Both familial ALS (FALS) and sporadic ALS (SALS) exhibit dysregulation of RNA metabolism, indicating that homeostasis of the transcriptome is crucial for motor neuron survival. Indeed, almost all ALS converges on TDP-43 pathology and thus, widespread transcriptome alterations. Given that changes in RNA metabolism occur in all forms of ALS, identifying key transcripts altered in the course of disease pathogenesis – particularly those caused by TDP-43 loss of function – could yield novel, broadly applicable therapeutic targets.
In collaboration with Dr. Wei Li (UC Irvine), we found that alternative polyadenylation (APA), an aspect of RNA metabolism understudied in ALS, occurs broadly upon TDP-43 mutation or depletion. APA affects RNA metabolism by producing distinct transcripts with shortened or lengthened 3′UTRs containing different cis-regulatory elements, such as binding sites for microRNAs (miRNAs) and RNA binding proteins (RBPs), leading to altered RNA stability, protein translation, or subcellular localization of a given transcript. We applied the dynamic analysis of polyadenylation from RNA-seq (DaPars) tool to ALS RNA-sequencing datasets, finding hundreds of previously unknown APA events in genes that function in pathways implicated in ALS pathogenesis, such as nucleocytoplasmic transport and the oxidative stress response. In addition, we found that APA of MARK3, a tau kinase implicated in Alzheimer’s disease, leads to increased MARK3 protein levels in neuronal cells depleted of TDP-43, reflecting a novel mechanistic link between TDP-43 and tau pathology. In a parallel study, we further implicated APA in the etiology of ALS, finding that genetic variants affecting APA of Ataxin-3 (ATXN3) are strongly associated with ALS. Knockdown of ATXN3 dramatically increased the accumulation of TDP-43 C-terminal fragments in cell models of ALS, including in iPSC-derived motor neurons. Moreover, we found that ATXN3 protein levels are reduced in postmortem frontal cortex of ALS/FTD patients and that ATXN3 levels are inversely correlated with the levels of phosphorylated TDP-43 in the same samples. Importantly, APA can be directly modulated by antisense oligonucleotides (ASOs); thus, newly identified APA genes as well as ATXN3 may be candidates for rapid therapy development in ALS.