Research Papers: General

Targeting telomere-containing chromosome ends with a near-infrared femtosecond laser to study the activation of the DNA damage response and DNA damage repair pathways

[+] Author Affiliations
Bárbara Alcaraz Silva

Beckman Laser Institute and Medical Clinic, 1002 Health Sciences Road East, Irvine, California 92612

University of California, School of Biological Sciences, Department of Developmental and Cell Biology, Irvine, California 92617

Jessica R. Stambaugh

Beckman Laser Institute and Medical Clinic, 1002 Health Sciences Road East, Irvine, California 92612

Michael W. Berns

Beckman Laser Institute and Medical Clinic, 1002 Health Sciences Road East, Irvine, California 92612

University of California, School of Biological Sciences, Department of Developmental and Cell Biology, Irvine, California 92617

University of California, Department of Biomedical Engineering, Irvine, California 92617

University of California, Department of Surgery, Irvine, California 92617

J. Biomed. Opt. 18(9), 095003 (Sep 24, 2013). doi:10.1117/1.JBO.18.9.095003
History: Received July 16, 2013; Revised August 27, 2013; Accepted August 29, 2013
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Abstract.  Telomeres are at the ends of chromosomes. Previous evidence suggests that laser-induced deoxyribose nucleic acid (DNA) breaks at chromosome ends during anaphase results in delayed cytokinesis. A possible explanation for this delay is that the DNA damage response (DDR) mechanism has been activated. We describe a live imaging method to study the effects of DDR activation following focal point near-infrared femtosecond laser microirradiation either at a single chromosome end or at a chromosome arm in mitotic anaphase cells. Laser microirradiation is used in combination with dual fluorescent labeling to monitor the co-localization of double-strand break marker γH2AX along with the DDR factors in PtK2 (Potorous tridactylus) cells. Laser-induced DNA breaks in chromosome ends as well as in chromosome arms results in recruitment of the following: poly(ADP-ribose) polymerase 1, checkpoint sensors (p-Chk1, p-Chk2), DNA repair protein Ku70/Ku80, and proliferating cell nuclear antigen. However, phosphorylated p53 at serine 15 is detected only at chromosome ends and not at chromosome arms. Full activation of DDR on damaged chromosome ends may explain previously published results that showed the delay of cytokinesis.

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© 2013 Society of Photo-Optical Instrumentation Engineers

Citation

Bárbara Alcaraz Silva ; Jessica R. Stambaugh and Michael W. Berns
"Targeting telomere-containing chromosome ends with a near-infrared femtosecond laser to study the activation of the DNA damage response and DNA damage repair pathways", J. Biomed. Opt. 18(9), 095003 (Sep 24, 2013). ; http://dx.doi.org/10.1117/1.JBO.18.9.095003


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