Structure of monomeric full-length ARC sheds light on molecular flexibility, protein interactions, and functional modalities.

The activity-regulated cytoskeleton-associated protein (ARC) is critical for long-term synaptic plasticity and memory formation. Acting as a protein interaction hub, ARC regulates diverse signalling events in postsynaptic neurons. A protein interaction site is present in the ARC C-terminal domain (CTD), a bilobar structure homologous to the retroviral Gag capsid domain. We hypothesized that detailed knowledge of the three-dimensional molecular structure of monomeric full-length ARC is crucial to understand its function; therefore, we set out to determine the structure of ARC to understand its various functional modalities. We purified recombinant ARC and analyzed its structure using small-angle X-ray scattering and synchrotron radiation circular dichroism spectroscopy. Monomeric full-length ARC has a compact, closed structure, in which the oppositely charged N-terminal domain (NTD) and CTD are juxtaposed, and the flexible linker between them is not extended. The modeled structure of ARC is supported by intramolecular live-cell Förster resonance energy transfer imaging in rat hippocampal slices. Peptides from several postsynaptic proteins, including stargazin, bind to the N-lobe, but not to the C-lobe, of the bilobar CTD. This interaction does not induce large-scale conformational changes in the CTD or flanking unfolded regions. The ARC NTD contains long helices, predicted to form an anti-parallel coiled coil; binding of ARC to phospholipid membranes requires the NTD. Our data support a role for the ARC NTD in oligomerization as well as lipid membrane binding. The findings have important implications for the structural organization of ARC with respect to distinct functions, such as postsynaptic signal transduction and virus-like capsid formation. Open Practices Open Science: This manuscript was awarded with the Open Materials Badge. For more information see: https://cos.io/our-services/open-science-badges/.