Regulation and impact of shortened TDP43 isoforms in FTD and ALS

University Of Michigan At Ann Arbor, Ann Arbor MI

Investigators

Abstract

ABSTRACT Mislocalization of the transactive response element DNA/RNA binding protein of 43 kDa (TDP43) is a signature event in an increasing number of degenerative diseases, including amyotrophic lateral sclerosis (ALS), frontotemporal lobar degeneration (FTLD), and inclusion body myositis (IBM). Despite this, however, the origins of TDP43 pathology remain obscure. Our previous experiments in neuronal models of hyperexcitability — a conserved and early feature of TDP43 proteinopathies — provided a new and unanticipated explanation for TDP43 pathology in these conditions. Specifically, we observed the accumulation of alternatively-spliced, shortened (s)TDP43 isoforms in hyperexcitable neurons. Unlike conventional, full-length TDP43, sTDP43 isoforms are cytoplasmic and highly prone to aggregation. sTDP43 deposition leads to flTDP43 sequestration within the cytoplasm and relative flTDP43 loss-of-function, recapitulating key pathological and molecular features of disease. Still, the factors responsible for sTDP43 deposition within hyperexcitable neurons, the connection between these pathways and disease pathogenesis, and whether sTDP43 exhibits a physiological or pathophysiological function, are all unknown. To address these gaps, here we will collaborate with an exceptional team of RNA and protein biologists to uncover the mechanisms underlying sTDP43 accumulation in disease, and the impact of sTDP43 on RNA homeostasis. During the previous cycle, we determined that sTDP43 is a byproduct of flTDP43 autoregulation, but its levels are severely limited by nonsense mediated RNA decay (NMD). In Aim 1, we test the hypothesis that sTDP43 accumulates in hyperexcitable neurons due to active TDP43 transcription and autoregulation, combined with NMD inhibition via the integrated stress response (ISR). We will also determine whether the ISR is necessary and/or sufficient for NMD inhibition and sTDP43 expression in human neurons. Working with Co-I Joshua Welch, in Aim 2 we utilize spatial transcriptomics to simultaneously examine neuronal hyperexcitability, NMD, and the ISR, in relation to sTDP43 and flTDP43 pathology in ALS/FTLD post-mortem material. We also take advantage of spatial proteomics to selectively investigate sTDP43 deposits and their content within ALS and control spinal neurons. These investigations, performed in collaboration with Co-I Wilfried Rossol, will be the first targeted study of sTDP43 interactions in human tissue. Lastly, in Aim 3 we employ genetic code expansion to non-invasively label sTDP43 and flTDP43 within human induced pluripotent stem cells. This platform provides a tractable means of visualizing, manipulating and purifying TDP43 variants in differentiated cells, enabling accurate studies of sTDP43/flTDP43 localization, trafficking and function in motor neurons and myocytes, vulnerable cell types affected in ALS and IBM. At the completion of these studies, we will have defined the mechanism as well as the functional impact of sTDP43 deposition in TDP43 proteinopathies, providing a glimpse into the origins of these conditions, and highlighted novel pathways that may serve as the basis for future therapies.