc as a hub protein and extends the range of Arc function to the endoplasmic reticulum, though the function of the Arc/calnexin interaction remains to be defined.
Highlights
Activity-regulated cytoskeleton-associated protein (Arc) has been established as a major regulator of protein synthesis-dependent synaptic plasticity (Bramham et al, 2010; Korb and Finkbeiner, 2011; Shepherd and Bear, 2011; Nikolaienko et al, 2017a)
Using a GST-based pull-down assay and mass spectrometry, we found that Arc interacts with the cytosolic domain of the transmembrane endoplasmic reticulum (ER) protein, calnexin
The band was identified as a transmembrane protein of the endoplasmic reticulum, calnexin (Peptide Identification Score: 142; Peptides identified: APVPTGEVYFADSFDR and KIPNPDFFEDLEPFR)
Summary
Activity-regulated cytoskeleton-associated protein (Arc) has been established as a major regulator of protein synthesis-dependent synaptic plasticity (Bramham et al, 2010; Korb and Finkbeiner, 2011; Shepherd and Bear, 2011; Nikolaienko et al, 2017a). Arc synthesis is essential for long-term forms of synaptic plasticity such as long-term potentiation (LTP), long-term depression (LTD; Park et al, 2008; Waung et al, 2008) and homeostatic scaling (Shepherd et al, 2006). These Arc-dependent changes in synaptic efficacy are thought to be important. Arc can regulate synaptic strength at the level of gene expression by localizing to nuclear promyelocytic leukemia bodies and controlling expression of the AMPA-type glutamate receptors (Bloomer et al, 2007; Korb et al, 2013)
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