Among known ALS genes, TDP-43 is of particular interest because its protein is hyper-phosphorylated and ubiquitinated in sporadic ALS. Compared with genetic mutation, post-translational modification is difficult to detect in patients and hard to reproduce in animal models. Therefore, a critical step toward understanding TDP-43 pathogenesis is revealing the pathways by which mutant TDP-43 causes neurodegeneration in ALS. TDP-43 binds to many RNAs and proteins, but how pathogenic mutation impacts TDP-43 function has not yet been determined, particularly in an appropriate animal model. Unexpectedly, overexpression of both WT and mutant TDP-43 in rodents causes indistinguishable phenotypes, indicating that excessive TDP-43 is neurotoxic. Because transgene expression is heavily influenced by position effect, the patterns and levels of transgene expression vary greatly from line to line. Whether mutant TDP-43 is more or less toxic than its WT form thus cannot be determined using existing transgenic models. The question regarding the impact of pathogenic mutation on TDP-43 function is further confounded by observations that TDP-43 depletion induces neurodegeneration in TDP-43 knock-out and knock-down mice. As TDP-43 binds to the 3'-untranslated region of its own mRNA and thereby regulates its own expression, the cDNA knock-in (KI) approach destroys TDP-43 self-regulatory machinery, rendering TDP-43 KI animals not very useful. Even 7 years after the discovery of TDP-43 mutation in ALS, whether mutant TDP-43 causes ALS through a gain or loss of function is still uncertain, and this unresolved issue is a roadblock to unraveling TDP-43 disease mechanisms. To resolve this critical issue, we created KI rats by introducing a single disease-linked point mutation into the rat genome. With the unprecedented rat models, we will determine how pathogenic mutation impacts TDP-43 function in a systematic manner.
Using novel knockin rats, this proposal will examine how pathogenic mutation of TDP-43 affects its function in a systematic manner, providing a critical insight into the mechanisms of amyotrophic lateral sclerosis caused by TDP-43 mutation.
