Change search
CiteExportLink to record
Permanent link

Direct link
Cite
Citation style
  • apa
  • ieee
  • modern-language-association-8th-edition
  • vancouver
  • Other style
More styles
Language
  • de-DE
  • en-GB
  • en-US
  • fi-FI
  • nn-NO
  • nn-NB
  • sv-SE
  • Other locale
More languages
Output format
  • html
  • text
  • asciidoc
  • rtf
Removal of heat-sensitive clustered damaged DNA sites is independent of double-strand break repair
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Medical Radiation Science.
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Medical Radiation Science.
2018 (English)In: PLoS ONE, ISSN 1932-6203, E-ISSN 1932-6203, Vol. 13, no 12, article id e0209594Article in journal (Refereed) Published
Abstract [en]

DNA double-strand breaks (DSBs) are the most deleterious lesions that can arise in cells after ionizing radiation or radiometric drug treatment. In addition to prompt DSBs, DSBs may also be produced during repair, evolving from a clustered DNA damaged site, which is composed of two or more distinct lesions that are located within two helical turns. A specific type of cluster damage is the heat-sensitive clustered site (HSCS), which transforms into DSBs upon treatment at elevated temperatures. The actual lesions or mechanisms that mediate the HSCS transformation into DSBs are unknown. However, there are two possibilities; either these lesions are transformed into DSBs due to DNA lesion instability, e.g., transfer of HSCS into single-strand breaks (SSBs), or they are formed due to local DNA structure instability, e.g., DNA melting, where two SSBs on opposite strands meet and transform into a DSB. The importance of these processes in living cells is not understood, but they significantly affect estimates of DSB repair capacity. In this study, we show that HSCS removal in human cells is not affected by defects in DSB repair or inhibition of DSB repair. Under conditions where rejoining of prompt DSBs was almost completely inhibited, heat-sensitive DSBs were successfully rejoined, without resulting in increased DSB levels, indicating that HSCS do not transfer into DSB in cells under physiological conditions. Furthermore, analysis by atomic force microscopy suggests that prolonged heating of chromosomal DNA can induce structural changes that facilitate transformation of HSCS into DSB. In conclusion, the HSCS do not generate additional DSBs at physiological temperatures in human cells, and the repair of HSCS is independent of DSB repair.

Place, publisher, year, edition, pages
2018. Vol. 13, no 12, article id e0209594
National Category
Cancer and Oncology
Identifiers
URN: urn:nbn:se:uu:diva-374120DOI: 10.1371/journal.pone.0209594ISI: 000454621900032PubMedID: 30592737OAI: oai:DiVA.org:uu-374120DiVA, id: diva2:1281985
Funder
Swedish Cancer Society, CAN2014/661Swedish Cancer Society, CAN2016/649Swedish Radiation Safety Authority, SSM2017-2374Swedish Radiation Safety Authority, SSM2018-2181Available from: 2019-01-23 Created: 2019-01-23 Last updated: 2019-03-08Bibliographically approved
In thesis
1. Induction and repair of clustered DNA damage sites after exposure to ionizing radiation
Open this publication in new window or tab >>Induction and repair of clustered DNA damage sites after exposure to ionizing radiation
2019 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

The mechanisms that maintain genomic stability safeguard cells from constant DNA damage produced by endogenous and external stressors. Therefore, this thesis aimed to specifically address questions regarding the requirement and involvement of DNA repair proteins in the repair of various types of radiation-induced DNA damage.

The first aim was to determine whether the phosphorylation of DNA-PKcs, a major kinase involved in non-homologous end joining pathway, can be utilized to score the DNA double-strand break (DSB) content in cells. DNA-PKcs phosphorylated (pDNA-PKcs) at T2609 was more sensitive to the cellular DSB content than ɣH2AX, as analyzed by flow cytometry. Further, pDNA-PKcs at T2609 could discriminate between DSB repair-compromised and normal cells, confirming that the pDNA-PKcs can be used as a DSB repair marker. In paper II, the DSB repair was assessed in cells with reduced levels of DNA-PKcs. The reduction in DNA-PKcs resulted in decreased cell survival and unaffected DSB repair. These results clearly indicate that DNA-PKcs plays an additional role in promoting cell survival in addition to its function in DSB repair.

The second part of the thesis focused on the characterization of complex DNA damage. DNA damage was investigated after exposure to α-particles originating from Ra-223. The Ra-223 treatment induced a nonrandom DSB distribution consistent with damage induced by high-linear energy transfer radiation. The exposure to Ra-223 significantly reduced cell survival in monolayers and 3D cell structures. The last paper unraveled the fate of heat-sensitive clustered DNA damage site (HSCS) repair in cells. HSCS repair was independent of DSB repair, and these lesions did not contribute to the generation of additional DSBs during repair. Prolonged heating of DNA at relatively low temperatures induced structural changes in the DNA that contributed to the production of DNA artifacts.

In conclusion, these results demonstrate that DNA-PKcs can be used to monitor DSB repair in cells after exposure to ionizing radiation. However, the functions of DNA-PKcs are not limited to DSB repair, as it can promote cell survival through other mechanisms. The complexity of the DNA damage produced by high-LET radiation is a major contributor to cell death. However, not all clusters produced in irradiated cells are converted into DSBs during repair.

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2019. p. 54
Series
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Medicine, ISSN 1651-6206 ; 1548
Keywords
NHEJ, DSB repair, clustered DNA damage, DNA repair, DNA-PKcs, HSCS, Ra-223, ionizing radiation
National Category
Medical and Health Sciences
Research subject
Medical Science
Identifiers
urn:nbn:se:uu:diva-378721 (URN)978-91-513-0591-2 (ISBN)
Public defence
2019-04-29, Rudbecksalen, Rudbecklaboratoriet, Dag Hammarskjölds v 20, Uppsala, 13:00 (English)
Opponent
Supervisors
Available from: 2019-04-04 Created: 2019-03-08 Last updated: 2019-05-07

Open Access in DiVA

fulltext(2086 kB)41 downloads
File information
File name FULLTEXT01.pdfFile size 2086 kBChecksum SHA-512
1b0842c5c789ce7dd1fea8791af69066afa2216580de658640c2640cc176061be74c1bf6288de555e98afc15f4f0919372fe0336d87562528e52c7a034590c20
Type fulltextMimetype application/pdf

Other links

Publisher's full textPubMed

Search in DiVA

By author/editor
Abramenkovs, AndrisStenerlöw, Bo
By organisation
Medical Radiation Science
In the same journal
PLoS ONE
Cancer and Oncology

Search outside of DiVA

GoogleGoogle Scholar
Total: 41 downloads
The number of downloads is the sum of all downloads of full texts. It may include eg previous versions that are now no longer available

doi
pubmed
urn-nbn

Altmetric score

doi
pubmed
urn-nbn
Total: 451 hits
CiteExportLink to record
Permanent link

Direct link
Cite
Citation style
  • apa
  • ieee
  • modern-language-association-8th-edition
  • vancouver
  • Other style
More styles
Language
  • de-DE
  • en-GB
  • en-US
  • fi-FI
  • nn-NO
  • nn-NB
  • sv-SE
  • Other locale
More languages
Output format
  • html
  • text
  • asciidoc
  • rtf