Influence of homologous recombinational repair on cell survival and chromosomal aberration induction during the cell cycle in γ-irradiated CHO cells

Paul F. Wilson, John M. Hinz, Salustra S. Urbin, Peter B. Nham, Larry H. Thompson

Research output: Contribution to journalArticle

12 Citations (Scopus)

Abstract

The repair of DNA double-strand breaks (DSBs) by homologous recombinational repair (HRR) underlies the high radioresistance and low mutability observed in S-phase mammalian cells. To evaluate the contributions of HRR and non-homologous end-joining (NHEJ) to overall DSB repair capacity throughout the cell cycle after γ-irradiation, we compared HRR-deficient RAD51D-knockout 51D1 to CgRAD51D-complemented 51D1 (51D1.3) CHO cells for survival and chromosomal aberrations (CAs). Asynchronous cultures were irradiated with 150 or 300 cGy and separated by cell size using centrifugal elutriation. Cell survival of each synchronous fraction (∼20 fractions total from early G1 to late G2/M) was measured by colony formation. 51D1.3 cells were most resistant in S, while 51D1 cells were most resistant in early G1 (with survival and chromosome-type CA levels similar to 51D1.3) and became progressively more sensitive throughout S and G2. Both cell lines experienced significantly reduced survival from late S into G2. Metaphases were collected from every third elutriation fraction at the first post-irradiation mitosis and scored for CAs. 51D1 cells irradiated in S and G2 had ∼2-fold higher chromatid-type CAs and a remarkable ∼25-fold higher level of complex chromatid-type exchanges compared to 51D1.3 cells. Complex exchanges in 51D1.3 cells were only observed in G2. These results show an essential role for HRR in preventing gross chromosomal rearrangements in proliferating cells and, with our previous report of reduced survival of G2-phase NHEJ-deficient prkdc CHO cells [Hinz et al., DNA Repair 4, 782-792, 2005], imply reduced activity/efficiency of both HRR and NHEJ as cells transition from S to G2.

Original languageEnglish (US)
Pages (from-to)737-744
Number of pages8
JournalDNA Repair
Volume9
Issue number7
DOIs
StatePublished - Jul 1 2010
Externally publishedYes

Fingerprint

Recombinational DNA Repair
CHO Cells
Aberrations
Chromosome Aberrations
Cell Survival
Cell Cycle
Repair
Cells
Joining
Chromatids
Irradiation
Double-Stranded DNA Breaks
G2 Phase
DNA
Metaphase
Chromosomes
S Phase
Cell Size
Mitosis
DNA Repair

Keywords

  • Cell cycle
  • Chromosomal aberrations
  • Complex exchange
  • Homologous recombinational repair
  • Non-homologous end-joining
  • Radiosensitivity

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

Cite this

Influence of homologous recombinational repair on cell survival and chromosomal aberration induction during the cell cycle in γ-irradiated CHO cells. / Wilson, Paul F.; Hinz, John M.; Urbin, Salustra S.; Nham, Peter B.; Thompson, Larry H.

In: DNA Repair, Vol. 9, No. 7, 01.07.2010, p. 737-744.

Research output: Contribution to journalArticle

Wilson, Paul F. ; Hinz, John M. ; Urbin, Salustra S. ; Nham, Peter B. ; Thompson, Larry H. / Influence of homologous recombinational repair on cell survival and chromosomal aberration induction during the cell cycle in γ-irradiated CHO cells. In: DNA Repair. 2010 ; Vol. 9, No. 7. pp. 737-744.
@article{f00c0d7a2275450cbb2076389999481c,
title = "Influence of homologous recombinational repair on cell survival and chromosomal aberration induction during the cell cycle in γ-irradiated CHO cells",
abstract = "The repair of DNA double-strand breaks (DSBs) by homologous recombinational repair (HRR) underlies the high radioresistance and low mutability observed in S-phase mammalian cells. To evaluate the contributions of HRR and non-homologous end-joining (NHEJ) to overall DSB repair capacity throughout the cell cycle after γ-irradiation, we compared HRR-deficient RAD51D-knockout 51D1 to CgRAD51D-complemented 51D1 (51D1.3) CHO cells for survival and chromosomal aberrations (CAs). Asynchronous cultures were irradiated with 150 or 300 cGy and separated by cell size using centrifugal elutriation. Cell survival of each synchronous fraction (∼20 fractions total from early G1 to late G2/M) was measured by colony formation. 51D1.3 cells were most resistant in S, while 51D1 cells were most resistant in early G1 (with survival and chromosome-type CA levels similar to 51D1.3) and became progressively more sensitive throughout S and G2. Both cell lines experienced significantly reduced survival from late S into G2. Metaphases were collected from every third elutriation fraction at the first post-irradiation mitosis and scored for CAs. 51D1 cells irradiated in S and G2 had ∼2-fold higher chromatid-type CAs and a remarkable ∼25-fold higher level of complex chromatid-type exchanges compared to 51D1.3 cells. Complex exchanges in 51D1.3 cells were only observed in G2. These results show an essential role for HRR in preventing gross chromosomal rearrangements in proliferating cells and, with our previous report of reduced survival of G2-phase NHEJ-deficient prkdc CHO cells [Hinz et al., DNA Repair 4, 782-792, 2005], imply reduced activity/efficiency of both HRR and NHEJ as cells transition from S to G2.",
keywords = "Cell cycle, Chromosomal aberrations, Complex exchange, Homologous recombinational repair, Non-homologous end-joining, Radiosensitivity",
author = "Wilson, {Paul F.} and Hinz, {John M.} and Urbin, {Salustra S.} and Nham, {Peter B.} and Thompson, {Larry H.}",
year = "2010",
month = "7",
day = "1",
doi = "10.1016/j.dnarep.2010.03.009",
language = "English (US)",
volume = "9",
pages = "737--744",
journal = "DNA Repair",
issn = "1568-7864",
publisher = "Elsevier",
number = "7",

}

TY - JOUR

T1 - Influence of homologous recombinational repair on cell survival and chromosomal aberration induction during the cell cycle in γ-irradiated CHO cells

AU - Wilson, Paul F.

AU - Hinz, John M.

AU - Urbin, Salustra S.

AU - Nham, Peter B.

AU - Thompson, Larry H.

PY - 2010/7/1

Y1 - 2010/7/1

N2 - The repair of DNA double-strand breaks (DSBs) by homologous recombinational repair (HRR) underlies the high radioresistance and low mutability observed in S-phase mammalian cells. To evaluate the contributions of HRR and non-homologous end-joining (NHEJ) to overall DSB repair capacity throughout the cell cycle after γ-irradiation, we compared HRR-deficient RAD51D-knockout 51D1 to CgRAD51D-complemented 51D1 (51D1.3) CHO cells for survival and chromosomal aberrations (CAs). Asynchronous cultures were irradiated with 150 or 300 cGy and separated by cell size using centrifugal elutriation. Cell survival of each synchronous fraction (∼20 fractions total from early G1 to late G2/M) was measured by colony formation. 51D1.3 cells were most resistant in S, while 51D1 cells were most resistant in early G1 (with survival and chromosome-type CA levels similar to 51D1.3) and became progressively more sensitive throughout S and G2. Both cell lines experienced significantly reduced survival from late S into G2. Metaphases were collected from every third elutriation fraction at the first post-irradiation mitosis and scored for CAs. 51D1 cells irradiated in S and G2 had ∼2-fold higher chromatid-type CAs and a remarkable ∼25-fold higher level of complex chromatid-type exchanges compared to 51D1.3 cells. Complex exchanges in 51D1.3 cells were only observed in G2. These results show an essential role for HRR in preventing gross chromosomal rearrangements in proliferating cells and, with our previous report of reduced survival of G2-phase NHEJ-deficient prkdc CHO cells [Hinz et al., DNA Repair 4, 782-792, 2005], imply reduced activity/efficiency of both HRR and NHEJ as cells transition from S to G2.

AB - The repair of DNA double-strand breaks (DSBs) by homologous recombinational repair (HRR) underlies the high radioresistance and low mutability observed in S-phase mammalian cells. To evaluate the contributions of HRR and non-homologous end-joining (NHEJ) to overall DSB repair capacity throughout the cell cycle after γ-irradiation, we compared HRR-deficient RAD51D-knockout 51D1 to CgRAD51D-complemented 51D1 (51D1.3) CHO cells for survival and chromosomal aberrations (CAs). Asynchronous cultures were irradiated with 150 or 300 cGy and separated by cell size using centrifugal elutriation. Cell survival of each synchronous fraction (∼20 fractions total from early G1 to late G2/M) was measured by colony formation. 51D1.3 cells were most resistant in S, while 51D1 cells were most resistant in early G1 (with survival and chromosome-type CA levels similar to 51D1.3) and became progressively more sensitive throughout S and G2. Both cell lines experienced significantly reduced survival from late S into G2. Metaphases were collected from every third elutriation fraction at the first post-irradiation mitosis and scored for CAs. 51D1 cells irradiated in S and G2 had ∼2-fold higher chromatid-type CAs and a remarkable ∼25-fold higher level of complex chromatid-type exchanges compared to 51D1.3 cells. Complex exchanges in 51D1.3 cells were only observed in G2. These results show an essential role for HRR in preventing gross chromosomal rearrangements in proliferating cells and, with our previous report of reduced survival of G2-phase NHEJ-deficient prkdc CHO cells [Hinz et al., DNA Repair 4, 782-792, 2005], imply reduced activity/efficiency of both HRR and NHEJ as cells transition from S to G2.

KW - Cell cycle

KW - Chromosomal aberrations

KW - Complex exchange

KW - Homologous recombinational repair

KW - Non-homologous end-joining

KW - Radiosensitivity

UR - http://www.scopus.com/inward/record.url?scp=77953322110&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=77953322110&partnerID=8YFLogxK

U2 - 10.1016/j.dnarep.2010.03.009

DO - 10.1016/j.dnarep.2010.03.009

M3 - Article

C2 - 20434408

AN - SCOPUS:77953322110

VL - 9

SP - 737

EP - 744

JO - DNA Repair

JF - DNA Repair

SN - 1568-7864

IS - 7

ER -