The critical and diverse roles of microRNAs in the brain are continuing to be discovered. To date, microRNAs have been implicated in neurogenesis, neurodevelopment, synaptic plasticity, and neuropsychiatric disorders. To fully understand how microRNAs carry out these functions, both in normal and pathological states, their mRNA targets must be identified and characterized. This project takes advantage of the RISC-trap assay, an excellent tool to address the roles of microRNAs in neurons. RISC-trap utilizes a dominant negative component of the RNAi-Induced Silencing Complex (RISC) - a central component in the microRNA pathway that is the bridge between microRNAs and their targets- to identify targets of individual microRNAs. This project will use RISC-trap to identify a comprehensive set of mRNAs targeted by miR-132, involved in neuronal maturation, mature function, and disease states, and miR-124, demonstrated to promote neural differentiation and recently shown to regulate plasticity. Neuron-specific microRNA targets may be in one of two categories. They may encode transcripts, such as synaptic components, that have neural-specific functions. Alternatively, and of particular interest, will be transcripts that are expressd in both neurons and HEK cells, but where the miR-132 or miR-124 interactions occur exclusively in neurons-these interactions may require the activities of specific RNA-binding proteins. Identifying targets specific to neurons will advance understanding of the mechanisms used by microRNAs in neurons. This global target-identification approach will be complemented by examining the regulation of a novel miR-132 target, ARHGEF11, which was identified in RISC-trap screen of HEK cells, and is also known to contribute to neuronal cell signaling. This PDZ-containing protein is interesting because it has been shown to activate Rho signaling and suppress neurite outgrowth. Additionally, ARHGEF11, like miR-132, has been linked to schizophrenia. This project takes advantage of both miR-132 knockout mice and conditional knockout mice, which provide the ability to target excision of the miR-132 locus in the adult newborn neurons of the conditional knockout mouse to examine how ARHGEF11 is regulated by miR-132 in these cells. A thorough investigation miR-132 and miR-124 targets will lead to a deeper understanding of the mechanism of action of these microRNAs and may reveal drug targets for neurodevelopmental disorders and potentially disorders of the adult nervous system.
Neural development, including targeting, dendritic arborization, and synaptogenesis, is critical for the proper functioning of the adult brain. Understanding the molecular mechanisms that allow these processes to occur properly is necessary to assess what is malfunctioning when development is abnormal and neurological disorders arise. Thus, a thorough investigation of the molecules involved in post-mitotic neural development and neuronal function, including miR-132 and its targets, may reveal drug targets for neurodevelopmental disorders and potentially disorders of the adult nervous system.
