Multivalent GU-rich oligonucleotides sequester TDP-43 in the nucleus by inducing high molecular weight RNP complexes

Abstract

ABSTRACTThe loss of nuclear TDP-43 localization and its accumulation in cytoplasmic aggregates are hallmarks of neurodegeneration and major therapeutic targets. We recently demonstrated that TDP-43 binding to endogenous nuclear GU-rich RNAs sequesters TDP-43 in the nucleus and restricts its passive nuclear export. Here, we tested the feasibility of synthetic RNA oligonucleotide-mediated augmentation of TDP-43 nuclear localization. Using biochemical assays, we compared the ability of GU-rich oligonucleotides to engage in multivalent, RRM-dependent binding with TDP-43 and identified (GU)16 as a strong multivalent binder. When transfected into cells, unlike monovalent oligonucleotides that displaced TDP-43 from the nucleus, (GU)16 preserved steady-state TDP-43 nuclear localization and prevented transcriptional blockade-induced TDP-43 mislocalization. RNA pulldowns from (GU)16-transfected cells confirmed that (GU)16 induced high molecular weight RNP complexes, incorporating TDP-43 and possibly other GU-binding proteins. Transfected (GU)16 caused partial failure of TDP-43 cryptic exon repression, likely because the high-affinity oligonucleotides diverted TDP-43 from endogenous RNAs. Thus, while GU-rich oligonucleotides can attenuate TDP-43 mislocalization, optimization is needed to avoid TDP-43 loss of function.