Activation of the Pre-replication Complex is Blocked by Mimosine through Reactive Oxygen Species-Activated ATM without DNA Damage [Cell Biology]

January 13th, 2014 by Kubota, S., Fukumoto, Y., Ishibashi, K., Soeda, S., , Yuki, R., Nakayama, Y., Aoyama, K., Yamaguchi, N.,

Mimosine is an effective cell-synchronization reagent used for arresting cells in late G1 phase. However, the mechanism underlying mimosine-induced G1 cell cycle arrest remains unclear. Using highly synchronous cell populations, we show here that mimosine blocks S-phase entry through ATM activation. HeLa S3 cells are exposed to thymidine for 15 h, released for 9 h by washing out the thymidine, and subsequently treated with 1 mM mimosine for further 15 h (Thymidine‚ÜíMimosine). In contrast to thymidine-induced S-phase arrest, mimosine treatment synchronizes >90% of cells at the G1-S phase boundary by inhibiting the transition of the pre-replication complex to the preinitiation complex. Mimosine treatment activates ATM/ATR-mediated checkpoint signaling without inducing DNA damage. Inhibition of ATM activity is found to induce mimosine-arrested cells to enter S phase. In addition, ATM activation by mimosine treatment is mediated by reactive oxygen species (ROS). These results suggest that upon mimosine treatment, ATM blocks S-phase entry in response to ROS, which prevents replication fork stalling-induced DNA damage.
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