Inter-domain interactions of TDP-43 as decoded by NMR

Wei, Yuanyuan; Lim, Liangzhong; Wang, Lu; Song, Jianxing

doi:10.1016/j.bbrc.2016.03.158

Cited by 30 publications

(44 citation statements)

References 22 publications

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“…However, likely due to the interactions of the RRM domains with both N- and C-terminal LC domains, in the full-length FUS the well-dispersed HSQC peaks of the folded RRM domain become undetectable due to significantly provoked µs-ms dynamics, or/and increased rotational tumbling time. This is reminiscent of our previous observation on another ALS-/FTD-causing protein TDP-43 29, 52 . The well-dispersed HSQC peaks of two folded RRM domains also disappeared in the full-length TDP-43 and even its truncated fragment with the N-domain (1–101) deleted, which was characterized to result from µs-ms exchanges between the closed and open conformations mediated by dynamic inter-domain interactions coordinated by intrinsically-disordered prion-like domain 29 .…”

Section: Discussionsupporting

confidence: 82%

“…This is reminiscent of our previous observation on another ALS-/FTD-causing protein TDP-43 29, 52 . The well-dispersed HSQC peaks of two folded RRM domains also disappeared in the full-length TDP-43 and even its truncated fragment with the N-domain (1–101) deleted, which was characterized to result from µs-ms exchanges between the closed and open conformations mediated by dynamic inter-domain interactions coordinated by intrinsically-disordered prion-like domain 29 . Despite being very challenging, in the future it would be important to identify the exact regions involved in inter-domain interactions of FUS.…”

Section: Discussionsupporting

confidence: 82%

“…Our current study reveals that like TDP-43 23, 24, 29, 52–55 , the dynamics and self-assembly of FUS are also modulated by more than one domain/region, as well as by their interactions. As a consequence, various perturbations including genetic, pathological and environmental factors to one domain may be relayed to other domains to solidifying the reversible and physiological FUS assembly, thus leading to ALS/FTD by “loss-of-function” or/and “gain-of-function” mechanisms.…”

Section: Discussionmentioning

confidence: 65%

“…Very amazingly, we have previously observed the same phenomenon on another ALS/FTD-causing protein TDP-43. The well-dispersed HSQC peaks of two well-folded RRM domains also became disappeared in the contexts of the full-length TDP-43 as well as its fragment with the N-terminal domain deleted, which has been characterized to mainly result from µs-ms exchanges between the open and closed conformations mediated by dynamic inter-domain interactions 29 .…”

Section: Resultsmentioning

confidence: 99%

“…The generation of the isotope-labelled proteins for NMR studies followed a similar procedure except that the bacteria were grown in M9 medium with the addition of ( 15 NH 4 ) 2 SO 4 for 15 N labeling and ( 15 NH 4 ) 2 SO 4 /[ 13 C]-glucose for double labelling 23, 24, 29, 31 . The purity of the recombinant proteins was checked by SDS–PAGE gels and their molecular weights were verified by a Voyager STR matrix-assisted laser desorption ionization time-of-flight-mass spectrometer (Applied Biosystems).…”

Section: Methodsmentioning

confidence: 99%

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RRM domain of ALS/FTD-causing FUS characteristic of irreversible unfolding spontaneously self-assembles into amyloid fibrils

Lim

Song

2017

Sci Rep

Self Cite

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526-residue FUS functions to self-assemble into reversible droplets/hydrogels, which could be further solidified into pathological fibrils. FUS is intrinsically prone to aggregation, composed of N-terminal low-sequence complexity (LC); RNA-recognition motif (RRM) and C-terminal LC domains. Intriguingly, previous in vivo studies revealed that its RRM is required for manifesting FUS cytotoxicity but the underlying mechanism remains unknown. Here, we characterized solution conformations of FUS and its five differentially dissected fragments, followed by detailed investigations on thermal unfolding, NMR dynamics and self-assembly of RRM. The results decipher: (1) the N- and C-terminal LC domains are intrinsically disordered, while RRM is folded. Intriguingly, well-dispersed HSQC peaks of RRM disappear in the full-length FUS, reminiscent of the previous observation on TDP-43. (2) FUS RRM is characteristic of irreversible unfolding. “Model-free” analysis of NMR relaxation data decodes that RRM has high ps-ns conformational dynamics even over some residues within secondary structure regions. (3) RRM spontaneously self-assembles into amyloid fibrils. Therefore, in addition to the well-established prion-like region, FUS RRM is also prone to self-assembly to form amyloid fibrils. Taken together, FUS RRM appears to play a crucial role in exaggerating the physiological/reversible self-assembly into pathological/irreversible fibrillization, thus contributing to manifestation of FUS cytotoxicity.

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Section: Discussionsupporting

confidence: 82%

Section: Discussionsupporting

confidence: 82%

Section: Discussionmentioning

confidence: 65%

Section: Resultsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

See 3 more Smart Citations

RRM domain of ALS/FTD-causing FUS characteristic of irreversible unfolding spontaneously self-assembles into amyloid fibrils

Lim

Song

2017

Sci Rep

Self Cite

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Immunolocalization of TAR DNA‐binding protein of 43 kDa (TDP‐43) in mouse seminiferous epithelium

Osuru

Pramoonjago

Abhyankar

et al. 2017

Molecular Reproduction Devel

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TDP-43 (TAR DNA-binding protein of 43 kDa) is an evolutionarily conserved, ubiquitously expressed, multi-functional DNA/RNA-binding protein with roles in gene transcription, mRNA splicing, stability, transport, micro RNA biogenesis, and suppression of transposons. Aberrant expression of TDP-43 in testis and sperm was recently shown to be associated with male infertility, which highlights the need to understand better the expression of TDP-43 in the testis. We previously cloned TDP-43 from a mouse testis cDNA library, and showed that it functions as a transcriptional repressor, and regulates the precise spatiotemporal expression of the Acrv1 gene, which encodes the acrosomal protein SP-10, during spermatogenesis. Here, we performed immunoblotting and immunohistochemistry of the mouse testis using four separate antibodies recognizing the amino and carboxyl termini of TDP-43. TDP-43 is present in the nuclei of germ cells as well as Sertoli cells. TDP-43 expression begins in type B / intermediate spermatogonia, peaks in preleptotene spermatocytes, and becomes undetectable in leptotene and zygotene spermatocytes. Pachytene spermatocytes and early round spermatids again express TDP-43, but its abundance diminishes later in spermatids (at steps 5 to 8). Interestingly, two of the four antibodies showed TDP-43 expression in spermatids at steps 9–10, which coincides with the initial phase of the histone-to-protamine transition. Immunoreactivity patterns observed in the study suggest that TDP-43 assumes different conformational states at different stages of spermatogenesis. TDP-43 pathology has been extensively studied in the context of neurodegenerative diseases; its role in spermatogenesis warrants further detailed investigation of the involvement of TDP-43 in male infertility.

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A complex network of interdomain interactions underlies the conformational ensemble of monomeric TDP‐43 and modulates its phase behavior

Mohanty,

Rizuan,

Kim

et al. 2024

Protein Science

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TAR DNA‐binding protein 43 (TDP‐43) is a multidomain protein involved in the regulation of RNA processing, and its aggregates have been observed in neurodegenerative diseases, including amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). Numerous studies indicate TDP‐43 can undergo liquid‐liquid phase separation (LLPS) in vitro and is a component of biological condensates. Homo‐oligomerization via the folded N‐terminal domain (aa:1‐77) and the conserved helical region (aa:319‐341) of the disordered, C‐terminal domain is found to be an important driver of TDP‐43 phase separation. However, a comprehensive molecular view of TDP‐43 phase separation, particularly regarding the nature of heterodomain interactions, is lacking due to the challenges associated with its stability and purification. Here, we utilize all‐atom and coarse‐grained (CG) molecular dynamics (MD) simulations to uncover the network of interdomain interactions implicated in TDP‐43 phase separation. All‐atom simulations uncovered the presence of transient, interdomain interactions involving flexible linkers, RNA‐recognition motif (RRM) domains and a charged segment of disordered C‐terminal domain (CTD). CG simulations indicate these inter‐domain interactions which affect the conformational landscape of TDP‐43 in the dilute phase are also prevalent in the condensed phase. Finally, sequence and surface charge distribution analysis coupled with all‐atom simulations (at high salt) confirmed that the transient interdomain contacts are predominantly electrostatic in nature. Overall, our findings from multiscale simulations lead to a greater appreciation of the complex interaction network underlying the structural landscape and phase separation of TDP‐43.This article is protected by copyright. All rights reserved.

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Inter-domain interactions of TDP-43 as decoded by NMR

Cited by 30 publications

References 22 publications

RRM domain of ALS/FTD-causing FUS characteristic of irreversible unfolding spontaneously self-assembles into amyloid fibrils

RRM domain of ALS/FTD-causing FUS characteristic of irreversible unfolding spontaneously self-assembles into amyloid fibrils

Immunolocalization of TAR DNA‐binding protein of 43 kDa (TDP‐43) in mouse seminiferous epithelium

A complex network of interdomain interactions underlies the conformational ensemble of monomeric TDP‐43 and modulates its phase behavior

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