The Mre11-CDK2 Interaction in the DNA Damage Response.

MRN (Mre11, Rad50, and NBS1) complex consists of highly conserved proteins integral to DNA double-strand break (DSB) signaling and repair. The unique MRN complex architecture allows direct binding to damaged DNA ends, DSB detection, and activation of cellular repair systems. Defects in the MRN complex lead to cancer predisposition, neurodegeneration, and immunodeficiency. MRN is required to initiate homologous recombination, the predominant repair pathway during the S and G2 phases of the cell cycle. Mre11 provides the nuclease activity needed for resection, while the NBS1 subunit interacts with ATM kinase, the master regulator of the DNA damage response (DDR) signaling cascade. The nuclease activity of Mre11 are required but alone are insufficient for resection; tumor suppressor BRCA1 and CtIP protein are also needed. CtIP is responsible for cell-cycle regulation of resection, while the major cyclin-dependent kinase (CDK) in S-phase is CDK2 bound to Cyclin A. Phosphorylation of CtIP by CDK2 allows the MRN-CtIP-BRCA1 resection complex to assemble, providing maximum resection capacity for homologous recombination. Mre11 controls these events through direct interaction with CDK2 which is required for CtIP phosphorylation and interaction with BRCA in normal cells. This observation demonstrates the important functions of MRN in both the DDR and in normal cell cycle regulation. The DDR has been extensively studied, but the role of CDK2 within the DDR remains unclear. The work presented here includes the following investigations: 1) the impact of DNA damage on the Mre11-CDK2/Cyclin A interaction, 2) whether change in the Mre11-CDK2/Cyclin A interaction is ATM-dependent, and 3) whether alterations in the activity of CDK2 indicate DDR participation. To examine the effect of DNA damage on the Mre11-CDK2/Cyclin A complex, I exposed mammalian cells to ionizing radiation and evaluated the interaction of endogenous Mre11 and CDK2/Cyclin A. These data indicate that DNA damage induces rapid dissociation of the Mre11-CDK2/Cyclin A complex dependent on ATM kinase activity and correlative to a reduction in CDK2 activity. This suggests that the Mre11-CDK2/Cyclin A complex dissociation causes a reduction in CDK2 activity, thereby contributing to the delay of cell cycle progression in S-phase, allowing for DNA repair proteins to restore DNA integrity.PhDToxicologyUniversity of Michigan, Horace H. Rackham School of Graduate Studieshttp://deepblue.lib.umich.edu/bitstream/2027.42/108856/1/toddafes_1.pd