Structure/function analysis of TDP-43 neurotoxicity in C. elegans

Abstract

TDP-43 is a conserved RNA binding protein with known roles in mRNA splicing and stability. Cytoplasmic deposition of TDP-43 has been linked to multiple neurodegenerative diseases, including ALS and frontotemporal lobar dementia (FTLD). We have engineered pan-neuronal expression of human TDP-43 protein in C. elegans, with the goal of generating a convenient in vivo model of TDP-43 neurotoxicity. Full-length (wild type) human TDP-43 expressed in C. elegans is nuclear as is observed in human cells. Transgenic worms with neuronal human TDP-43 expression exhibit an uncoordinated phenotype and have abnormal motorneuron synapses. By using this uncoordinated phenotype as a read-out of TDP-43 neurotoxicity, we have investigated the contribution of specific TDP-43 domains as well as TDP-43 sub-cellular localization to toxicity. Deletion of either RNA recognition domain (RRM1 or RRM2) completely blocks neurotoxicity, as does deletion of the C-terminal region. These deleted TDP-43 variants still accumulate in the nucleus, although their subnuclear distribution is altered. In contrast, N-terminal deletions result in the formation of toxic cytoplasmic aggregates. Mutation of the TDP-43 nuclear localization signal (NLS) results in cytoplasmic deposition of fulllength TDP-43, which is not toxic. Mutations that alter two TDP-43 caspase cleavage sites (D89/219E), however, do not reverse TDP43 toxicity. Our results demonstrate that TDP-43 neurotoxicity can result from either nuclear activity of the full-length protein or accumulation of cytoplasmic aggregates composed of C-terminal fragments. These results suggest that there may be (at least) two different mechanisms of TDP-43 neurotoxicity.