TDP-43 NTD can be induced while CTD is significantly enhanced by ssDNA to undergo liquid-liquid phase separation

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

doi:10.1016/j.bbrc.2018.03.121

Cited by 37 publications

(52 citation statements)

References 24 publications

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“…His tag removal consistently resulted in low yields of purified protein, possibly due to the high aggregation propensity at conditions necessary for efficacious thrombin activity. Although the presence of a histidine (with a pK a of ϳ6) likely has an influence on the pH dependence of the LCD (which does not naturally contain histidine), use of a His-tagged protein is in line with almost all previously published studies exploring the LLPS or aggregation properties of purified TDP-43 LCD (13,36,67). A notable exception is the study by Conicella et al (35) in which the authors removed the His tag but which still included artificial GH residues at the N terminus.…”

Section: Experimental Procedures Expression Purification and Preparmentioning

confidence: 62%

The role of liquid–liquid phase separation in aggregation of the TDP-43 low-complexity domain

Babinchak

Raza

Dumm

et al. 2019

Journal of Biological Chemistry

280

287

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Pathological aggregation of the transactive response DNAbinding protein of 43 kDa (TDP-43) is associated with several neurodegenerative disorders, including ALS, frontotemporal dementia, chronic traumatic encephalopathy, and Alzheimer's disease. TDP-43 aggregation appears to be largely driven by its low-complexity domain (LCD), which also has a high propensity to undergo liquid-liquid phase separation (LLPS). However, the mechanism of TDP-43 LCD pathological aggregation and, most importantly, the relationship between the aggregation process and LLPS remains largely unknown. Here, we show that amyloid formation by the LCD is controlled by electrostatic repulsion. We also demonstrate that the liquid droplet environment strongly accelerates LCD fibrillation and that its aggregation under LLPS conditions involves several distinct events, culminating in rapid assembly of fibrillar aggregates that emanate from within mature liquid droplets. These combined results strongly suggest that LLPS may play a major role in pathological TDP-43 aggregation, contributing to pathogenesis in neurodegenerative diseases.

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Section: Experimental Procedures Expression Purification and Preparmentioning

confidence: 62%

The role of liquid–liquid phase separation in aggregation of the TDP-43 low-complexity domain

Babinchak

Raza

Dumm

et al. 2019

Journal of Biological Chemistry

280

287

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“…Unlike pathological aggregates, which are hyperphosphorylated and ubiquitinated (Arai et al, 2006;Hasegawa et al, 2008), reversible formation of TDP-43 polymers through the NTD has been shown to be required for splicing activity (Afroz et al, 2017;Jiang et al, 2017;Mompeán et al, 2017; and to contribute to phase separation via liquid-droplet formation (Afroz et al, 2017;, thought to contribute to formation of cytoplasmic stress granules (SGs) (Molliex et al, 2015). The NTD is also the site of one of three predicted mitochondrial targeting sequences (Wang et al, 2016), conserved phosphosite Ser 48 (Rigbolt et al, 2011;, as well as potential, albeit weak, nucleotide binding (Chang et al, 2012;Qin et al, 2014;Mompeán et al, 2016c;Wang L. et al, 2018). Thus far, five 3-dimensional structures of the NTD have been published, including three monomeric (Mompeán et al, 2016c(Mompeán et al, , 2017Jiang et al, 2017) and two dimeric structures (Afroz et al, 2017;.…”

Section: The N-terminal Domain and Nuclear Localization Signal Organimentioning

confidence: 99%

“…Further complicating the picture is the recent identification of the role of reversible self-association, generally termed LLPS (liquid-liquid phase separation) which is thought to initiate formation of SGs and to which both the NTD and CTD seem to contribute (Conicella et al, 2016;Schmidt and Rohatgi, 2016;Afroz et al, 2017;Li et al, 2018;Wang L. et al, 2018;Babinchak et al, 2019). The conditions under which TDP-43 undergoes LLPS are highly sensitive to the conditions of the experiment (Conicella et al, 2016;Li et al, 2018;Wang L. et al, 2018). Moreover, interaction between the CTD and charged nucleic acids, specifically ssDNA, can increase the CTD's ability to undergo LLPS, thought to occur via the many aromatic and pi interactions (Wang L. et al, 2018).…”

Section: Liquid-liquid Phase Separation and Aggregationmentioning

confidence: 99%

“…The conditions under which TDP-43 undergoes LLPS are highly sensitive to the conditions of the experiment (Conicella et al, 2016;Li et al, 2018;Wang L. et al, 2018). Moreover, interaction between the CTD and charged nucleic acids, specifically ssDNA, can increase the CTD's ability to undergo LLPS, thought to occur via the many aromatic and pi interactions (Wang L. et al, 2018). Despite a lack of direct modulation of LLPS formation by charged residues, it was found that arginine residues played an integral role in changing the material properties and dynamics of the droplets formed by TDP-43.…”

Section: Liquid-liquid Phase Separation and Aggregationmentioning

confidence: 99%

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Structural Insights Into TDP-43 and Effects of Post-translational Modifications

François‐Moutal

Perez‐Miller

Scott

et al. 2019

Front. Mol. Neurosci.

107

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Transactive response DNA binding protein (TDP-43) is a key player in neurodegenerative diseases. In this review, we have gathered and presented structural information on the different regions of TDP-43 with high resolution structures available. A thorough understanding of TDP-43 structure, effect of modifications, aggregation and sites of localization is necessary as we develop therapeutic strategies targeting TDP-43 for neurodegenerative diseases. We discuss how different domains as well as posttranslational modification may influence TDP-43 overall structure, aggregation and droplet formation. The primary aim of the review is to utilize structural insights as we develop an understanding of the deleterious behavior of TDP-43 and highlight locations of established and proposed post-translation modifications. TDP-43 structure and effect on localization is paralleled by many RNA-binding proteins and this review serves as an example of how structure may be modulated by numerous compounding elements.

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“…Therefore for the LC domains, we have tested TDP-43 Cterminal LC domain over residues 263-414, FUS N-terminal 1-165 and 1-267 we previously characterized (15,28). Unfortunately, we found that the TDP-43 C-terminal LC domain has two disadvantages: 1) it only formed a small number of small liquid droplets at 25 ºC exactly as previously observed (26,30); 2) most likely due to the presence of a hydrophobic fragment 311-341 (15,26), visible aggregates formed upon adding ALS-causing mutants of SOD1 or PFN1. For FUS containing only the GQSY-rich prion-like domain, it only formed small liquid droplets at 25 ºC as previously reported (25), while FUS (1-267), the entire Nterminal LC domain of FUS with an extra Gly-rich region 166-267 could form dynamic droplets with large numbers and sizes at 25 ºC ( Fig.…”

Section: Identification Of a System Suitable For High-resolution Nmr mentioning

confidence: 58%

Misfolded proteins share a common capacity in disrupting LLPS organizing membrane-less organelles

Kang

Song

2018

Preprint

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Profilin-1 mutants cause ALS by gain of toxicity but the underlying mechanism remains unknown. Here we showed that three PFN1 mutants have differential capacity in disrupting dynamics of FUS liquid droplets underlying the formation of stress granules (SGs). Subsequently we extensively characterized conformations, dynamics and hydrodynamic properties of C71G-PFN1, FUS droplets and their interaction by NMR spectroscopy. C71G-PFN1 co-exists between the folded (55.2%) and unfolded (44.8%) states undergoing exchanges at 11.7 Hz, while its unfolded state non-specifically interacts with FUS droplets. Results together lead to a model for dynamic droplets to recruit misfolded proteins, which functions seemingly at great cost: simple accumulation of misfolded proteins within liquid droplets is sufficient to reduce their dynamics. Further aggregation of misfolded proteins within droplets might irreversibly disrupt/destroy structures and dynamics of droplets, as increasingly observed on SGs, an emerging target for various neurodegenerative diseases. Therefore, our study implies that other misfolded proteins might also share the capacity in disrupting LLPS.Amyotrophic lateral sclerosis (ALS) is the most common motor neuron disease, which was first described in 1869 but its mechanism still remains a great mystery (1). Human superoxide dismutases 1 (SOD1) was the first gene identified to be associated with familial ALS and so far more than 20 ALS-causative genes have been found (1-3). Like all other neurodegenerative diseases (4), ALS is also characterized by severe aggregation of these proteins in vivo and in vitro (1-9). The ALS-associated proteins can be categorized into two major groups: the first including SOD1 and profilin-1 (PFN1) whose wild types are wellfolded and only the ALS-associated mutants become aggregation-prone and toxic (1-3,7-10); while the second including FUS and TDP-43 containing the low-complexity (LC) domains whose wild types are already aggregation-prone and toxic (1,10-15). Very intriguingly, some ALS-causing mutants of the first group such as C71G-PFN1 have been previously revealed to trigger seed-dependent co-aggregation with the proteins in the second group including TDP-43 to manifest the prion-like propagandation (16), but so far the mechanism underlying this observation remains completely unknown.Progressive aggregation of thousands of non-specific proteins has also been shown to be associated with aging down to unicellular organisms (17,18). Although it remains largely elusive whether the aggregation of non-specific proteins also leads to aging by gain of toxicity, emerging evidence implies that to clean up aggregated proteins represents a key step for cells to rejuvenate as evidenced by the previous result that upon division of E. coli cells, the mother cell keeps protein aggregates so as to allow the daughter cell to be free of protein aggregates for rejuvenation (18); as well as by a recent report that for Caenorhabditis elegans, upon fertilization the sperm will send a signal to the egg to ...

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TDP-43 NTD can be induced while CTD is significantly enhanced by ssDNA to undergo liquid-liquid phase separation

Cited by 37 publications

References 24 publications

The role of liquid–liquid phase separation in aggregation of the TDP-43 low-complexity domain

The role of liquid–liquid phase separation in aggregation of the TDP-43 low-complexity domain

Structural Insights Into TDP-43 and Effects of Post-translational Modifications

Misfolded proteins share a common capacity in disrupting LLPS organizing membrane-less organelles

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