Tau variants in FUS-mediated ALS

About This Grant

Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disease with no effective treatment. Among the identified ALS-causing genes, several encode RNA binding proteins including FUsed in Sarcoma (FUS). Mutant FUS protein is mis-localized to the cytoplasm where it forms inclusions, a pathological hallmark of ALS. We and others have studied the FUS protein under physiological and pathological conditions in various models. Notably, we recently identified six individuals in an extended ALS kindred who carry the ALS-linked FUS R521G mutation but remain free of ALS well into their 60s and beyond (Unaffected Mutation Carriers, UMCs). We generated induced pluripotent stem cell (iPSC) lines from four UMCs, six FUS ALS patients, and four healthy controls, differentiated them to motor neurons (iMNs), and measured their electrophysiological properties. FUS ALS iMNs demonstrated disease-related hyperexcitability, whereas UMC and control iMNs displayed normal electrophysiological properties. Using an integrated analysis of whole genome sequencing and RNA-Seq data, we identified a cluster of variants in the 3’ untranslated region (3’-UTR) of the microtubule- associated protein tau (MAPT) gene. This set of linked variants was highly expressed in all UMCs but was absent in all ALS patients. More excitingly, an isogenic line with these variants incorporated into MAPT 3’-UTR of an ALS iPSC showed normal electrophysiological properties, supporting that these variants are critical to mitigating the hyperexcitability phenotype. iMNs from FUS ALS patients had increased Tau protein levels as compared to those from UMCs with the MAPT variants. In addition, overexpression of Tau induced neuronal hyperexcitability whereas Tau knockdown restored neuronal excitability to normal. We thus hypothesize that the MAPT variants reduce Tau protein expression, mitigate iMN hyperexcitability, and ultimately protect UMCs from developing ALS. Three specific aims are designed to test the hypothesis. Aim 1 is to establish the causative role of the MAPT 3’-UTR variants in reducing the hyperexcitability phenotype. We will generate additional isogenic lines by introducing the wild-type MAPT allele to UMC iPSCs to solidify the causative role of the MAPT variants in determining ALS or UMC phenotypes. We will generate and characterize two isogenic lines carrying 4 rare variants or 10 common variants to delineate their contributions. Aim 2 is to determine how the FUS mutation increases Tau expression and alters other properties, and how the MAPT variants prevent these abnormalities. Aim 3 is to determine which ion channel or receptor is responsible for the ALS-related hyperexcitability phenotype, and to examine whether up- and down-regulation of Tau expression changes the specific ion channel or receptor activity. The proposed studies are highly innovative conceptually and technically. The results will provide insights into the mechanisms underlying the protective effects of the MAPT variants and identify potential novel therapeutic targets. This project has a broader impact beyond ALS since Tau plays a critical role in other neurodegenerative diseases.