Background: The prevalence of autism spectrum disorders (ASD) has risen in the past three decades such that 1 in 68 children are diagnosed with ASD. New genetic technologies have implicated multiple genes in ASD pathogenesis. Many of these genes produce proteins that function at the post-synaptic density (PSD). One central protein of the PSD is SHANK3, which serves as a scaffold, linking transmembrane proteins of the PSD such as NMDA receptors and neuroligins with the underlying cytoskeleton. As such, SHANK3 is critical for organization and function of the synapse. One percent of all children with ASD have loss of function mutations in SHANK3, but another 15% of individuals with ASD have epigenetic dysregulation of the SHANK3 gene. Thus, therapies to normalize SHANK3 abundance in the PSD could be beneficial for hundreds of thousands of individuals with ASD.Hypothesis/Objective: We hypothesize that we can increase the abundance of SHANK3 in the post-synaptic density by inhibiting molecular pathways that normally promote post-translational degradation of SHANK3. We will identify these molecular pathways using an unbiased cell-based screen.Specific Aims:Aim 1: Perform a forward genetic screen for modifiers of SHANK3 stability. Aim 1A: Complete primary cell based screen of druggable genome. Aim 1B: Mapping network/pathways of SHANK3 modifiers and their interactions with SHANK3. Aim 2: Validate proteins that regulate SHANK3 in neurons and mouse models of SHANKopathies. Study Design: We will begin by using an unbiased cell-based screen to identify genes and molecular pathways that regulate the abundance of SHANK3 through post-translational mechanisms. As any screen of this type can be complicated by false positive hits, we will validate candidates from the primary screen in a second cell line. Those candidates passing both the primary and secondary screens will be prioritized bioinformatically and then tested in cultured neurons. Finally, our best candidates that pass all three levels of our screen will be tested for rescue of SHANK3 haploinsufficiency phenotype in vivo in a mouse model of ASD.Impact: Perhaps as many as 500,000 individuals in the United States with ASD have either mutation or dysregulation of SHANK3. This work could potentially lead to targeted therapies for these individuals. Moreover, this approach could serve as a blueprint for other neurodevelopmental disorders due to haploinsuffiency.Innovation: This approach is innovative in two ways. First, instead of focusing on downstream molecular effects of a syndromic autism, we are focusing on the upstream regulation of SHANK3. Any therapy we employ to increase SHANK3 stability will improve all downstream dysregulation not just a subset. Second, our approach uses a genome-wide screen rather than cherry-picking molecular pathways based on our incomplete understanding of the synapse and thus is unbiased.
Interagency Autism Coordinating Committee (IACC)
Autism Research Database (AFD)
