Opto-CLIP reveals dynamic FMRP regulation of mRNAs upon CA1 neuronal activation

Neuronal diversity and function are intricately linked to the dynamic regulation of RNA metabolism, including splicing, localization, and translation. Electrophysiologic studies of synaptic plasticity, models for learning and memory, are disrupted in Fragile X Syndrome (FXS). FXS is characterized by the loss of FMRP, an RNA-binding protein (RBP) known to bind and suppress translation of specific neuronal RNAs. Since molecular studies have demonstrated that synaptic plasticity in CA1 excitatory hippocampal neurons is protein-synthesis dependent, together these observations have suggested a potential role for FMRP in synaptic plasticity in FXS. To explore this model, we developed a new experimental platform, Opto-CLIP, to integrate optogenetics with cell-type specific FMRP CLIP and RiboTag in CA1 hippocampal neurons, allowing investigation of FMRP-regulated dynamics after neuronal activation. We tracked changes in FMRP binding and ribosome-associated RNA profiles 30 minutes after neuronal activation. Our findings reveal a significant reduction in FMRP-RNA binding to transcripts encoding nuclear proteins, suggesting FMRP translational inhibition may be de-repressed to allow rapid translational responses required for neuronal homeostasis. In contrast, FMRP binding to transcripts encoding synaptic targets were generally stable after activation, but all categories of targets demonstrated variability in FMRP translational control. Opto-CLIP revealed differential regulation of subsets of transcripts within CA1 neurons rapidly after depolarization, and offers promise as a generally useful platform to uncover mechanisms of RBP-mediated RNA regulation in the context of synaptic plasticity.