Characterization of p53 p.T253I as a pathogenic mutation underlying Li-Fraumeni Syndrome

Nathaniel C. Holcomb; Amanda M. Harrington; Hong Pu; Berina Halilovic; Nathan R. Shelman; Shulin Zhang; Catherine Sears; Terra Armstrong; Brent Shelton; Lauren Corum; John A. D'Orazio

doi:10.1371/journal.pone.0320036

Characterization of p53 p.T253I as a pathogenic mutation underlying Li-Fraumeni Syndrome

Nathaniel C. Holcomb, Amanda M. Harrington, Hong Pu, Berina Halilovic, Nathan R. Shelman, Shulin Zhang, Catherine Sears, Terra Armstrong, Brent Shelton, Lauren Corum, John A. D'Orazio

Research output: Contribution to journal › Article › peer-review

Abstract

We identified a germline TP53 c.758C > T (p.T253I) mutation in the TP53 tumor suppressor gene in a pediatric adrenocortical carcinoma (ACC) patient. Characteristic of pathogenic p53 mutations, we observed upregulation of total p53 protein levels in the patient’s ACC and concurrent suppression of the wild-type (WT) TP53 allele. As ACC can be associated with Li-Fraumeni Syndrome (LFS) and the mutation has not yet been linked to LFS, we sought to characterize the functionality of the T253I mutation. We acquired p53^-/- HEK293 cells and stably transduced them with GFP-tagged wild type (T253) or T253I p53 as well as two established pathogenic p53 mutants (C176Y and R213X). Compared to p53 WT, levels of T253I p53 increased while MDM2 levels decreased, suggesting a loss of MDM2-mediated regulation of T253I p53. Additionally, T253I showed a reduction in DNA damage responsive events, diminished DNA binding capabilities, and blunted transactivation capacity. These experimental data lead us to conclude that T253I represents a pathologic variant in TP53 that may predispose to LFS-associated tumors.

Original language	English
Article number	e0320036
Journal	PLoS ONE
Volume	20
Issue number	12 December
DOIs	https://doi.org/10.1371/journal.pone.0320036
State	Published - Dec 2025

Bibliographical note

Publisher Copyright:
Copyright: © 2025 Holcomb et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

Funding

Kentucky Pediatric Cancer Research Trust Fund NCI P30 CA177558 NIH P20 GM121327. We are grateful to the Biostatistics and Bioinformatics, Flow Cytometry and Immune Monitoring, Biospecimen Procurement and Translational Pathology, Cancer Research and Informatics, and Oncogenomics Shared Resource Facilities of the University of Kentucky Markey Cancer Center for their technical and scientific contributions to the work. We thank the COBRE Imaging Core facility at the University of Kentucky for providing service with microscopes and imaging systems. We are indebted to the Clinical Research Office of the Division of Pediatric Hematology/Oncology at the University of Kentucky for their efforts in accrual and data management for Project Inherited Cancer Risk, the clinical study to identify and manage children, adolescents and young adults with cancer predisposition syndromes. Finally, we are immensely grateful to the patients and families who consented to take part in the “Project Inherited Cancer Risk” study at the University of Kentucky by which these variants were identified. Funding: Kentucky Pediatric Cancer Research Trust Fund NCI P30 CA177558 NIH P20 GM121327.

Funders	Funder number
Departmentof Bioinformatics and Biostatistics
University of Kentucky Markey Comprehensive Cancer Center
Clinical Research Office of the Division of Pediatric Hematology
University of Kentucky
Kura Oncology Incorporated
Kentucky Pediatric Cancer Research Trust Fund NCI	P30 CA177558 NIH P20 GM121327

ASJC Scopus subject areas

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title = "Characterization of p53 p.T253I as a pathogenic mutation underlying Li-Fraumeni Syndrome",

abstract = "We identified a germline TP53 c.758C > T (p.T253I) mutation in the TP53 tumor suppressor gene in a pediatric adrenocortical carcinoma (ACC) patient. Characteristic of pathogenic p53 mutations, we observed upregulation of total p53 protein levels in the patient{\textquoteright}s ACC and concurrent suppression of the wild-type (WT) TP53 allele. As ACC can be associated with Li-Fraumeni Syndrome (LFS) and the mutation has not yet been linked to LFS, we sought to characterize the functionality of the T253I mutation. We acquired p53-/- HEK293 cells and stably transduced them with GFP-tagged wild type (T253) or T253I p53 as well as two established pathogenic p53 mutants (C176Y and R213X). Compared to p53 WT, levels of T253I p53 increased while MDM2 levels decreased, suggesting a loss of MDM2-mediated regulation of T253I p53. Additionally, T253I showed a reduction in DNA damage responsive events, diminished DNA binding capabilities, and blunted transactivation capacity. These experimental data lead us to conclude that T253I represents a pathologic variant in TP53 that may predispose to LFS-associated tumors.",

author = "Holcomb, \{Nathaniel C.\} and Harrington, \{Amanda M.\} and Hong Pu and Berina Halilovic and Shelman, \{Nathan R.\} and Shulin Zhang and Catherine Sears and Terra Armstrong and Brent Shelton and Lauren Corum and D'Orazio, \{John A.\}",

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AU - Shelman, Nathan R.

AU - Zhang, Shulin

AU - Sears, Catherine

AU - Armstrong, Terra

AU - Shelton, Brent

AU - Corum, Lauren

AU - D'Orazio, John A.

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N2 - We identified a germline TP53 c.758C > T (p.T253I) mutation in the TP53 tumor suppressor gene in a pediatric adrenocortical carcinoma (ACC) patient. Characteristic of pathogenic p53 mutations, we observed upregulation of total p53 protein levels in the patient’s ACC and concurrent suppression of the wild-type (WT) TP53 allele. As ACC can be associated with Li-Fraumeni Syndrome (LFS) and the mutation has not yet been linked to LFS, we sought to characterize the functionality of the T253I mutation. We acquired p53-/- HEK293 cells and stably transduced them with GFP-tagged wild type (T253) or T253I p53 as well as two established pathogenic p53 mutants (C176Y and R213X). Compared to p53 WT, levels of T253I p53 increased while MDM2 levels decreased, suggesting a loss of MDM2-mediated regulation of T253I p53. Additionally, T253I showed a reduction in DNA damage responsive events, diminished DNA binding capabilities, and blunted transactivation capacity. These experimental data lead us to conclude that T253I represents a pathologic variant in TP53 that may predispose to LFS-associated tumors.

AB - We identified a germline TP53 c.758C > T (p.T253I) mutation in the TP53 tumor suppressor gene in a pediatric adrenocortical carcinoma (ACC) patient. Characteristic of pathogenic p53 mutations, we observed upregulation of total p53 protein levels in the patient’s ACC and concurrent suppression of the wild-type (WT) TP53 allele. As ACC can be associated with Li-Fraumeni Syndrome (LFS) and the mutation has not yet been linked to LFS, we sought to characterize the functionality of the T253I mutation. We acquired p53-/- HEK293 cells and stably transduced them with GFP-tagged wild type (T253) or T253I p53 as well as two established pathogenic p53 mutants (C176Y and R213X). Compared to p53 WT, levels of T253I p53 increased while MDM2 levels decreased, suggesting a loss of MDM2-mediated regulation of T253I p53. Additionally, T253I showed a reduction in DNA damage responsive events, diminished DNA binding capabilities, and blunted transactivation capacity. These experimental data lead us to conclude that T253I represents a pathologic variant in TP53 that may predispose to LFS-associated tumors.

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Characterization of p53 p.T253I as a pathogenic mutation underlying Li-Fraumeni Syndrome

Abstract

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