Publicação
TDP−43 self-assembly in neurodegenerative proteinopathies: Chaperone regulation of phase separation and aggregation of the TDP−43 low complexity domain
| Resumo: | TDP−43 is a ubiquitous nucleic acid-binding protein that undergoes physiological phase sepa-ration, essential for its roles in RNA metabolism. TDP−43 proteinopathy is a hallmark of several neu-rodegenerative diseases (NDDs), including amyotrophic lateral sclerosis (ALS) and frontotemporal de-mentia, with increasing evidence linking it to Alzheimer’s disease. It is characterized by nuclear deple-tion, cytoplasmic mislocalization, and accumulation of C-terminal fragments primarily composed of its disordered low-complexity domain (TDP−43−LCD). Once mislocalized to the cytoplasm, TDP−43−LCD undergoes aberrant phase separation and forms amyloid aggregates. S100B is an abundant calcium-binding protein in the brain, implicated in NDDs. Recent find-ings have revealed its activity as a molecular chaperone. S100B binds disordered, amyloidogenic pro-teins such as Aβ and tau in a calcium-dependent manner, inhibiting their pathological self-assembly. Given that S100B and TDP−43 are expressed in overlapping cellular compartments, and considering preliminary in silico data supporting a direct interaction between TDP−43 and S100B at the chaperone substrate-binding cleft, this study aimed to investigate whether S100B plays a regulatory role in the aggregation and phase separation of TDP−43−LCD. By combining molecular structural modelling with experimental determination of binding af-finity, we demonstrate that TDP−43−LCD acts as a client protein within the regulatory cleft of S100B, binding with high affinity to its Ca2+-bound form. Furthermore, S100B inhibits RNA-induced phase separation of TDP−43−LCD and can partially dissolve preformed condensates, without altering their liquid dynamics. Strikingly, S100B promotes both condensation and decondensation of TDP−43−LCD, a dual effect mediated by distinct interaction modes that favour either assembly or disassembly. Struc-tural analysis suggests that key residues involved in TDP−43−LCD phase separation directly interact with the substrate-binding cleft of S100B. Finally, we show that S100B inhibits aggregation of TDP−43−LCD. Altogether, our results advanced the understanding of S100B’s role in TDP−43 self-assembly, revealing its dual function: as an inhibitor of pathological aggregation and as a regulator of physiological phase separation dynamics. |
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| Autores principais: | Silva,Carolina Filipa Monteiro |
| Assunto: | Phase separation amyloid aggregation protein-protein interactions molecular chaperones RNA-binding proteins |
| Ano: | 2025 |
| País: | Portugal |
| Tipo de documento: | dissertação de mestrado |
| Tipo de acesso: | acesso embargado |
| Instituição associada: | Universidade de Lisboa |
| Idioma: | inglês |
| Origem: | Repositório da Universidade de Lisboa |
| Resumo: | TDP−43 is a ubiquitous nucleic acid-binding protein that undergoes physiological phase sepa-ration, essential for its roles in RNA metabolism. TDP−43 proteinopathy is a hallmark of several neu-rodegenerative diseases (NDDs), including amyotrophic lateral sclerosis (ALS) and frontotemporal de-mentia, with increasing evidence linking it to Alzheimer’s disease. It is characterized by nuclear deple-tion, cytoplasmic mislocalization, and accumulation of C-terminal fragments primarily composed of its disordered low-complexity domain (TDP−43−LCD). Once mislocalized to the cytoplasm, TDP−43−LCD undergoes aberrant phase separation and forms amyloid aggregates. S100B is an abundant calcium-binding protein in the brain, implicated in NDDs. Recent find-ings have revealed its activity as a molecular chaperone. S100B binds disordered, amyloidogenic pro-teins such as Aβ and tau in a calcium-dependent manner, inhibiting their pathological self-assembly. Given that S100B and TDP−43 are expressed in overlapping cellular compartments, and considering preliminary in silico data supporting a direct interaction between TDP−43 and S100B at the chaperone substrate-binding cleft, this study aimed to investigate whether S100B plays a regulatory role in the aggregation and phase separation of TDP−43−LCD. By combining molecular structural modelling with experimental determination of binding af-finity, we demonstrate that TDP−43−LCD acts as a client protein within the regulatory cleft of S100B, binding with high affinity to its Ca2+-bound form. Furthermore, S100B inhibits RNA-induced phase separation of TDP−43−LCD and can partially dissolve preformed condensates, without altering their liquid dynamics. Strikingly, S100B promotes both condensation and decondensation of TDP−43−LCD, a dual effect mediated by distinct interaction modes that favour either assembly or disassembly. Struc-tural analysis suggests that key residues involved in TDP−43−LCD phase separation directly interact with the substrate-binding cleft of S100B. Finally, we show that S100B inhibits aggregation of TDP−43−LCD. Altogether, our results advanced the understanding of S100B’s role in TDP−43 self-assembly, revealing its dual function: as an inhibitor of pathological aggregation and as a regulator of physiological phase separation dynamics. |
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