Investigating the synaptic pathology of Autism

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PROJECT SUMMARY Genetic mutations that confer autism (ASD) risk often occur in genes that comprise signal transduction networks that link synaptic transmission to downstream changes in gene expression. However, the dynamic, network- scale behavior of these complex and interconnected signaling networks in normal or disease states is poorly understood. This is the first renewal application of a highly productive R01 grant that built a quantitative multiplex co-immunoprecipitation or QMI panel to study the dynamic activity of a 20-member protein interaction network (PIN), consisting of glutamate receptors, scaffolds, and signal transduction molecules; mutations in the genes encoding all target proteins have been genetically linked to autism. Using QMI, we discovered that this PIN encodes information by varying the composition and intensity of modules of coordinated interactions in response to incoming signals. Moreover, we found that mutations that contribute to autism risk disrupt synaptic PINs by causing them to assume a network state that resembles the state of a wildtype neuron that has undergone homeostatic scaling. This results in a reduced dynamic range of the network to change in response to subsequent stimuli, and leads to a systems-level disturbance in basal glutamate tone (as reflected in disrupted E/I balance). In the second cycle we focus on the question of, can we normalize ASD PINs, and will this normalization correlate with functional rescue of phenotypes? In Aim 1, we focus on normalization via FYN kinase, which we identified as a dysregulated network hub in FMR1-/y mice downstream of synaptic plasticity inputs. Preliminary data demonstrate that inhibition of hyperactive FYN signaling normalizes hyperactive protein synthesis and behavior; we propose to perform an extensive battery of molecular, cellular and behavioral assays to investigate the potential of FYN inhibition to treat the phenotypes of FMR1 deficiency. In Aim 2, we extend our network-scale analysis to a second PIN critical to synaptic plasticity, the mTOR network. We use pharmacological and genetic inhibition of mTOR to model information flow through the mTOR PIN during synaptic plasticity, and to establish which components of the pathway are required for homeostatic scaling, in vivo or in vitro. In Aim 3, we focus on PIN normalization by manipulating synaptic activity. We attempt to `un- scale' the synaptic PIN of ASD-mutation-carrying mice, and measure if this treatment is able to restore normal PIN activity and the ability of the neuron to undergo normal homeostatic scaling. Critically, this last experiment will reveal whether altered levels of neuronal activity downstream of developmental mechanisms cause disrupted synaptic and mTOR signal transduction, or conversely if ongoing deficits in signal transduction are independent, or even causative, of altered basal activity levels. Overall, this renewal would continue our investigations into the molecular network mechanisms by which ASD risk genes disrupt synaptic signal transduction.