Gene amplification in human cells knocked down for RAD54

Aurora Ruiz-Herrera, Alexandra Smirnova, Lela Khouriauli, Solomon G. Nergadze, Chiara Mondello, Elena Giulotto

Research output: Contribution to journalArticle

8 Citations (Scopus)

Abstract

Background: In mammalian cells gene amplification is a common manifestation of genome instability promoted by DNA double-strand breaks (DSBs). The repair of DSBs mainly occurs through two mechanisms: non-homologous end-joining (NHEJ) and homologous recombination (HR). We previously showed that defects in the repair of DSBs via NHEJ could increase the frequency of gene amplification. In this paper we explored whether a single or a combined defect in DSBs repair pathways can affect gene amplification.Results: We constructed human cell lines in which the expression of RAD54 and/or DNA-PKcs was constitutively knocked-down by RNA interference. We analyzed their radiosensitivity and their capacity to generate amplified DNA. Our results showed that both RAD54 and DNA-PKcs deficient cells are hypersensitive to γ-irradiation and generate methotrexate resistant colonies at a higher frequency compared to the proficient cell lines. In addition, the analysis of the cytogenetic organization of the amplicons revealed that isochromosome formation is a prevalent mechanism responsible for copy number increase in RAD54 defective cells.Conclusions: Defects in the DSBs repair mechanisms can influence the organization of amplified DNA. The high frequency of isochromosome formation in cells deficient for RAD54 suggests that homologous recombination proteins might play a role in preventing rearrangements at the centromeres.

Original languageEnglish
Article number5
JournalGenome Integrity
Volume2
DOIs
Publication statusPublished - Mar 18 2011

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Gene Amplification
Isochromosomes
Homologous Recombination
DNA
Cell Line
Double-Stranded DNA Breaks
Centromere
Genomic Instability
Cytogenetic Analysis
Radiation Tolerance
RNA Interference
Methotrexate
Proteins

ASJC Scopus subject areas

  • Genetics
  • Molecular Biology

Cite this

Ruiz-Herrera, A., Smirnova, A., Khouriauli, L., Nergadze, S. G., Mondello, C., & Giulotto, E. (2011). Gene amplification in human cells knocked down for RAD54. Genome Integrity, 2, [5]. https://doi.org/10.1186/2041-9414-2-5

Gene amplification in human cells knocked down for RAD54. / Ruiz-Herrera, Aurora; Smirnova, Alexandra; Khouriauli, Lela; Nergadze, Solomon G.; Mondello, Chiara; Giulotto, Elena.

In: Genome Integrity, Vol. 2, 5, 18.03.2011.

Research output: Contribution to journalArticle

Ruiz-Herrera, A, Smirnova, A, Khouriauli, L, Nergadze, SG, Mondello, C & Giulotto, E 2011, 'Gene amplification in human cells knocked down for RAD54', Genome Integrity, vol. 2, 5. https://doi.org/10.1186/2041-9414-2-5
Ruiz-Herrera A, Smirnova A, Khouriauli L, Nergadze SG, Mondello C, Giulotto E. Gene amplification in human cells knocked down for RAD54. Genome Integrity. 2011 Mar 18;2. 5. https://doi.org/10.1186/2041-9414-2-5
Ruiz-Herrera, Aurora ; Smirnova, Alexandra ; Khouriauli, Lela ; Nergadze, Solomon G. ; Mondello, Chiara ; Giulotto, Elena. / Gene amplification in human cells knocked down for RAD54. In: Genome Integrity. 2011 ; Vol. 2.
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