Spinal muscular atrophy (SMA) is the leading genetic cause of infant mortality and the second most common autosomal recessive genetic disorder worldwide. Characterized by the gradual loss of motor neurons and a denervation of muscles, SMA is directly caused by a deficiency of survival of motor neuron protein (SMN). Low levels of SMN often results in progressive muscle weakness and eventual death due to respiratory distress. SMN and its associated complex proteins, function to assemble spliceosomal small nuclear ribonucleoprotein (snRNP) complexes. Accordingly, reduction of SMN impairs snRNP assembly, producing splicing defects observed in SMA patients and animal models. However, research examining splicing defects has been unable to resolve the selective vulnerability of spinal motor neurons in SMA patients. SMN localizes to axons and dendrites in spinal motor neurons. In the axon, SMN colocalizes and co-transports with multiple mRNA binding proteins that direct axonal mRNA transport. Preliminary data from our lab using a severe SMA mouse model demonstrate an axonal mRNA localization defect in these cultured spinal motor neurons, without effects on the steady state levels of mRNAs. Given the importance of mRNA transport in facilitating localized mRNA translation needed for axonal function, we predict that this defect in mRNA localization is present in SMA motor neurons in vivo and contributes to disease progression. We hypothesize that SMN mediates axonal localization of mRNAs required for growth and maintenance of the axon, through the assembly and/or transport of mRNA and associated protein (mRNP) granules. We will first interrogate the role of SMN in mRNA processing and localization.
In specific aim one, we will employ motor-neuron specific analysis of the ribosome-associated ?translatome? to examine mRNA regulation in axons and cell bodies of a well-characterized mouse model of SMA. Identification of highly altered transcripts will expand the list of known mislocalized axonal mRNAs in SMA motor neurons, and will be further explored using in situ approaches.
In specific aim two, we will use immunofluorescence and RNA fluorescence in situ hybridization to evaluate the axonal localization of specific mRNA binding proteins and their growth promoting transcripts in susceptible muscle populations in vivo. Through the identification of novel dysregulated transcripts in the somatic and axonal translatome and examining the localization of mRNA binding protein in SMA motor neurons in vivo, this work will contribute to our long-term goal of understanding the importance of mRNA processing and axonal localization as a regulator of disease pathophysiology in SMA.
Spinal muscular atrophy (SMA) is the leading genetic cause of infant mortality worldwide, and is the direct result of a deficiency in the survival of motor neuron protein (SMN). With no effective cure or treatment available, it is imperative to examine aspects of SMN that may play a pivotal role in modifying disease pathology in vivo. The results from this study will further define the non-canonical role of SMN in mRNA localization and processing, and reveal novel targets dysregulated in SMA that will continue to advance the development of therapeutic interventions to treat SMA.
