One-Dimensional Search Dynamics of Tumor Suppressor p53 Regulated by a Disordered C-Terminal Domain

Murata, Agato; Itoh, Yuji; Mano, Eriko; Kanbayashi, Saori; Igarashi, Chihiro; Takahashi, Hiroto; Takahashi, Satoshi; Kamagata, Kiyoto

doi:10.1016/j.bpj.2017.04.038

Cited by 28 publications

(52 citation statements)

References 80 publications

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“…We used a thermostable mutant termed a full-length pseudo wild-type p53 (FL-p53) to reduce the irreversible aggregation found in wild-type p53 5 . At a FL-p53 concentration of 100 μM and pH 7.9 [5][6][7][8][9] , we detected an increase in light scattering at 350 nm. However, imaging of the solutions using a differential interference contrast (DIC) microscope could not identify any μm-sized assemblies.…”

Section: Resultsmentioning

confidence: 81%

“…p53 droplet formation is mediated by the NT and CT disordered domains. In order to identify the domains that participate in droplet formation, we investigated three deletion p53 mutants: the NTCoreTet mutant (residues 1-363) lacking the CT domain, the CoreTetCT mutant (residues 94-393) lacking the NT domain, and the TetCT mutant (residues 293-393) lacking the NT and core domains 6 (Fig. 2).…”

Section: Resultsmentioning

confidence: 99%

“…2). The three mutants were purified as described in our previous report 6 . We investigated droplet formation of the deletion mutants at 12 μM at varying pH and salt concentrations and in the absence of dextran.…”

Section: Resultsmentioning

confidence: 99%

“…The results suggested that the specific p53/dsDNA sp complex, in which p53 is bound to DNA via the core domain [51][52][53][54][55] , did not form droplets probably due to the electrostatic repulsion between DNAs present in the complexes. Similarly, the non-specific 30-bp double-stranded DNA, dsDNA nsp , suppressed droplet formation at 2.5 μM, suggesting that the non-specific p53/ dsDNA nsp complex, in which p53 is bound to DNA via the CT and core domains 3,5,6 , did not form the droplet (Fig. 5d).…”

Section: Droplet Formation Of P53 Is Regulated By Crowding Nucleic Amentioning

confidence: 91%

“…The TetCT mutant corresponds to residues 293−393 of FL-p53 with an additional N-terminal cysteine 5 . The NTCoreTet mutant corresponds to residues 1-363 6 . The S392E mutant was constructed based on FL-p53 8 .…”

Section: Methodsmentioning

confidence: 99%

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Liquid-like droplet formation by tumor suppressor p53 induced by multivalent electrostatic interactions between two disordered domains

Kamagata

Kanbayashi

Honda

et al. 2020

Sci Rep

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early in vivo studies demonstrated the involvement of a tumor-suppressing transcription factor, p53, into cellular droplets such as Cajal and promyelocytic leukemia protein bodies, suggesting that the liquid-liquid phase separation (LLPS) might be involved in the cellular functions of p53. To examine this possibility, we conducted extensive investigations on the droplet formation of p53 in vitro. First, p53 itself was found to form liquid-like droplets at neutral and slightly acidic pH and at low salt concentrations. Truncated p53 mutants modulated droplet formation, suggesting the importance of multivalent electrostatic interactions among the N-terminal and C-terminal domains. Second, FRET efficiency measurements for the dimer mutants of p53 revealed that distances between the core domains and between the C-terminal domains were modulated in an opposite manner within the droplets. Third, the molecular crowding agents were found to promote droplet formation, whereas ssDNA, dsDNA, and ATP, to suppress it. Finally, the p53 mutant mimicking posttranslational phosphorylation did not form the droplets. We conclude that p53 itself has a potential to form droplets that can be controlled by cellular molecules and by posttranslational modifications, suggesting that LLPS might be involved in p53 function. Tumor suppressor p53 is a multifunctional transcription factor that induces cell cycle arrest, DNA repair or apoptosis upon binding to its target DNA sequence. In 50% of human cancers, mutations on p53 are found to hamper its binding to the target sequence. Accordingly, extensive investigations have been conducted to characterize the functions as well as malfunctions of p53. However, an important aspect of p53, namely its involvement in liquid-like droplets, is still largely unresolved. In fact, p53 has long been known to be uptaken into cellular droplets such as Cajal and promyelocytic leukemia protein (PML) bodies. In this report, we describe that p53 itself can form liquid-like droplets upon the control of solution conditions, suggesting a possible involvement of the p53 droplets in the cellular environment. The primary function of p53 is the accommodation of various posttranslational modifications, termed activation, which in turn triggers the search for and the binding to its target DNA sequence, leading to the expression of downstream genes 1. p53 is composed of the N-terminal (NT) (residues 1-95), the core (95-293), the linker (293-326), the tetramerization (Tet) (326-357), and the C-terminal (CT) (357-393) domains. p53 slides along nonspecific DNA by attaching the CT domain to the DNA and by hopping the core domain 2-4. The sliding of p53 occurs in two modes 5,6 , in which the CT, core and linker domains are differently in contact with the DNA 7. The recognition efficiency of the target sequence by the sliding p53 is low, but is enhanced by the activation of p53 8. Furthermore, the sliding p53 can transfer from one DNA strand to another using the CT domain 9. In addition,

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Section: Resultsmentioning

confidence: 81%

Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

Section: Droplet Formation Of P53 Is Regulated By Crowding Nucleic Amentioning

confidence: 91%

Section: Methodsmentioning

confidence: 99%

See 3 more Smart Citations

Liquid-like droplet formation by tumor suppressor p53 induced by multivalent electrostatic interactions between two disordered domains

Kamagata

Kanbayashi

Honda

et al. 2020

Sci Rep

Self Cite

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A mini-review of the diffusion dynamics of DNA-binding proteins: experiments and models

Park

Lee

Durang

2021

J. Korean Phys. Soc.

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In the course of various biological processes, specific DNA-binding proteins must efficiently find a particular target sequence/ protein or a damaged site on the DNA. DNA-binding proteins perform this task based on diffusion. Nevertheless, investigations over recent decades have found that the diffusion dynamics of DNA-binding proteins are generally complicated and, further, protein specific. In this review, we collect experimental and theoretical studies that quantify the diffusion dynamics of DNA-binding proteins and review them from the viewpoint of diffusion processes.

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Ser392 phosphorylation modulated a switch between p53 and transcriptional condensates

Dai

Chen

et al. 2022

Biochimica et Biophysica Acta (BBA) - Gene Regulatory Mechanism

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One-Dimensional Search Dynamics of Tumor Suppressor p53 Regulated by a Disordered C-Terminal Domain

Cited by 28 publications

References 80 publications

Liquid-like droplet formation by tumor suppressor p53 induced by multivalent electrostatic interactions between two disordered domains

Liquid-like droplet formation by tumor suppressor p53 induced by multivalent electrostatic interactions between two disordered domains

A mini-review of the diffusion dynamics of DNA-binding proteins: experiments and models

Ser392 phosphorylation modulated a switch between p53 and transcriptional condensates

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