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The remarkable capacity of neurons to reorganize their structure, function, and connections in an activity-dependent manner is supported by an extensive network of molecules and effectors. Among those, Activity-regulated cytoskeleton-associated protein (Arc, also known as Arg3.1) is considered one of the central, most versatile, players. Arc is an immediate early gene product that support neuroplastic changes important for cognitive function and memory formation, while also oligomerizing to form capsids that can be released and can move independent of the synapse. Although a lot has been uncovered about the contributions of Arc to neuron biology and behavior, very little is known about how different functions of Arc are regulated both temporally and spatially in neurons. To answer to this question, the regulation of Arc expression must be monitored at various levels. This thesis analyzes how Arc can be transcriptionally and post-translationally regulated and how this confers neurobiological specificity. Being an immediate early gene, transcription of Arc needs to act rapidly in response to a neuronal stimulus, such as BDNF. The first aim of this thesis provides a model to describe how the Arc promoter can overcome topological constraints to facilitate rapid and persistent transcription, up to 6 hours post-BDNF exposure. Using chromatin analysis and biochemical techniques we have mapped regions of the Arc promoter that undergo double stranded DNA breaks to facilitate rapid transcription, and that BDNF-induced Arc non-coding RNA elements interact with DNA repair proteins such as Brd4 to effectively end transcription. The second aim of this thesis aims to determine how phosphorylation can spatiotemporally dictate the function of Arc. Mass spectrometry and sequence prediction strategies were used to map novel Arc phosphorylation sites. This approach led us to recognize S67 and T278 as residues that can be modified by TNIK, which is a kinase abundantly expressed in neurons and been implicated as a risk factor for psychiatric disorders. Characterization of TNIK-dependant phosphorylation at these two sites can strongly influence Arc's subcellular distribution and self-assembly as capsids. The multifaceted nature of Arc’s neurobiology ranges from being a key component of long-term memory consolidation to participating in viral capsid intracellular communication. This thesis provides insight into Arc’s moonlighting capabilities both at the transcription and post-translational levels.

Activity-regulated cytoskeleton-associated protein (Arc) is an immediate early gene product that support neuroplastic changes important for cognitive function and memory formation. As a protein with homology to the retroviral Gag protein, a particular characteristic of Arc is its capacity to self-assemble into virus-like capsids that can package mRNAs and transfer those transcripts to other cells. Although a lot has been uncovered about the contributions of Arc to neuron biology and behavior, very little is known about how different functions of Arc are coordinately regulated both temporally and spatially in neurons. The answer to this question we hypothesized must involve the occurrence of different protein post-translational modifications acting to confer specificity. In this study, we used mass spectrometry and sequence prediction strategies to map novel Arc phosphorylation sites. Our approach led us to recognize serine 67 (S67) and threonine 278 (T278) as residues that can be modified by TNIK, which is a kinase abundantly expressed in neurons that shares many functional overlaps with Arc and has, along with its interacting proteins such as the NMDA receptor, and been implicated as a risk factor for psychiatric disorders. Furthermore, characterization of each residue using site-directed mutagenesis to create S67 and T278 mutant variants revealed that TNIK action at those amino acids can strongly influence Arc's subcellular distribution and self-assembly as capsids. Together, our findings reveal an unsuspected connection between Arc and TNIK. Better understanding of the interplay between these two proteins in neuronal cells could lead to new insights about apparition and progression of psychiatric disorders. Cover Image for this issue: https://doi.org/10.1111/jnc.15077.