Introduction: Spinal and bulbar muscular atrophy (SBMA) is a neurodegenerative disease caused by a polyglutamine (polyQ) expansion in the androgen receptor (AR). Instead of being exported out and degraded, mutant polyQ-AR remains in the nucleus, forming aggregates. We hypothesize that the cellular toxicity in SBMA is caused by aberrant intra-nuclear interactions (both gained and lost) of polyQ-AR aggregates.
Methods: We utilized SILAC proteomics on a cell model of SBMA to develop an AR interactome and investigate which intra-nuclear proteins and protein networks are affected through interaction with polyQ-AR (population C18) in comparison to wild type AR (C21). A candidate protein was validated using Western Blot and proximity ligation assay (PLA). 100 cells per condition were analyzed for mean gray value and integrated density across the cell.
Results: From the SILAC screen, we chose aromatic-L-amino-acid decarboxylase (DDC), which was elevated 45-fold (p<0.0001) in C21s, to further validate. A one-way ANOVA revealed similar DDC expression across both cell lines in the Western Blot. A Kolmogorov-Smirnov test revealed a significant increase in both mean gray value (p<0.0001) and integrated density (p<0.006) of the C18 cell population compared to the C21 cells.
Discussion: While the SILAC screen suggested that AR-DDC interaction was downregulated in SBMA, our validation studies implied the opposite. The next steps would be to repeat these studies in vitro and also in vivo mouse models. We hope to eventually develop new therapeutic targets for SBMA by further studying this interaction as well as its downstream effects on other protein networks.
Recommended CitationChatterjee, Debotri, "Using SILAC-based proteomics to investigate pathogenic mechanisms in spinal and bulbar muscular atrophy" (2021). Phase 1. Paper 77.