FUS/TLS GAIN AND LOSS OF FUNCTION IN ALS: ANIMAL AND CELLULAR MODELS OF DISEASE

DESCRIPTION (provided by applicant): Amyotrophic Lateral Sclerosis (ALS) is a progressive neurodegenerative disorder in which preferential loss of motor neurons (MNs) results in paralysis and death. Although ALS is largely a sporadic disease, research has focused on heritable forms of the disorder because clinical and pathological evidence suggests common pathogenic mechanisms. Mutations in the gene FUS (or TLS) were recently reported in rare ALS families, and FUS pathology has since been found in sporadic ALS, suggesting that FUS may provide a mechanistic link between familial and sporadic disease. Structural and functional similarities between FUS and TDP-43 - another RNA/DNA-binding protein involved in the pathogenesis of sporadic and familial ALS - have also led to speculation that the molecular pathways regulated by both of these factors are vital to our understanding of common disease mechanisms. We know very little about how mutations in FUS cause motor neuron degeneration. Dominant inheritance of FUS mutations suggests a novel gain of function that is selectively toxic to motor neurons. Alternatively, mutant FUS may act as a dominant negative, inhibiting the normal activity of wild type protein, perhaps by sequestering it in abnormal FUS-positive, cytoplasmic aggregates that are a hallmark of sporadic and familial ALS. If ALS results as a consequence of FUS deficiency, then it is critical to understand more about the normal functions of FUS in the central nervous system, and specifically in the motor circuits affected in the disease. In this project, loss and gain of function strategies are used to explore the role of FUS in normal motor neuron development in animal and cellular models, and to relate that function to mutant FUS-mediated ALS. In vitro studies will take advantage of our ability to generate FUS mutant embryonic stem cell-derived motor neurons in large numbers. In Aim 1, FUS knockout mice will be used to support the hypothesis that motor neuron degeneration in ALS is a consequence of FUS deficiency. We will test the effect of FUS loss on motor neuron differentiation and survival and on the functional development of spinal motor circuits required for normal motor activity. By high-throughput RNA sequencing (RNA Seq), we will explore the normal role of FUS in the regulation of gene expression in the nervous system. In Aim 2, we will use overexpression studies of mutant FUS to characterize the effect on motor neuron survival and function in vivo, and to determine how mutations alter the functional properties of FUS in ALS. RNA Seq analysis will be used to identify molecular pathways involved in the pathogenesis of disease. In Aim 3, we will use motor neurons derived from mouse embryonic stem cells to study cellular and molecular mechanism of FUS-mediated motor neuron degeneration. This project will address fundamental questions about the role of FUS in ALS, and generate novel models of FUS-mediated disease in mice and cultured motor neurons that will be critical tools for future studies of disease mechanism and drug discovery in the ALS research community.