Proteomic characterisation of TDP-43 inclusion pathology in amyotrophic lateral sclerosis and frontotemporal dementia
thesisposted on 28.03.2022, 23:20 by Thomas James Hedl
Amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD) are part of a spectrum of adult-onset fatal neurodegenerative diseases. ALS begins in either the upper motor neurons of the motor cortex or the lower motor neurons of the brain-stem and spinal cord, resulting in progressive paralysis. FTD however presents in the frontal and temporal lobes of the brain, causing debilitating changes to personality, language and behaviour. The pathological hallmark of both diseases is the presence of low-solubility protein aggregates (which form large inclusions in affected neurons), usually within the cytoplasm. These inclusions consist of both misfolded non-functional proteins as well as functioning proteins that are sequestered due to abnormal interactions or endogenous aggregate-clearance mechanisms. The major inclusion-forming protein in the vast majority of ALS and approximately half of all FTD cases is the DNA/RNA-binding protein TAR DNA-binding protein of 43 kDa (TDP-43), however the other components of TDP-43 pathology in disease remain largely unknown. The aim of this thesis is to characterise the populations of proteins that are present within ALS/FTD-linked TDP-43 inclusions. Various forms of human TDP-43 (wildtype, cytoplasmic-targeted and post-translational mimetics) were expressed in neuronal NSC-34 and SH-SY5Y cells, and brain tissue from a TDP-43 transgenic mouse model was also used. By developing a biochemical low-solubility protein fractionation method, the types of proteins present with the TDP-43 pathology in these cells and brain samples were isolated for shotgun mass spectrometry proteomics. This approach identified many potential disease-associated as well as previously unidentified proteins that may play a role in ALS/FTD pathology. By identifying the types of proteins that are affected in these terrible diseases, it will be possible to understand the pathways and mechanisms through which they act to ultimately target therapeutics and identify biomarkers for people living with ALS and FTD.