Identifying and characterizing new therapy targets in TDP-43 proteinopathies - Project Summary: Age-related neurodegenerative diseases are a rapidly growing cause of mortality and morbidity worldwide. The overwhelming majority of neurodegenerative disease, referred to as ‘sporadic,’ is caused by poorly understood interactions between genetic and environmental risk factors. Due in part to the complex etiology of neurodegenerative disease, broadly effective therapies are lacking. While the clinical symptoms of these disorders are heterogenous, reflecting selective neuronal death in distinct brain regions, there are pathophysiological features that link some neurodegenerative diseases. One such commonly observed pathological hallmark is the aberrant nuclear clearance and cytoplasmic aggregation of TAR DNA-binding protein 43 (TDP-43), which, in addition to being the defining pathology of limbic-predominant age-related TDP-43 encephalopathy (LATE), is observed in 97% of patients with amyotrophic lateral sclerosis (ALS), 50% of patients suffering from frontotemporal dementia (FTD), and up to 57% of Alzheimer’s disease (AD) patients. Together, the neurodegenerative disorders characterized by TDP-43 pathology can be referred to as ‘TDP-43 proteinopathies.’ Therapies that slow or reverse TDP-43 dysfunction thus have the potential to impact a broad group of neurogenerative disease patients; however, potent modifiers of TDP-43 pathology are lacking. Moreover, the mechanism by which TDP-43 mislocalization results in neuronal death is incompletely understood, and it is likely that therapy targets that mediate this disease process remain undiscovered. We recently found that genetic variants impacting alternative polyadenylation (APA) of ATXN3 are a novel genetic risk factor for ALS. Subsequent experiments revealed that modulation of ATXN3 substantially impacts TDP-43 pathology in multiple cell types, including in human iPSC-derived neurons and in ALS/FTD patient brain tissue. In Aim 1 of this proposal, I will determine the functional significance of ATXN3 alternative polyadenylation and identify cellular pathways to modulate ATXN3 expression in human neurons. TDP-43 is a prolific RNA-binding protein, directly impacting the metabolism of over 6,000 RNAs. Recent high-profile studies have identified RNAs that are differentially expressed (e.g., stmn2) or alternatively spliced (e.g., unc13a) upon nuclear depletion of TDP-43; however, transcripts alternatively polyadenylated upon TDP- 43 nuclear loss remain comparatively unexplored, despite the fact that regulation of APA is a key function of TDP-43. We have identified hundreds of previously unknown APA genes in ALS/FTD patient neurons exhibiting nuclear clearance of TDP-43. Notably, we found significant APA of MARK3, a tau kinase associated with early tau S262 phosphorylation in AD, reflecting a possible mechanistic link between TDP-43 and tau pathology. In Aim 2, I propose to identify and characterize new TDP-43 target genes in human neurons, first by studying the function of MARK3 APA, and second, by developing a new cellular tool to precisely define the transcriptome of human neurons undergoing TDP-43 nuclear clearance in a cell-type and temporally-controllable manner.
