Loss of wild-type p53 function is responsible for upregulated homologous recombination in immortal rodent fibroblasts.
OBJECTIVE
A correlation between mutations in the tumour suppressor gene p53 and high rates of homologous recombination were previously found in immortal rodent fibroblasts. In the current study, direct evidence was sought that loss of p53 function is mainly responsible for upregulated levels of homologous recombination.


METHODS
Homologous recombination was assessed in vitro using DNA plasmid substrates that stably integrated into the genome of mouse and rat embryonic fibroblasts.


RESULTS
Primary fibroblasts with wild-type p53 displayed a recombination rate of about 1 x 10(-4). This number increased by 33- to 93-fold after spontaneous cellular immortalization, accompanied by loss of p53 function. To exclude potential bias from other gene mutations, wild-type p53 was experimentally disrupted in primary fibroblasts leading to an increase in recombination by one order of magnitude. Conversely, re-introduction of wild-type p53 into p53-null immortal cells reconstituted suppressed recombination rates. Finally, early-passage fibroblast cultures from p53-knock-out mice showed elevated recombination rates, which did not increase further following immortalization.


CONCLUSIONS
Loss of wild-type p53 is the major genetic determinant of increased homologous recombination frequencies in immortal rodent fibroblasts. Cellular p53 status will be an important factor to consider when performing functional analysis of the increasing number of mammalian proteins that are found to be involved in homologous recombination.
Willers
H
H
Department of Radiooncology, University of Hamburg, Germany.
McCarthy
E E
EE
Alberti
W
W
Dahm-Daphi
J
J
Powell
S N
SN
D000818:Animals; D002455:Cell Division / Q000235:genetics; D002471:Cell Transformation, Neoplastic / Q000235:genetics*; D002478:Cells, Cultured; D005347:Fibroblasts / Q000473:pathology* / Q000502:physiology*; D016158:Genes, p53; D051379:Mice; D009154:Mutation; D010957:Plasmids; D051381:Rats; D011995:Recombination, Genetic; D015854:Up-Regulation