A functional screen uncovers circular RNAs regulating excitatory synaptogenesis in hippocampal neurons

Circular RNAs (circRNAs) are an expanding class of largely unexplored RNAs which are prominently enriched in the mammalian brain. Here, we systematically interrogate their role in excitatory synaptogenesis of rat hippocampal neurons using RNA interference. Thereby, we identify seven circRNAs as negative regulators of excitatory synapse formation, many of which contain high-affinity microRNA binding sites. Knockdown of one of these candidates, circRERE, promotes the formation of electrophysiologically silent synapses. Mechanistically, circRERE knockdown results in a preferential upregulation of synaptic mRNAs containing binding sites for miR-128-3p. Overexpression of circRERE stabilizes miR-128-3p and rescues exaggerated synapse formation upon circRERE knockdown in a miR-128-3p binding site-specific manner. Overall, our results uncover circRERE-mediated stabilization of miR-128-3p as a means to restrict the formation of silent excitatory synaptic co-clusters and more generally implicate circRNA-dependent microRNA regulation in the control of synapse development and function.