Cyclin D1 overexpression perturbs DNA replication and induces replication-associated DNA double-strand breaks in acquired radioresistant cells

Abstract

Fractionated radiotherapy (RT) is widely used in cancer treatment, because it preserves normal tissues. However, repopulation of radioresistant tumors during fractionated RT limits the efficacy of RT. We recently demonstrated that a moderate level of long-term fractionated radiation confers acquired radioresistance to tumor cells, which is caused by DNA-PK/AKT/GSK3β-mediated cyclin D1 overexpression. The resulting cyclin D1 overexpression leads to forced progression of the cell cycle to S-phase, concomitant with induction of DNA double-strand breaks (DSBs). In this study, we investigated the molecular mechanisms underlying cyclin D1 overexpression-induced DSBs during DNA replication in acquired radioresistant cells. DNA fiber data demonstrated that replication forks progressed slowly in acquired radioresistant cells compared with corresponding parental cells in HepG2 and HeLa cell lines. Slowly progressing replication forks were also observed in HepG2 and HeLa cells that overexpressed a nondegradable cyclin D1 mutant. We also found that knockdown of Mus81 endonuclease, which is responsible for resolving aberrant replication forks, suppressed DSB formation in acquired radioresistant cells. Consequently, Mus81 created DSBs to remove aberrant replication forks in response to replication perturbation triggered by cyclin D1 overexpression. After treating cells with a specific inhibitor for DNA-PK or ATM, apoptosis rates increased in acquired radioresistant cells but not in parental cells by inhibiting the DNA damage response to cyclin D1-mediated DSBs. This suggested that these inhibitors might eradicate acquired radioresistant cells and improve fractionated RT outcomes.

Profiling of the Chromatin-associated Proteome Identifies HP1BP3 as a Novel Regulator of Cell Cycle Progression

“…3B-C), which is characteristic of global transcriptional activation (53). In addition, cycline D1 expression regulates G 1 -S phase progression (54), and expression levels of this protein are modulated by the chromatin-modeling factor SNF2H and the CCAAT-binding protein C/EBP (55). Our quantitative data FIG.…”

Nuclear accumulation of cyclin D1 following long-term fractionated exposures to low-dose ionizing radiation in normal human diploid cells

“…12 Cell cycledependent proteolysis of cyclin D1 is essential for S-phase progression, and cyclin D1 accumulation during S phase perturbs DNA replication. 15,20 These perturbations in DNA replication, e.g., DNA re-replication and the suppression of replication-fork progression, lead to DSBs. 15,20 Here we observed that normal human fibroblasts exposed to long-term FR accumulated nuclear cyclin D1 and suffered chromosome damage in the form of micronuclei (Fig.…”

“…12 AKT is transiently activated after SR exposure 13,14 but constitutively activated in cells exposed to FR for >14 d. 7 These results suggest that exposure to FR affects the AKT signaling pathway. 7,15 Constitutive AKT activation following FR exposures downregulates the nuclear export and proteolysis of cyclin D1, which results in the nuclear retention of cyclin D1 during S phase. 7 We recently reported that the abnormal expression of cyclin D1 during S phase damages DNA by blocking DNA replication.…”

“…7 We recently reported that the abnormal expression of cyclin D1 during S phase damages DNA by blocking DNA replication. 15,16 These cyclin D1-mediated DSBs activate DNA-activated protein Cyclin D1 is a mitogenic sensor that responds to growth signals from the extracellular environment and regulates the G 1 -to-S cell cycle transition. When cells are acutely irradiated with a single dose of 10 Gy, cyclin D1 is degraded, causing cell cycle arrest at the G 1 /S checkpoint.…”

X-Ray Induced Mutation Frequency at the <i>Hypoxanthine Phosphoribosyltransferase</i> Locus in Clinically Relevant Radioresistant Cells

“…The molecular basis of radioresistance remains to be elucidated. The radioresistant phenotype has been correlated with characteristics such as the enhanced repair of IR-induced DNA damage [1] [2] [3] [4], overexpression of cyclin D1 [5] [6], higher rates of cell growth and migration [7], a lower concentration of reactive oxygen species (ROS) in cells [8] [9] [10], inactivation of apoptotic proteins [11] and alterations in radiation-induced autophagy [12] [13]. A disadvantage of studying radioresistance is the lack of adequate radioresistant models reflecting the clinical features of this phenomenon.…”