Skip to main content
Key Specialties
Cardiology Diabetology Endocrinology Gastroenterology Geriatrics and Gerontology Gynecology and Obstetrics Hematology Infectious Disease Internal Medicine Nephrology Neurology Oncology Pediatrics Respiratory Medicine Rheumatology Surgery
Congress Coverage
2024 ACC Congress 2023 ESMO Congress 2023 EASD Congress 2023 ERS Congress 2023 ESC Congress
Clinical Collections
Advances in Alzheimer’s

Keynote webinar | Spotlight on medication adherence

LIVE: Thursday 27th June 2024, 18:00-19:30 (CEST)
Join our expert panel to discover why you need to understand the drivers of non-adherence in your patients, and how you can optimize medication adherence in your clinics to drastically improve patient outcomes.
ADVANCED SEARCH
Log in
Top
Radiation Oncology
Published in: Radiation Oncology 1/2022

Open Access 01-12-2022 | Research

ΔNp63α transcriptionally represses p53 target genes involved in the radiation-induced DNA damage response

ΔNp63α may cause genomic instability in epithelial stem cells

Authors: Ken-ichi Kudo, Naohiro Tsuyama, Kento Nagata, Tatsuhiko Imaoka, Daisuke Iizuka, Misaki Sugai-Takahashi, Moe Muramatsu, Akira Sakai

Published in: Radiation Oncology | Issue 1/2022

Login to get access

Abstract

Background

The DNA damage response (DDR) is a mechanism that protects cells against radiation-induced oxidative DNA damage by causing cell cycle arrest and apoptosis. TP63 is a member of the tumour suppressor TP53 gene family, and ΔNp63α, a TP63 splicing variant, is constitutively expressed in the stem cell-containing basal layer of stratified epithelial tissues, including the mammary gland, where it plays a critical role in stemness and tissue development. ΔNp63α has been reported to transcriptionally inhibit the tumour suppression protein p53. This p53-repressive activity may cause genomic instability in epithelial stem cells exposed to radiation. In this study, we analysed the inhibitory effect of ΔNp63α on radiation-induced DDR.

Methods

To elucidate the role of the p53-repressive effect of ΔNp63α in radiation response, we performed a p63-siRNA knockdown experiment using human mammary epithelial cells (HMECs) expressing ΔNp63α and then performed ectopic and entopic expression experiments using human induced pluripotent stem cells (hiPSCs). After irradiation, the expression of DDR-related genes and proteins in ΔNp63α-expressing and control cells was analysed by RT–qPCR, Western blotting, and flow cytometry.

Results

The mRNA/protein expression levels of BAX and p21 were significantly increased in p63-siRNA-treated HMECs (sip63) after X-ray irradiation (4 Gy, 0.7 Gy/min) but not in scramble-siRNA treated HMECs (scr). Transcriptomic analysis showed decreased RNA expression of cell cycle-related genes and increased expression of programmed cell death-related genes in sip63 cells compared to scr cells. Furthermore, flow cytometric analysis revealed an increase in apoptotic cells and a decrease in 5-ethynyl-2´-deoxyuridine uptake in sip63 cells compared to scr cells. On the other hand, both the ectopic and entopic expression of ΔNp63α in apoptosis-sensitive hiPSCs reduced the expression levels of BAX after irradiation and significantly decreased the number of apoptotic cells induced by radiation.

Conclusion

Taken together, these results indicate that ΔNp63α represses p53-related radiation-induced DDR, thereby potentially causing genomic instability in epithelial stem cells.
Appendix
Available only for authorised users
Literature
1.
Haupt Y, Maya R, Kazaz A, Oren M. Mdm2 promotes the rapid degradation of p53. Nature. 1997;387:296–9.PubMedCrossRef
2.
Valerie K, Povirk LF. Regulation and mechanisms of mammalian double-strand break repair. Oncogene. 2003;22:5792–812.PubMedCrossRef
3.
El-Deiry WS, Tokino T, Velculescu VE, Levy DB, Parsons R, Trent JM, et al. WAF1, a potential mediator of p53 tumor suppression. Cell. 1993;75:817–25.PubMedCrossRef
4.
5.
Kroemer G, Galluzzi L, Brenner C. Mitochondrial membrane permeabilization in cell death. Physiol Rev. 2007;87:99–163.PubMedCrossRef
6.
Yang A, Kaghad M, Wang Y, Gillett E, Fleming MD, Dötsch V, et al. p63, a p53 homolog at 3q27-29, encodes multiple products with transactivating, death-inducing, and dominant-negative activities. Mol Cell. 1998;2:305–16.PubMedCrossRef
7.
Osada M, Ohba M, Kawahara C, Ishioka C, Kanamaru R, Katoh I, et al. Cloning and functional analysis of human p51, which structurally and functionally resembles p53. Nat Med. 1998;4:839–43.PubMedCrossRef
8.
Westfall MD, Mays DJ, Sniezek JC, Pietenpol JA. The Delta Np63 alpha phosphoprotein binds the p21 and 14-3-3 sigma promoters in vivo and has transcriptional repressor activity that is reduced by Hay-Wells syndrome-derived mutations. Mol Cell Biol. 2003;23:2264–76.PubMedPubMedCentralCrossRef
9.
10.
11.
Cai J, Chen S, Yi M, Tan Y, Peng Q, Ban Y, et al. ∆Np63α is a super enhancer-enriched master factor controlling the basal-to-luminal differentiation transcriptional program and gene regulatory networks in nasopharyngeal carcinoma. Carcinogenesis. 2020;41:1282–93.PubMedCrossRef
12.
Gaiddon C, Lokshin M, Ahn J, Zhang T, Prives C. A subset of tumor-derived mutant forms of p53 down-regulate p63 and p73 through a direct interaction with the p53 core domain. Mol Cell Biol. 2001;21:1874–87.PubMedPubMedCentralCrossRef
13.
Cam M, Gardner HL, Roberts RD, Fenger JM, Guttridge DC, London CA, et al. ∆Np63 mediates cellular survival and metastasis in canine osteosarcoma. Oncotarget. 2016;7:48533–46.PubMedPubMedCentralCrossRef
14.
Min S, Oyelakin A, Gluck C, Bard JE, Song EC, Smalley K, et al. p63 and its target follistatin maintain salivary gland stem/progenitor cell function through TGF-β/Activin signaling. iScience. 2020;23:101524.PubMedPubMedCentralCrossRef
15.
Gallant-Behm CL, Ramsey MR, Bensard CL, Nojek I, Tran J, Liu M, et al. ∆Np63α represses anti-proliferative genes via H2A.Z deposition. Genes Dev. 2012;26:2325–36.PubMedPubMedCentralCrossRef
16.
Melino G, Memmi -EM, Pelicci PG, Bernassola F. Maintaining epithelial stemness with p63. Sci Signal. 2015;8:re9.PubMedCrossRef
17.
Stingl J, Eirew P, Ricketson I, Shackleton M, Vaillant F, Choi D, et al. Purification and unique properties of mammary epithelial stem cells. Nature. 2006;439:993–7.PubMedCrossRef
18.
Shackleton M, Vaillant F, Simpson KJ, Stingl J, Smyth GK, Asselin-Labat ML, et al. Generation of a functional mammary gland from a single stem cell. Nature. 2006;439:84–8.PubMedCrossRef
19.
Kurinna S, Seltmann K, Bachmann AL, Schwendimann A, Thiagarajan L, Hennig P, et al. Interaction of the NRF2 and p63 transcription factors promotes keratinocyte proliferation in the epidermis. Nucleic Acids Res. 2021;49:3748–63.PubMedPubMedCentralCrossRef
20.
Centonze A, Lin S, Tika E, Sifrim A, Fioramonti M, Malfait M, et al. Heterotypic cell-cell communication regulates glandular stem cell multipotency. Nature. 2020;584:608–13.PubMedPubMedCentralCrossRef
21.
22.
Niwa O, Barcellos-Hoff MH, Globus RK, Harrison JD, Hendry JH, Jacob P, et al. ICRP publication 131: stem cell biology with respect to carcinogenesis aspects of radiological protection. Ann ICRP. 2015;44:7–357.PubMedCrossRef
23.
Preston DL, Ron E, Tokuoka S, Funamoto S, Nishi N, Soda M, et al Solid cancer incidence in atomic bomb survivors: 1958–1998. Radiat Res. 2007;168:1–64.Kim M, Mun H, Sung CO, Cho EJ, Jeon HJ, Chun SM, et al. Patient-derived lung cancer organoids as in vitro cancer models for therapeutic screening. Nat Commun. 2019;10:3991.
24.
Travis WD, Brambilla E, Noguchi M, Nicholson AG, Geisinger K, Yatabe Y, et al. Diagnosis of lung cancer in small biopsies and cytology: implications of the 2011 International Association for the Study of Lung Cancer/American Thoracic Society/European Respiratory Society classification. Arch Pathol Lab Med. 2013;137:668–84.PubMedCrossRef
25.
Kim M, Mun H, Sung CO, Cho EJ, Jeon HJ, Chun SM, et al. Patient-derived lung cancer organoids as in vitro cancer models for therapeutic screening. Nat Commun. 2019;10:3991.PubMedPubMedCentralCrossRef
26.
Chang CH, Zhang M, Rajapakshe K, Coarfa C, Edwards D, Huang S, et al. Mammary stem cells and tumor-initiating cells are more resistant to apoptosis and exhibit increased DNA repair activity in response to DNA damage. Stem Cell Reports. 2015;5:378–91.PubMedPubMedCentralCrossRef
27.
Kudo KI, Takabatake M, Nagata K, Nishimura Y, Daino K, Iizuka D, et al. Flow cytometry definition of rat mammary epithelial cell populations and their distinct radiation responses. Radiat Res. 2020;194:22–37.PubMedCrossRef
28.
Tsuyama N, Abe Y, Yanagi A, Yanai Y, Sugai M, Katafuchi A, et al. Induction of t(11;14) IgH enhancer/promoter-cyclin D1 gene translocation using CRISPR/Cas9. Oncol Lett. 2019;18:275–82.PubMedPubMedCentral
29.
Ge SX, Son EW, Yao R. iDEP: an integrated web application for differential expression and pathway analysis of RNA-Seq data. BMC Bioinformatics. 2018;19:534.PubMedPubMedCentralCrossRef
30.
Miyake T, Shimada M, Matsumoto Y, Okino A. DNA damage response after ionizing radiation exposure in skin keratinocytes derived from human-induced pluripotent stem cells. Int J Radiat Oncol Biol Phys. 2019;105:193–205.PubMedCrossRef
31.
Romano RA, Ortt K, Birkaya B, Smalley K, Sinha S. An active role of the DeltaN isoform of p63 in regulating basal keratin genes K5 and K14 and directing epidermal cell fate. PLoS One. 2009;4:e5623.PubMedPubMedCentralCrossRef
32.
Imaoka T, Nishimura M, Daino K, Hosoki A, Takabatake M, Kokubo T, et al. Age modifies the effect of 2-MeV fast neutrons on rat mammary carcinogenesis. Radiat Res. 2017;188:419–25.PubMedCrossRef
33.
Asatsuma-Okumura T, Ando H, De Simone M, Yamamoto J, Sato T, Shimizu N, et al. p63 is a cereblon substrate involved in thalidomide teratogenicity. Nat Chem Biol. 2019;15:1077–84.PubMedCrossRef
34.
Leonard MK, Kommagani R, Payal V, Mayo LD, Shamma HN, Kadakia MP. ∆Np63α regulates keratinocyte proliferation by controlling PTEN expression and localization. Cell Death Differ. 2011;18:1924–33.PubMedPubMedCentralCrossRef
35.
Linnemann JR, Miura H, Meixner LK, Irmler M, Kloos UJ, Hirschi B, et al. Quantification of regenerative potential in primary human mammary epithelial cells. Development. 2015;142:3239–51.PubMedPubMedCentral
36.
Latina A, Viticchiè G, Lena AM, Piro MC, Annicchiarico-Petruzzelli M, Melino G, et al. ∆Np63 targets cytoglobin to inhibit oxidative stress-induced apoptosis in keratinocytes and lung cancer. Oncogene. 2016;35:1493–503.PubMedCrossRef
37.
Yan W, Chen X. GPX2, a direct target of p63, inhibits oxidative stress-induced apoptosis in a p53-dependent manner. J Biol Chem. 2006;281:7856–62.PubMedCrossRef
38.
Oleksiewicz U, Liloglou T, Field JK, Xinarianos G. Cytoglobin: biochemical, functional and clinical perspective of the newest member of the globin family. Cell Mol Life Sci. 2011;68:3869–83.PubMedCrossRef
39.
Olive PL, Wlodek D, Banáth JP. DNA double-strand breaks measured in individual cells subjected to gel electrophoresis. Cancer Res. 1991;51:4671–6.PubMed
40.
Driessens N, Versteyhe S, Ghaddhab C, Burniat A, De Deken X, Van Sande J, et al. Hydrogen peroxide induces DNA single- and double-strand breaks in thyroid cells and is therefore a potential mutagen for this organ. Endocr Relat Cancer. 2009;16:845–56.PubMedCrossRef
41.
Rogakou EP, Pilch DR, Orr AH, Ivanova VS, Bonner WM. DNA double-stranded breaks induce histone H2AX phosphorylation on serine 139. J Biol Chem. 1998;273:5858–68.PubMedCrossRef
42.
Burma S, Chen BP, Murphy M, Kurimasa A, Chen DJ. ATM phosphorylates histone H2AX in response to DNA double-strand breaks. J Biol Chem. 2001;276:42462–7.PubMedCrossRef
43.
Lebel CP, Bondy SC. Sensitive and rapid quantitation of oxygen reactive species formation in rat synaptosomes. Neurochem Int. 1990;17:435–40.PubMedCrossRef
44.
Kawamura F, Inaki M, Katafuchi A, Abe Y, Tsuyama N, Kurosu Y, et al. Establishment of induced pluripotent stem cells from normal B cells and inducing AID expression in their differentiation into hematopoietic progenitor cells. Sci Rep. 2017;7:1659.PubMedPubMedCentralCrossRef
45.
Deckbar D, Jeggo PA, Löbrich M. Understanding the limitations of radiation-induced cell cycle checkpoints. Crit Rev Biochem Mol Biol. 2011;46:271–83.PubMedPubMedCentralCrossRef
46.
Landsverk KS, Patzke S, Rein ID, Stokke C, Lyng H, et al. Three independent mechanisms for arrest in G2 after ionizing radiation. Cell Cycle. 2011;10:1–11.CrossRef
47.
Nik-Zainal S, Davies H, Staaf J, Ramakrishna M, Glodzik D, Zou X, et al. Landscape of somatic mutations in 560 breast cancer whole-genome sequences. Nature. 2016;534:47–54.PubMedPubMedCentralCrossRef
Metadata
Title
ΔNp63α transcriptionally represses p53 target genes involved in the radiation-induced DNA damage response
ΔNp63α may cause genomic instability in epithelial stem cells
Authors
Ken-ichi Kudo
Naohiro Tsuyama
Kento Nagata
Tatsuhiko Imaoka
Daisuke Iizuka
Misaki Sugai-Takahashi
Moe Muramatsu
Akira Sakai
Publication date
01-12-2022
Publisher
BioMed Central
Published in
Radiation Oncology / Issue 1/2022
Electronic ISSN: 1748-717X
DOI
https://doi.org/10.1186/s13014-022-02139-7

Other articles of this Issue 1/2022

Radiation Oncology 1/2022 Go to the issue

Research

The prognostic significance of pretreatment squamous cell carcinoma antigen levels in cervical cancer patients treated by concurrent chemoradiation therapy and a comparison of dosimetric outcomes and clinical toxicities between tomotherapy and volumetric modulated arc therapy

Research

Early-onset adverse events after stereotactic radiosurgery for jugular foramen schwannoma: a mid-term follow-up single-center review of 46 cases

Research

The safety of an MRI simulation-guided boost after short-course preoperative radiotherapy for unresectable rectal cancer (SUNRISE): interim analysis of a randomized phase II trial

Research

18F-FET PET radiomics-based survival prediction in glioblastoma patients receiving radio(chemo)therapy

Short report

PET/CT-based adaptive radiotherapy of locally advanced non-small cell lung cancer in multicenter yDEGRO ARO 2017-01 cohort study

Research

Endoscopic resection as an independent predictive factor of local control in patients with T1bN0M0 esophageal squamous cell carcinoma treated with chemoradiotherapy: a retrospective study