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PhiC31 integrase induces a DNA damage response and chromosomal rearrangements in human adult fibroblasts

Liu, Jian LU ; Skjorringe, Tina; Gjetting, Torben and Jensen, Thomas G. (2009) In BMC Biotechnology 9.
Abstract
Background: PhiC31 integrase facilitates efficient integration of transgenes into human and mouse genomes and is considered for clinical gene therapy. However recent studies have shown that the enzyme can induce various chromosomal abnormalities in primary human embryonic cells and mammalian cell lines. The mechanisms involved are unknown, but it has been proposed that PhiC31 attachment sites in the host genome recombine leading to chromosomal translocations. Results: We have studied possible effects of the PhiC31 integrase expression in human adult fibroblasts by karyotyping. All control cells were cytogenetically normal, whereas cells expressing PhiC31 integrase show chromosomal abnormalities confirming our previous results using primary... (More)
Background: PhiC31 integrase facilitates efficient integration of transgenes into human and mouse genomes and is considered for clinical gene therapy. However recent studies have shown that the enzyme can induce various chromosomal abnormalities in primary human embryonic cells and mammalian cell lines. The mechanisms involved are unknown, but it has been proposed that PhiC31 attachment sites in the host genome recombine leading to chromosomal translocations. Results: We have studied possible effects of the PhiC31 integrase expression in human adult fibroblasts by karyotyping. All control cells were cytogenetically normal, whereas cells expressing PhiC31 integrase show chromosomal abnormalities confirming our previous results using primary embryonic fibroblasts. In order to study the early mechanisms involved we measured H2AX phosphorylation-a primary event in the response to DNA double-strand-breaks. Transient transfection with PhiC31 integrase encoding plasmids lead to an elevated number of cells positive for H2AX phosphorylation detected by immunofluorescence. Western blot analysis confirmed the upregulated H2AX phosphorylation, whereas markers for apoptosis as well as p53 and p21 were not induced. Cells transfected with plasmids encoding the Sleeping Beauty transposase remained cytogenetically normal, and in these cells less upregulation of H2AX phosphorylation could be detected. Conclusion: In primary human fibroblasts expression of PhiC31 integrase leads to a DNA damage response and chromosomal aberrations. (Less)
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author
organization
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type
Contribution to journal
publication status
published
subject
in
BMC Biotechnology
volume
9
publisher
BioMed Central
external identifiers
  • wos:000266607100002
  • scopus:65849090971
ISSN
1472-6750
DOI
10.1186/1472-6750-9-31
language
English
LU publication?
yes
id
9eba8e46-7fcd-4f41-b279-8a0fced41ba5 (old id 1444091)
date added to LUP
2009-07-17 09:42:03
date last changed
2017-01-01 05:27:56
@article{9eba8e46-7fcd-4f41-b279-8a0fced41ba5,
  abstract     = {Background: PhiC31 integrase facilitates efficient integration of transgenes into human and mouse genomes and is considered for clinical gene therapy. However recent studies have shown that the enzyme can induce various chromosomal abnormalities in primary human embryonic cells and mammalian cell lines. The mechanisms involved are unknown, but it has been proposed that PhiC31 attachment sites in the host genome recombine leading to chromosomal translocations. Results: We have studied possible effects of the PhiC31 integrase expression in human adult fibroblasts by karyotyping. All control cells were cytogenetically normal, whereas cells expressing PhiC31 integrase show chromosomal abnormalities confirming our previous results using primary embryonic fibroblasts. In order to study the early mechanisms involved we measured H2AX phosphorylation-a primary event in the response to DNA double-strand-breaks. Transient transfection with PhiC31 integrase encoding plasmids lead to an elevated number of cells positive for H2AX phosphorylation detected by immunofluorescence. Western blot analysis confirmed the upregulated H2AX phosphorylation, whereas markers for apoptosis as well as p53 and p21 were not induced. Cells transfected with plasmids encoding the Sleeping Beauty transposase remained cytogenetically normal, and in these cells less upregulation of H2AX phosphorylation could be detected. Conclusion: In primary human fibroblasts expression of PhiC31 integrase leads to a DNA damage response and chromosomal aberrations.},
  articleno    = {31},
  author       = {Liu, Jian and Skjorringe, Tina and Gjetting, Torben and Jensen, Thomas G.},
  issn         = {1472-6750},
  language     = {eng},
  publisher    = {BioMed Central},
  series       = {BMC Biotechnology},
  title        = {PhiC31 integrase induces a DNA damage response and chromosomal rearrangements in human adult fibroblasts},
  url          = {http://dx.doi.org/10.1186/1472-6750-9-31},
  volume       = {9},
  year         = {2009},
}