Depletion of Werner helicase results in mitotic hyperrecombination and pleiotropic homologous and nonhomologous recombination phenotypes.
Werner syndrome (WS) is a rare, segmental progeroid syndrome caused by defects in the WRN gene, which encodes a RecQ helicase. WRN has roles in many aspects of DNA metabolism including DNA repair and recombination. In this study, we exploited two different recombination assays previously used to describe a role for the structure-specific endonuclease ERCC1-XPF in mitotic and targeted homologous recombination. We constructed Chinese hamster ovary (CHO) cell lines isogenic with the cell lines used in these previous studies by depleting WRN using shRNA vectors. When intrachromosomal, mitotic recombination was assayed in WRN-depleted CHO cells, a hyperrecombination phenotype was observed, and a small number of aberrant recombinants were generated. Targeted homologous recombination was also examined in WRN-depleted CHO cells using a plasmid-chromosome targeting assay. In these experiments, loss of WRN resulted in a significant decrease in nonhomologous integration events and ablation of recombinants that required random integration of the corrected targeting vector. Aberrant recombinants were also recovered, but only from WRN-depleted cells. The pleiotropic recombination phenotypes conferred by WRN depletion, reflected in distinct homologous and nonhomologous recombination pathways, suggest a role for WRN in processing specific types of homologous recombination intermediates as well as an important function in nonhomologous recombination.
Rahn
Jennifer J
JJ
University of Texas M.D. Anderson Cancer Center, Department of Carcinogenesis, Science Park Research Division, P.O. Box 389, Smithville, TX 78597, United States.
Lowery
Megan P
MP
Della-Coletta
Luis
L
Adair
Gerald M
GM
Nairn
Rodney S
RS
D000818:Animals; D016466:CHO Cells; D002875:Chromosomes / Q000648:ultrastructure; D006224:Cricetinae; D003412:Cricetulus; D004268:DNA-Binding Proteins / Q000235:genetics; D004720:Endonucleases / Q000235:genetics; D005090:Exodeoxyribonucleases / Q000235:genetics* / Q000502:physiology*; D006801:Humans; D051379:Mice; D008938:Mitosis; D010641:Phenotype; D034741:RNA, Small Interfering / Q000378:metabolism; D053484:RecQ Helicases / Q000235:genetics* / Q000378:metabolism / Q000502:physiology*; D011995:Recombination, Genetic; D000071657:Werner Syndrome Helicase
D004268:DNA-Binding Proteins; D034741:RNA, Small Interfering; C071447:ERCC1 protein, human; D004720:Endonucleases; D005090:Exodeoxyribonucleases; D053484:RecQ Helicases; C098785:WRN protein, human; D000071657:Werner Syndrome Helicase; C499253:Wrn protein, mouse