RNA-binding tunes the conformational plasticity and intradomain stability of TDP-43 tandem RNA recognition motifs

Abstract

TAR DNA binding protein 43 (TDP-43) is a nuclear RNA/DNA-binding protein with pivotal roles in RNA-related processes such as splicing, transcription, transport, and stability. The high binding affinity and specificity of TDP-43 towards its cognate RNA sequences (GU-rich) is mediated by highly conserved residues in its tandem RNA recognition motif (RRM) domains (aa:104-263). Importantly, the loss of RNA-binding to the tandem RRMs caused by physiological stressors and chemical modifications promotes cytoplasmic mislocalization and pathological aggregation of TDP-43. Despite the substantial implications of RNA in TDP-43 function and pathology, a comprehensive characterization of the effect of RNA-binding on conformational dynamics, interdomain interactions and intradomain stability of the tandem RRMs has not yet been conducted. Here, we employed all-atom molecular dynamics (MD) simulations to assess the effect of RNA-binding on the conformational landscape and intradomain stability of TDP-43 tandem RRMs. Our simulations reveal a high intrinsic conformational plasticity of the tandem RRMs in the absence of RNA which surprisingly, is accompanied by a tendency of RRM1 to adopt partially-unfolded conformations. While binding to RNA limits the overall conformational space of the tandem RRMs and promotes intradomain stability, several RRM-RNA contacts mediated by highly conserved residues are observed to be far more dynamic than previously inferred from NMR structural ensemble. Overall, our simulations reveal how RNA dynamically tunes the structural and conformational landscape of TDP-43 tandem RRMs, contributing to physiological function and mitigating pathological aggregation. SIGNIFICANCEThe cytoplasmic mislocalization and aggregation of TDP-43 due to loss of its RNA-binding capability is associated with the onset and progression of neurodegenerative diseases such as Amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). Due to the flexible nature of RNA and the presence of a disordered linker between RRM domains, characterizing the dynamic interactions between RRMs-RNA and/or RRM1-RRM2 by experiments alone has remained challenging. In this study, we performed all-atom simulations initiated from the NMR conformers of RNA-bound tandem RRMs of TDP-43 to investigate their underlying structural and conformational dynamics. Our findings indicate that RNA binding effectively reduces conformational heterogeneity in the tandem RRMs and acts as a protective factor for the unfolding and aggregation of RRM1. These effects are achieved through a combination of stable and dynamic protein-RNA interactions which involve highly conserved amino acids.