AbstractThe transition between the native and amyloid states of proteins can proceed via a deposition pathway through oligomeric intermediates or via a condensation pathway through liquid droplet intermediates generated through liquid-liquid phase separation. The maturation of these droplet intermediates into ordered assemblies has been associated with human disease, including in particular amyotrophic lateral sclerosis (ALS), although the mechanisms of toxicity have not been yet clarified. Here we investigate the processes by which ALS-related mutations give rise to cytotoxicity along the condensation pathway. Based on the sequence-determinants of the different types of interactions stabilising the droplet and amyloid states, we accurately predict the levels of toxicity of about 50,000 deep mutagenesis variants of TDP-43 prion-like domain. We find that condensation is not typically initiated by structural ordering, but rather through non-specific interactions, and that the cytotoxicity of ALS-related TDP-43 mutations stems from promiscuous interactions within the droplet intermediates, rather than from the mature aggregates. These results provide insights into the mechanisms by which condensates convert into amyloids and their links with human disease.SignificanceProtein liquid-liquid phase separation underlies the formation of functional protein condensates, which upon dysregulation can mature into cytotoxic amyloid-containing aggregates. The sequence-based principles governing this pathway, and the mechanisms giving rise to cytotoxicity, however, are still not known in detail. Here, based on the different amino acid codes leading to the droplet and amyloid states, we show how one can predict the toxicity of the intermediate states along the condensation pathway. Our results highlight that this toxicity originates from the interaction promiscuity of amyloid-containing intermediates, in particular those with ALS-related mutations, rather than from the mature amyloid state. These results contribute to our understanding of the mechanisms through which TDP-43 mutations are linked to ALS.
Liquid-liquid phase separation in secondary organic aerosol particles produced from α-pinene ozonolysis and α-pinene photo-oxidation with/without ammonia
