Amyotrophic lateral sclerosis (ALS) is an adult-onset neurodegenerative disorder in which premature loss of upper and lower motor neurons leads to fatal paralysis with a typical disease course of one to five years. Mutations in two DNA/RNA binding proteins, TAR DNA-binding protein (TDP-43) and Fused in Sarcoma (FUS) have recently been identified as primary causes of inherited ALS and have led to what is likely to be a paradigm shift in efforts to understand the pathogenesis of ALS. Yet there remains a need to examine the functions of these proteins in cell types relevant for ALS progression, both motor neurons and glial cells. While a contribution from surrounding cells is well established in inherited ALS caused by SOD1 mutations, the impact of non-cell autonomous mechanisms in disease progression following mutation in TDP-43 or FUS is completely unknown. This proposal seeks to use embryonic stem cells derived from existing transgenic mice to examine the consequences of either reduction or mutation of TDP-43 or FUS in purified motor neurons. Cell intrinsic consequences of alterations in these genes will be determined by inducing differentiation of embryonic stem cells into motor neurons which either lack or express mutant TDP-43 or FUS. Rather than take a candidate approach to pursue the few genes that have already been identified as targets of TDP-43 or FUS in proliferating cell lines, isolated motor neuron cultures will be used to identify the complete set of RNAs that are altered by loss of TDP-43 or FUS in Aim 1, and then in Aim 2 to ask whether any RNAs are altered upon mutation of TDP-43 or FUS and if so, if they are the same as those observed upon loss of function. By taking such a comprehensive and systematic approach from purified starting material it is likely that the specific targets and signaling pathways affected can be uncovered - discoveries that might elucidate underlying mechanisms for disease and provide a basis for future therapeutic developments. Success in these goals will lead to greater understanding of ALS disease mechanism and will provide rationale for future studies in glial cell types to determine whether manipulation of supporting non-neuronal cells would provide therapeutic benefit in ALS patients with mutations in TDP-43 or FUS.

Public Health Relevance

Amyotrophic lateral sclerosis (ALS) is an adult-onset neurodegenerative disorder that affects neurons of the brain and spinal cord controlling muscle movement and leads to fatal paralysis within the course of one to five years. Mutations in two proteins have recently been identified as a primary cause of ALS, yet the mechanism by which they contribute to disease is unknown. This project seeks to understand the functions of normal or mutant forms of these proteins in purified motor neurons to uncover the specific pathways affected, and may have wider implications for other nervous system diseases with similar pathology.

National Institute of Health (NIH)
National Institute of Neurological Disorders and Stroke (NINDS)
Postdoctoral Individual National Research Service Award (F32)
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Special Emphasis Panel (ZRG1-F03A-F (20))
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Gubitz, Amelie
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Ludwig Institute for Cancer Research Ltd
La Jolla
United States
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Ditsworth, Dara; Maldonado, Marcus; McAlonis-Downes, Melissa et al. (2017) Mutant TDP-43 within motor neurons drives disease onset but not progression in amyotrophic lateral sclerosis. Acta Neuropathol 133:907-922
Foust, Kevin D; Salazar, Desirée L; Likhite, Shibi et al. (2013) Therapeutic AAV9-mediated suppression of mutant SOD1 slows disease progression and extends survival in models of inherited ALS. Mol Ther 21:2148-59