Issue 21, 2021

Phase separation of p53 precedes aggregation and is affected by oncogenic mutations and ligands

Abstract

Mutant p53 tends to form aggregates with amyloid properties, especially amyloid oligomers inside the nucleus, which are believed to cause oncogenic gain-of-function (GoF). The mechanism of the formation of the aggregates in the nucleus remains uncertain. The present study demonstrated that the DNA-binding domain of p53 (p53C) underwent phase separation (PS) on the pathway to aggregation under various conditions. p53C phase separated in the presence of the crowding agent polyethylene glycol (PEG). Similarly, mutant p53C (M237I and R249S) underwent PS; however, the process evolved to a solid-like phase transition faster than that in the case of wild-type p53C. The data obtained by microscopy of live cells indicated that transfection of mutant full-length p53 into the cells tended to result in PS and phase transition (PT) in the nuclear compartments, which are likely the cause of the GoF effects. Fluorescence recovery after photobleaching (FRAP) experiments revealed liquid characteristics of the condensates in the nucleus. Mutant p53 tended to undergo gel- and solid-like phase transitions in the nucleus and in nuclear bodies demonstrated by slow and incomplete recovery of fluorescence after photobleaching. Polyanions, such as heparin and RNA, were able to modulate PS and PT in vitro. Heparin apparently stabilized the condensates in a gel-like state, and RNA apparently induced a solid-like state of the protein even in the absence of PEG. Conditions that destabilize p53C into a molten globule conformation also produced liquid droplets in the absence of crowding. The disordered transactivation domain (TAD) modulated both phase separation and amyloid aggregation. In summary, our data provide mechanistic insight into the formation of p53 condensates and conditions that may result in the formation of aggregated structures, such as mutant amyloid oligomers, in cancer. The pathway of mutant p53 from liquid droplets to gel-like and solid-like (amyloid) species may be a suitable target for anticancer therapy.

Graphical abstract: Phase separation of p53 precedes aggregation and is affected by oncogenic mutations and ligands

Supplementary files

Article information

Article type
Edge Article
Submitted
27 Mar 2021
Accepted
19 Apr 2021
First published
26 Apr 2021
This article is Open Access

All publication charges for this article have been paid for by the Royal Society of Chemistry
Creative Commons BY-NC license

Chem. Sci., 2021,12, 7334-7349

Phase separation of p53 precedes aggregation and is affected by oncogenic mutations and ligands

E. C. Petronilho, M. M. Pedrote, M. A. Marques, Y. M. Passos, M. F. Mota, B. Jakobus, G. D. S. D. Sousa, F. Pereira da Costa, A. L. Felix, G. D. S. Ferretti, F. P. Almeida, Y. Cordeiro, T. C. R. G. Vieira, G. A. P. de Oliveira and J. L. Silva, Chem. Sci., 2021, 12, 7334 DOI: 10.1039/D1SC01739J

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