Project Summary Spinal muscular atrophy (SMA) is an inherited neurodegenerative disease characterized by motor neuron loss and skeletal muscle atrophy. SMA is caused by reduced levels of the SMN protein due to homozygous mutations in the SMN1 gene and is the leading genetic cause of infant mortality. Studies in mouse models revealed that SMA pathology involves dysfunction of sensory-motor circuits comprising multiple neuron types and possibly non-neuronal cells. Beyond motor neuron death, dysfunction and loss of neuromuscular junctions (NMJs) as well as central synapses from proprioceptive sensory neurons are established early drivers of motor system pathology in SMA. Mechanistically, the SMN protein functions in the assembly of small nuclear ribonucleoproteins (snRNPs) that mediate pre-mRNA splicing and 3’-end processing of histone mRNAs; and our previous studies directly linked dysregulation of these RNA pathways to SMA pathology. Importantly, three different SMN-inducing therapies have been approved for SMA treatment which prevent early death and slow progression depending on disease severity and time of intervention. However, these therapies alone do not provide a cure and significant motor deficits persist in SMA patients, especially those treated late. Therefore, incomplete correction of disease symptoms combined with variability in the clinical response to treatment represent urgent unmet needs of SMA patients. In this context, it remains essential to further our understanding of SMA disease mechanisms as a means to identify novel therapeutic targets and to develop new pharmacological approaches that can enhance the clinical benefit of SMN-inducing drugs through combinatorial treatment. Here, we propose a multidisciplinary research project that addresses these outstanding issues by building on published studies and preliminary data gathered during the previous award period. In Aim 1, we will establish the conservation of RNA-mediated mechanisms of synaptic pathology across mouse models of SMA with varying disease severity as a necessary step to determine their relevance to the human disease. We will employ validated means for selective restoration of individual RNA pathways and downstream targets of SMN deficiency by AAV9-mediated gene delivery to link the loss of proprioceptive synapses and NMJ denervation, respectively, to disruption of U12 splicing and U7-mediated histone mRNA processing in both severe and milder models of SMA. In Aim 2, guided by our hypothesis that dysregulation of the glutamatergic system contributes to SMA pathology, we will determine the therapeutic potential of a novel disease-modifying pharmacological approach aimed at stimulating glutamatergic neurotransmission and improving sensory-motor circuit activity in mouse models of SMA. This will be tested either alone or together with SMN-inducing drugs to highlight synergistic effects relevant for combinatorial treatment of SMA. In Aim 3, we will investigate intrinsic...