Deficits in proper synapse formation, elimination, and maintenance, are hypothesized to underlie numerous neurocognitive disorders, including mental retardation and autism, and are also thought to contribute to behaviors associated with drug addiction. Neuronal activity promotes synapse elimination to prune excess synapses formed during development, and to homeostatically maintain a steady-state number of synaptic connections. The transcription factor myocyte enhancer factor 2 (MEF2) is a critical activity-dependent regulator of excitatory synapse elimination in developing and adult brains. However, the molecular and cellular mechanisms by which MEF2 controls synapse number are not well understood. Our lab recently discovered that the RNA-binding protein, Fragile X Mental Retardation Protein (FMRP) is required for MEF2-induced elimination of synapses. Fragile X Syndrome (FXS), the most prevalent inherited form of autism, results from inactivation of the Fmr1 gene, which codes for FMRP. Individuals with FXS and a mouse model for the disorder, Fmr1 knockout mice, display increased cortical dendritic spine density, indicating that deficits in synapse elimination may underlie FXS. We propose that MEF2 and FMRP function together to regulate common transcripts to induce elimination of excitatory synapses and propose Specific Aim 1 to investigate this hypothesis. In this aim, we will utilize CLIP-seq, a technique pairing cross-linking immunoprecipitation (CLIP) followed by high throughput sequencing to identify MEF2-regulated gene targets whose transcripts associate with FMRP. We will then test a requirement for MEF2-regulated and FMRP-associated candidates in structural and functional synapse elimination by performing 2-photon live-cell dendritic spine imaging and whole-cell patch clamp recordings of mEPSCs.
In Specific Aim 2, we will characterize the process of MEF2-dependent synapse elimination, examining the kinetics of MEF2-induced structural and functional synapse elimination, as well as the morphological process of MEF2-mediated structural synapse elimination by performing 2-photon microscopy and whole-cell electrophysiology recordings. As synapse elimination defects may contribute to autism, Fragile X Syndrome, and behavioral adaptations associated with drug abuse, we believe that our studies will provide significant implications towards understanding these and related disorders.
The transcription factor myocyte enhancer factor 2 (MEF2) has critical roles in the proper development and maintenance of synaptic connections, in learning and memory, and in behaviors associated with drug addiction;however, the molecular and cellular mechanisms by which MEF2 mediates these effects are not well understood. Here, we propose to investigate the mechanisms by which MEF2 regulates the formation of proper connections in the brain, a study which will have broad implications for understanding autism, Fragile X Syndrome, drug addiction, and related disorders.
