(507g) The Disordered Domain of the Tumor Suppressor p53 Is a Passive Bystander in Mesoscopic Aggregation and Fibrillization.

Cancer is one of the leading causes of death worldwide. The protein p53 is an important tumor suppressor and is known as the guardian of the genome. This protein is a transcription factor that binds to DNA and controls multiple signaling pathway to determine the cell fate. More than 50 % of human cancers are related to mutations in the p53 DNA-binding domain of p53. Recent studies suggest that p53 aggregation is a key factor in cancer development and the majority of the p53 mutants have an exaggerated propensity to aggregate. Mechanistic details on the nucleation and growth of p53 amyloid fibrils, however, are missing. One of the crucial open questions is on the role of the substantial (it holds about 60% of the aminoacid residues) disordered domain of p53 in aggregation; disordered domains in other proteins are deemed indispensable for the formation of macroscopic dense liquids and the function of numerous membrane-less organelles in the nucleoplasm and cytoplasm. We explore the aggregation mechanism of the p53 DBD, a construct limited to the ordered core domain of the protein. We demonstrate that this p53 construct does not form dense liquids, but rather assembles into mesoscopic p53-rich clusters compromised of about 1000 p53 molecules. Transmission electron microscopy confirms the cluster size and likely liquid consistency and demonstrates that the clusters host and facilitate the nucleation and growth of p53 amyloid fibrils, linear aggregates that irreversibly store p53 chains. Two-step nucleation of p53 amyloids suggests means to control fibrillization and the associated pathologies through modifying the cluster characteristics. Importantly, cluster formation by p53 DBD implies that disordered segments are not a sine qua non for clustering and enhanced fibrillization, in sharp contradistinction with dense liquid of other proteins. This later conclusion is supported by molecular simulations of the structural destabilization and aggregation of the core domain. In a broader context, our findings exemplify interactions between distinct protein phases that activate complex physicochemical mechanisms operating in biological systems.