Molecular Dynamics simulation of TDP-43 RRM in the presence and absence of RNA

Scott, David D.; Mowrey, David; Nagarajan, Karthi; François‐Moutal, Liberty; Nair, Anil; Khanna, May

doi:10.1101/2022.03.15.484514

Cited by 5 publications

(3 citation statements)

References 22 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Our simulation results of the apo-structure of the RRM1-2 domains with EGCG showed different binding regions during the 800 ns simulation run which might be the result of the dynamic conformation changes and exposure of certain buried residues during the simulation. Our results are consistent with the previous reports showing the dynamic conformation changes during the MD simulation run for the apo-structure of tandem RRM1-2 domains [58]. However, EGCG remained stably bound to the hydrophobic, non-planar, amino terminal region of the ensemble structure of the low complexity C-terminal domain of TDP-43 (PDB ID: 7KWZ, aa: 276-414) for the 300 ns simulation performed, unlike during the simulation performed with both the N-terminal (PDB ID: 5MDI, aa: 1-80) and the RRM1-2 domain (PDB ID: 4BS2, aa: 96-269).…”

Section: Discussionsupporting

confidence: 94%

In silicoandin vitrostudies suggest epigallocatechin gallate (EGCG), a polyphenol in green tea, can bind and modulate the aggregation and cytotoxicity of the full-length TDP-43 protein implicated in TDP-43 proteinopathies

Meshram,

Balaji,

Saravanan

et al. 2023

Preprint

View full text Add to dashboard Cite

Misfolding and aggregation of TDP-43 protein are implicated in several proteinopathies like ALS and FTLD. Extracellular TDP-43 is also proposed to propagate in a prion-like pathogenic manner to the neighbouring cells. Here, using turbidity and sedimentation assay, we show that a polyphenol in green tea, epigallocatechin gallate (EGCG), can inhibit thein vitroaggregation of the full-length TDP-43 protein. Furthermore, Alexa-Fluor-labelled TDP-43 protein failed to show aggregates in the presence of EGCG in fluorescence microscopy. Also, AFM imaging revealed that EGCG co-incubation with TDP-43 allows formation of only small oligomers in contrast to the larger TDP-43 aggregates formed otherwise. A physical binding of EGCG with TDP-43 was observed using triphenyl tetrazolium chloride (TTC) staining and isothermal titration calorimetry (ITC). ITC also revealed a high-affinity binding site for EGCG on TDP-43 with a Kdvalue of 7.8 µM and a binding free energy of -6.9 kcal/mol. Furthermore,in silicomolecular docking and molecular dynamic simulation (MDS) studies using different available structures of the N-terminal, RRM1-2 and C-terminal domains of TDP-43, predicted a preferable and stable binding of EGCG to the structure of the aggregation prone C-terminal domain (CTD) (PDB ID:7KWZ). Also, EGCG complexed with CTD of TDP-43 yielded a negative ΔG value of -20.29 kcal/mol using MM-PBSA analysis of the MDS data thereby further suggesting a stable complex formation. Also, in MDS, EGCG interacted with the amino acids Phe-313 and Ala-341 of TDP-43, which were previously projected to be important for the recruitment of monomers for the amyloid formation by CTD, thereby suggesting a possible mechanism of EGCG’s inhibition of the TDP-43 aggregation. Notably, while thein vitro-made aggregates of full-length TDP-43 caused mild cytotoxicity to the HEK293 cells, the small oligomers of TDP-43 formed in presence of EGCG did not. In totality, EGCG canin vitrointeract with TDP-43 and inhibit its aggregation, possiblyviainteraction with the amyloidogenic domain, thereby preventing it from assuming cytotoxic conformations. As EGCG is a natural molecule, it could be relevant to the therapeutic quest against the TDP-43 proteinopathies.

show abstract

Section: Discussionsupporting

confidence: 94%

In silicoandin vitrostudies suggest epigallocatechin gallate (EGCG), a polyphenol in green tea, can bind and modulate the aggregation and cytotoxicity of the full-length TDP-43 protein implicated in TDP-43 proteinopathies

Meshram,

Balaji,

Saravanan

et al. 2023

Preprint

View full text Add to dashboard Cite

show abstract

“…Given, as mentioned above, the ability of a 20-mer PS-ASO to nucleate the assembly of PSPs into nuclear condensates lacking NEAT1 ( Shen et al, 2014 ), can strategies be designed (for instance via machine learning) to uncover similar PS-ASOs that condense NEAT1 and TDP-43? In this regard, it is noteworthy that an attempt to deploy AlphaFold and other computational approaches in expanding the targetable chemical space of TDP-43 has already been reported ( Scott et al, 2022 ). However, the recent early termination of promising ASO clinical trials for HD, due to lack of efficacy ( Kwon, 2021 ), is a reminder of how challenging the effective deployment of ASO in any neurodegenerative disorders is likely to remain for some time.…”

Section: Therapeutic Horizons: Oligonucleotide Approachesmentioning

confidence: 99%

TDP-43 and NEAT long non-coding RNA: Roles in neurodegenerative disease

Sekar

Tusubira²,

Ross³

2022

Front. Cell. Neurosci.

View full text Add to dashboard Cite

Understanding and ameliorating neurodegenerative diseases represents a key challenge for supporting the health span of the aging population. Diverse protein aggregates have been implicated in such neurodegenerative disorders, including amyloid-β, α-synuclein, tau, fused in sarcoma (FUS), and transactivation response element (TAR) DNA-binding protein 43 (TDP-43). Recent years have seen significant growth in our mechanistic knowledge of relationships between these proteins and some of the membrane-less nuclear structures that fulfill key roles in the cell function. These include the nucleolus, nuclear speckles, and paraspeckles. The ability of macromolecular protein:RNA complexes to partition these nuclear condensates through biophysical processes that involve liquid–liquid phase separation (LLPS) has also gained attention recently. The paraspeckle, which is scaffolded by the architectural long-non-coding RNA nuclear enriched abundant transcript 1 (NEAT1) plays central roles in RNA processing and metabolism and has been linked dynamically to TDP-43. In this mini-review, we outline essential early and recent insights in relation to TDP-43 proteinopathies. We then appraise the relationships between TDP-43 and NEAT1 in the context of neuronal paraspeckles and neuronal stress. We highlight key areas for investigation based on recent advances in our understanding of how TDP-43 affects neuronal function, especially in relation to messenger ribosomal nucleic acid (mRNA) splicing. Finally, we offer perspectives that should be considered for translational pipelines in order to improve health outcomes for the management of neurodegenerative diseases.

show abstract

“…Alternatively, the integration of knowledge obtained by the nuclear magnetic resonance (NMR) experiments with molecular dynamics (MD) simulations shows promising results in providing a more refined representation of the protein-RNA complexes [37,39]. While previous studies have utilized MD simulations to investigate the intradomain stability of TDP-43 RRMs and their interactions with RNA [40][41][42][43][44], a comprehensive understanding of the tandem RRM conformational landscape and how RNA modifies it to promote TDP-43 function and inhibit pathological aggregation, is still lacking. To this end, we conducted multi-microsecond long MD simulations of the NMR-derived conformer ensemble of TDP-43 tandem RRMs which was previously determined in the presence of a bound GU-rich RNA fragment [2] to investigate and elaborate on the changes in the interdomain flexibility and intradomain stability which occur upon RNA-binding.…”

Section: Introductionmentioning

confidence: 99%

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

Ozguney,

Mohanty,

Mittal

2024

Preprint

View full text Add to dashboard Cite

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.

show abstract

Molecular Dynamics simulation of TDP-43 RRM in the presence and absence of RNA

Cited by 5 publications

References 22 publications

In silicoandin vitrostudies suggest epigallocatechin gallate (EGCG), a polyphenol in green tea, can bind and modulate the aggregation and cytotoxicity of the full-length TDP-43 protein implicated in TDP-43 proteinopathies

In silicoandin vitrostudies suggest epigallocatechin gallate (EGCG), a polyphenol in green tea, can bind and modulate the aggregation and cytotoxicity of the full-length TDP-43 protein implicated in TDP-43 proteinopathies

TDP-43 and NEAT long non-coding RNA: Roles in neurodegenerative disease

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

Contact Info

Product

Resources

About