Analysis of the DNA Damage Response in Mitotic Cells Following Laser Micro-Irradiation

DNA damage can occur naturally through endogenous metabolic by products, DNA replication errors and from environmental factors such as the sun’s harmful UV rays. Unrepaired DNA damage can lead to cell senescence, apoptosis or even up-regulated cell divisions resulting in cancer. Therefore, organisms have evolved the ability to recognize and repair damaged DNA. The field of DNA repair has demonstrated a growing interest in utilizing lasers to induce DNA damage in a specified submicron linear region of interphase nuclei.  However, few studies have looked at the DNA damage response in mitotic cells. The few published studies that do exist with regards to mitotic DNA damage have used radiomimetic drugs and ionizing radiation. The results from those studies have only showed recruitment of initial response proteins. We show that mitotic rat kangaroo (Ptk2) and human (U2OS) cells, damaged by the femtosecond near-IR laser lead to the formation of double strand breaks, yH2AX, which are capable of recruiting a variety of factors involved in the damage response and repair pathways.  Additionally, our results show that cells damaged with the laser are capable of initiating repair in mitotic chromatin. In conclusion, we demonstrate for the first time, that laser induced mitotic DNA damage not only recruits initial responding factors, but also initiates the actual repair proces