Identifying microRNAs that bind to cytoplasmic TDP43

TDP43 is involved in microRNA biogenesis and found in cytoplasmic aggregates in amyotrophic lateral sclerosis (ALS), and microRNAs are important for regulation of gene expression and represent potential biomarkers and therapeutic targets. Therefore, we examined microRNAs that preferentially bind cytoplasmic TDP43 using cellular models expressing TDP43 variants and NanoString miRNA profiling analyses. We identified cytoplasmic TDP43-associated miRNAs and predicted genes and pathways to gain insights into potentially relevant disease pathways, biomarkers, and reversible therapeutic targets for ALS. Stable HeLa cell lines were established that expressed eGFP-His-tagged TDP43 variants (wild-type or ∆NLS) or an empty eGFP-His-tagged control construct. Cells were fractionated into nuclear and cytoplasmic fractions, cytoplasmic TDP43 was pulled down from the cytoplasmic fractions via its His tags, and microRNAs associated with cytoplasmic TDP43 were profiled by NanoString analysis.