DNA damage responses in human induced pluripotent stem cells and embryonic stem cells.
BACKGROUND
Induced pluripotent stem (iPS) cells have the capability to undergo self-renewal and differentiation into all somatic cell types. Since they can be produced through somatic cell reprogramming, which uses a defined set of transcription factors, iPS cells represent important sources of patient-specific cells for clinical applications. However, before these cells can be used in therapeutic designs, it is essential to understand their genetic stability.


RESULTS
Here, we describe DNA damage responses in human iPS cells. We observe hypersensitivity to DNA damaging agents resulting in rapid induction of apoptosis after γ-irradiation. Expression of pluripotency factors does not appear to be diminished after irradiation in iPS cells. Following irradiation, iPS cells activate checkpoint signaling, evidenced by phosphorylation of ATM, NBS1, CHEK2, and TP53, localization of ATM to the double strand breaks (DSB), and localization of TP53 to the nucleus of NANOG-positive cells. We demonstrate that iPS cells temporary arrest cell cycle progression in the G(2) phase of the cell cycle, displaying a lack of the G(1)/S cell cycle arrest similar to human embryonic stem (ES) cells. Furthermore, both cell types remove DSB within six hours of γ-irradiation, form RAD51 foci and exhibit sister chromatid exchanges suggesting homologous recombination repair. Finally, we report elevated expression of genes involved in DNA damage signaling, checkpoint function, and repair of various types of DNA lesions in ES and iPS cells relative to their differentiated counterparts.


CONCLUSIONS
High degrees of similarity in DNA damage responses between ES and iPS cells were found. Even though reprogramming did not alter checkpoint signaling following DNA damage, dramatic changes in cell cycle structure, including a high percentage of cells in the S phase, increased radiosensitivity and loss of DNA damage-induced G(1)/S cell cycle arrest, were observed in stem cells generated by induced pluripotency.
Momcilovic
Olga
O
Department of Human Genetics, Graduate School of Public Health, University of Pittsburgh, Pittsburgh, Pennsylvania, United States of America.
Knobloch
Leah
L
Fornsaglio
Jamie
J
Varum
Sandra
S
Easley
Charles
C
Schatten
Gerald
G
D015153:Blotting, Western; D002453:Cell Cycle; D018797:Cell Cycle Proteins / Q000378:metabolism; D002454:Cell Differentiation; D002460:Cell Line; D004249:DNA Damage; D004260:DNA Repair; D053595:Embryonic Stem Cells / Q000166:cytology* / Q000378:metabolism; D005434:Flow Cytometry; D020869:Gene Expression Profiling; D006801:Humans; D007150:Immunohistochemistry; D018613:Microscopy, Confocal; D010766:Phosphorylation; D039904:Pluripotent Stem Cells / Q000166:cytology* / Q000378:metabolism; D016133:Polymerase Chain Reaction
D018797:Cell Cycle Proteins