The Mechanisms and Consequences of DNA Re-replication

Abstract

DNA replication is a key event during cellular proliferation. In order to maintain genetic stability, cells have evolved different mechanisms to ensure the precise duplication of chromosomes and prevent DNA re-replication. The regulation of DNA replication initiation is critical for preventing re-replication. In mammalian cells, stabilization and activation of the replication initiator Cdt1 leads to DNA re-replication.In this dissertation, I examine the consequences of the re-replication.The single stranded DNA (ssDNA) initially generated during re-replication activates an ATR/Chk1 mediated pathway and arrests cells in G2/M phase. The checkpoint is essential for the accumulation of re-replicated cells, which can further activate ATM/Chk2, p53, and apoptosis. Our study suggests that cells can have a chance to repair relatively minor DNA damage caused by microscopic re-replication, and only induce apoptosis through the later acquisition of double strand breaks and activation of Chk2 and p53 when re-replication persists. We also identified HDAC6 deacetylase as a new player in re-replication induced checkpoint pathways, presumably by regulating Chk1 protein level and phosphorylation, directly or indirectly.MLN4924, a new anti-cancer drug, stabilizes Cdt1 and causes re-replication in a variety of human cancer cells. Transient exposure of MLN4924 is sufficient to induce rereplication, which activates checkpoint pathways, apoptosis, and senescence, contributing to the anti-proliferative effect of MLN4924 in cancer therapy. Intriguingly, unlike other III DNA damaging agents used for chemotherapy, p53-negative cells remain susceptible to MLN4924 induced cell death, suggesting an important clinical application.