Research Projects

TDP-43 is implicated in the fatal neurodegenerative diseases ALS and FTLD [1]. Depletion of TDP-43 from the nucleus and mislocalization to the cytoplasm, where it accumulates into phosphorylated and ubiquitinated inclusions are disease hallmarks in affected neurons [2]. These inclusions trap other key metabolism proteins and disturb the cellular ubiquitin pool, further aggravating the disease conditions. The frequency of ALS cases in India is 5 in 100,000, a figure established by the Foundation for Research on Rare Diseases and Disorders. Furthermore, the age of ALS onset in India is estimated to be up to a decade earlier than the worldwide average, at about 40 years. It is estimated that nearly 3 lakh people die of ALS every year in India [3,4]. The interplay of inadequate UPS response and TDP-43 deposition is being increasingly recognized [5,6]. TDP-43 has been identified as substrate for ubiquitin ligases such as Parkin, UBE2E, and RNF4. There are about 100 deubiquitinases (DUB) in the human proteome [7]. The DUBs USP13 and USP10 have been identified to affect TDP-43 aggregation, although independent of ubiquitin peptidase activity [8,9]. Recently, the ER membrane embedded USP19 was shown to assist the secretion of misfolded mutant TDP-43 via its DUB activity [10]. Mutation in the DUB CLYD has been found to promote cytoplasmic mislocalization of TDP-43 [11]. Another DUB, Ataxin-3 was identified as a significant gene in a transcriptome-wide association study for ALS. Ataxin-3 knockdown increased the accumulation of ubiquitinated TDP-43 C-term fragments [12]. These findings suggest a role for ubiquitin-mediated response in TDP-43 pathology. Soluble cytoplasmic TDP-43 microaggregates, which are small in size and highly mobile, leads to degradation by UPS and autophagy-lysosomal pathways [13]. In disease conditions, larger aggregates found in inclusions, bearing complex ubiquitination patterns, overburden these pathways. In this research project we seek to understand the clearance mechanism of TDP-43 aggregates through UPS. Specifically, we seek to investigate whether DUBs could be tasked with maintaining TDP-43 levels. We hypothesize that priming TDP-43 for UPS-mediated degradation with molecular glues could improve neuronal health. Conversely, improving TDP-43 stability by cleaving ubiquitin chains off TDP-43 aggregates, could restore TDP-43 functions and restore free ubiquitin in the cytoplasm. Using the novel chemo-proteomics approach of targeted protein degradation (TPD), TDP-43 protein turnover could be improved (Table-1). Using Ataxin-3 as a model DUB, we could investigate its interaction and deubiquitination efficiency with TDP-43 aggregates. We have generated stable cell Ataxin-3 model in SHSY5Y neuroblastoma cells and previously used it to study Ataxin-3 involvement in stress granule formation under arsenite and hyperosmotic stress. Our work could be expanded to other DUBs associated with proteostasis (Table-2).