Molecular mechanisms of transthyretin-induced cytotoxicity
Creators: name: Hou, Xu
Publisher: name: Monash University
Year: 2017
Subjects: Uncategorized

description: Familial amyloidotic polyneuropathy (FAP) is a fatal neurodegenerative disease characterized by the deposition of transthyretin (TTR) amyloid. The present study aimed to investigate the molecular mechanisms for TTR-induced cytotoxicity and factors involved in TTR amyloidogenesis. It was hypothesized that binding of TTR to the plasma membrane plays a crucial role in the molecular pathogenesis of FAP, whereby TTR alters the organization of the plasma membrane, inducing changes in the activity of membrane proteins which ultimately lead to cell death. To examine the role of the plasma membrane in TTR-induced cytotoxicity, the binding of TTR to a plasma membrane-enriched fraction isolated neuroblastoma cells was examined by surface plasmon resonance (SPR). TTR bound predominantly to the lipids on membrane surface via electrostatic interactions. A correlation was found between membrane binding of TTR and TTR-induced cytotoxicity, suggesting that binding to membrane lipids may be an initial step in the cascade of TTRinduced cytotoxic events. Upon binding to membrane lipids, TTR was found to increase the fluidity of the plasma membrane. The effect of TTR on cytoplasmic Ca2+ homeostasis was also examined in neuroblastoma cells. TTR rapidly induced an increase in the concentration of intracellular Ca2+ ([Ca2+]i) due to an influx of extracellular Ca2+, mainly via L- and N-type voltage-gated calcium channels (VGCCs), suggesting that dysregulation of [Ca2+]i may play an important role in TTR-induced cytotoxicity. As the effect on [Ca2+]i was correlated with the concentration of TTR oligomers (&lt;100 nm in diameter) in the preparation, oligomers are likely to be the major cytotoxic form of TTR. Based on these results, a hypothetical mechanism has been proposed that binding of TTR to the plasma membrane may disrupt lipid organization in the membrane, altering the activity of ion channels. In addition, the effects of phospholipids and heparin on TTR aggregation were examined. Anionic phospholipids accelerated nucleation (oligomerization) of TTR without influencing the total amount of amyloid formation, while zwitterionic phospholipids had no effect. The promoting effect of phospholipids was correlated with their binding capacities for TTR. Heparin and a number of anions also promoted TTR aggregation by increasing both the rate of nucleation and the amount of aggregation. It is possible that electrostatic interactions between positively charged residues of TTR and negatively charged headgroups of anionic phospholipids or negatively charged groups of heparin/heparan sulfate (e.g. sulfate and carboxylate) may be involved in TTR amyloidogenesis and deposition by providing a favourable local environment for TTR misfolding and aggregation. The findings of the present study have provided insights into the molecular mechanisms of TTR-induced cytotoxicity and TTR amyloidogenesis, and identified TTR oligomers as a potential target for therapeutic intervention. In addition, by supporting the idea that soluble oligomers are the common cytotoxic species in amyloidotic conditions, the present study has expanded our understanding of the broader field of amyloidosis.