Defining and modulating the stress granule proteome as a therapeutic strategy in Amyotrophic Lateral Sclerosis

ended

How do you study a disease with no known cause? Amyotrophic lateral sclerosis (ALS) is a devastating neurodegenerative disease. Patients typically die 3-5 years after symptom onset. There is no cure. Finding a cure is hindered by the lack of obvious causes: although 10% of patients show familial inheritance (fALS), 90% of patients exhibit a sporadic form of ALS with no known cause (sporadic ALS, sALS). Almost all ALS patients demonstrate intracellular inclusions of the RNA binding protein TDP-43. However, it is not clear what process allows TDP-43 to aggregate, especially in sALS. This will be the focus of the proposed StressOME project. TDP-43, and other ALS-associated proteins are recruited into stress granules, transient structures that form in response to cellular stresses. Stress granules are thought to constitute a microenvironment with a high local concentration of TDP-43, sufficient to allow its aggregation; however this is prevented in healthy neurons. Therefore, the composition of stress granules may be crucial in the pathogenesis of ALS. To determine whether the dynamics of stress granule assembly and disassembly are different in patient cells, I will derive skin cells from fALS and sALS cases and compare them to age-matched controls. In parallel, I will use a new technique called ‘biotinylation by antibody recognition’ to define the stress granule proteome in sALS and fALS patient cells for the first time. This will allow me to identify candidate genes that modulate stress granule dynamics. I will generate stress granule reporter lines and misexpress candidate proteins, using live cell imaging to determine their effect on the dynamics of stress granules and the recruitment of TDP-43. Candidate genes will also be misexpressed in Drosophila models expressing TDP-43 in order to test their involvement in aggregation and toxicity. Through this approach I will identify novel targets that affect the aggregation of TDP-43 not only in fALS but also sALS.