Doctor of Philosophy
School of Biological Sciences
Brown, Rebecca Anne, The effects of clusterin on the aggregation and pathogenicity of TDP-43, a protein implicated in amyotrophic lateral sclerosis, Doctor of Philosophy thesis, School of Biological Sciences, University of Wollongong, 2015. http://ro.uow.edu.au/theses/4664
This study aimed to examine whether TDP-43 was capable of cell-to-cell spread in cell culture and in vivo using a Drosophila model, and if so whether the extracellular chaperone clusterin could prevent this spread. Additionally it was determined whether the expression of TDP-43 in a subset of Drosophila glial cells was sufficient to induce motor neuron defects and mortality, and whether the co-expression of clusterin was able to reduce this. The ability of clusterin to inhibit the aggregation of TDP-43 in vitro was also tested. Finally, thoracic spinal cord sections from ALS patients were examined via immunohistochemistry to determine if clusterin was a constituent of TDP-43 inclusions, which would be consistent with the two proteins interacting during ALS progression.
Results from both cell-culture and in vivo experiments demonstrated that TDP-43 was able to spread from the glial cells where it was originally expressed, to surrounding cells which included motor neurons. Taken together these results suggest that like SOD1, TDP-43 may have prion-like modes of transmission. In Drosophila larvae the co-expression of clusterin was able to reduce this spread. Expression of TDP-43 in the glial cells of Drosophila caused a significant reduction of lifespan and locomotor defects. While the co-expression of clusterin was not able to extend lifespan, it did result in the preservation of locomotor ability. This effect may have resulted from clusterin inhibiting TDP-43 aggregation, as it was found that clusterin could potently inhibit the aggregation of TDP-43286-331, a synthetic peptide corresponding to residues 286-331 of TDP-43. Finally, clusterin was identified in the cytoplasm of human motor neurons where it was found to be co-localised with aggregates containing TDP-43 in spinal cord tissue from ALS but not control patients. Collectively, the results suggest that clusterin may be involved in preventing the aggregation of TDP-43 in ALS, possibly by interacting with TDP-43 in the extracellular space during prion-like transmission between cells, or in the intracellular space following retrotranslocation of clusterin to the cytosol.