Abstract
In response to DNA damage, the cellular decision of life versus death involves an intricate network of multiple factors that play critical roles in regulation of DNA repair, cell cycle, and cell death. DNA damage checkpoint proteins are crucial for maintaining DNA integrity and normal cellular functions, but they may also reduce the effectiveness of cancer treatment. Here we report the involvement of Cdk5 activator p35-binding protein C53 in regulation of apoptosis induced by genotoxic stress through modulating Cdk1-cyclin B1 function. C53 was originally identified as a Cdk5 activator p35-binding protein and a caspase substrate. Importantly, our results demonstrated that C53 deficiency conferred partial resistance to genotoxic agents such as etoposide and x-ray irradiation, whereas ectopic expression of C53 rendered cells susceptible to multiple genotoxins that usually trigger G(2)/M arrest. Furthermore, we found that Cdk1 activity was required for etoposide-induced apoptosis of HeLa cells. Overexpression of C53 promoted Cdk1 activity and nuclear accumulation of cyclin B1, whereas C53 deficiency led to more cytoplasmic retention of cyclin B1, suggesting that C53 acts as a pivotal player in modulating the G(2)/M DNA damage checkpoint. Finally, C53 and cyclin B1 co-localize and associate in vivo, indicating a direct role of C53 in regulating the Cdk1-cyclin B1 complex. Taken together, our results strongly indicate that in response to genotoxic stress, C53 serves as an important regulatory component of the G(2)/M DNA damage checkpoint. By overriding the G(2)/M checkpoint-mediated inhibition of Cdk1-cyclin B1 function, ectopic expression of C53 may represent a novel approach for chemo- and radio-sensitization of cancer cells.
Highlights
In response to DNA damage, the cellular decision of life versus death involves an intricate network of multiple factors that play critical roles in regulation of DNA repair, cell cycle, and cell death
We report the involvement of Cdk5 activator-binding protein C53 in regulation of apoptosis induced by genotoxic stress via modulating the G2/M DNA damage checkpoint
Our data demonstrated that C53 deficiency conferred partial resistance to genotoxic agents such as etoposide and x-ray irradiation, whereas ectopic expression of C53 rendered cells susceptible to multiple genotoxins
Summary
In response to DNA damage, the cellular decision of life versus death involves an intricate network of multiple factors that play critical roles in regulation of DNA repair, cell cycle, and cell death. We report the involvement of Cdk activator p35-binding protein C53 in regulation of apoptosis induced by genotoxic stress through modulating Cdk1-cyclin B1 function. Stress, DNA damage checkpoints play crucial roles in regulation of cell cycle arrest and cell death. In response to DNA damage, these checkpoints inhibit cell cycle progression from G1 to S (the G1/S checkpoint), DNA replication (the intra-S checkpoint), or G2 to mitosis (the G2/M checkpoint), respectively [2]. In response to DNA damage, the G2/M checkpoint system functions to arrest cell cycle in part by inhibiting Cdk1-cyclin B1 activity. Phosphorylated CDC25C is sequestered by 14-3-3 away from Cdk, thereby preventing dephosphorylation and activation of Cdk1 [8, 9] Another mechanism of the G2/M checkpoint control is nuclear exclusion of cyclin B1 [10, 11]. At the onset of mitosis, phosphorylation of four serine residues (Ser-126, -128, -133, and -147) at the N-terminal region of human cyclin B1 that contains both cytoplasmic retention sequence and nuclear export sequence causes an inhibition of nuclear export and enhances nuclear import, resulting in a net accumulation of cyclin B in nucleus
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