Abstract P1-06-05: Mre11 mediates a p53-independent quiescence program in response to mammary oncogene activation
Authors: Kumar RJ , Xiao HC , Fagan-Solis K , Purvis J , Gupta GP 
Published: May 2019

Abstract
Background: Oncogenic mutations drive uncontrolled proliferation, imposing significant replication stress and activating the DNA damage response (DDR) in early tumorigenesis. The accumulation of double strand breaks (DSBs) is recognized by the Mre11-Nbs-Rad50 (MRN) Complex, a critical sensor that mediates damage signaling and cell cycle checkpoint responses. The mechanism by which the DDR acts as an anti-tumor barrier is in part due to activation of p53 tumor suppressor. However, the role of the DDR in a p53 null or deficient setting has been poorly investigated. In basal-like breast cancer (BLBC) where p53 loss is ubiquitous, the functional consequence of the DDR is in need of further delineation. Previously, we have observed that Mre11 suppresses proliferation in hyperplastic mammary epithelium through a p53-independent mechanism. Here, we evaluate how Mre11 affects cell cycle regulation in p53 deficient primary mammary epithelium induced to overexpress the oncogene c-Myc, a known driver of BLBC.
Methods: We performed single-cell live imaging on primary murine mammary epithelial cells (mMECs) induced to overexpress c-Myc and be p53 deficient, with or without additional mutations in Mre11. Cells were further infected with a fluorescent cell cycle reporter: Proliferating Cell Nuclear Antigen (PCNA)-mCherry. By tracking PCNA foci changes via time-lapse imaging modalities, we were able to calculate distinct cell cycle phase lengths, identify disruption of cell cycle, visualize mitotic aberrancies, and characterize the cell cycle profile of p53 deficient, c-Myc activated primary mammary epithelial cells with or without intact DDR.
Results: Our results indicate that the presence of Mre11 mediates a quiescence phenotype even in the absence of p53 tumor suppressor with up to 75% of the cell fraction becoming quiescent. This fraction is significantly reduced (p&lt;0.0001) when Mre11 is targeted using gene editing giving rise to highly aberrant multinucleated mMECs capable of unrestrained replication and entry into mitosis. Cells with intact Mre11 undergo cell cycle arrest post abnormal mitoses and are characterized by quiescent cells with classic micronuclei formation. In contrast, mMECs without Mre11 are likely to be multinucleated with reduced micronuclei formation and rarely undergo cell cycle arrest. The loss of Mre11 enables these cells to continue cell cycle progression despite tremendous mitotic aberrancies. Impact: Mre11 is an important mediator of a p53 independent quiescence program in c-Myc expressing cells. It regulates the proliferative activity of multinucleated daughter cells and their ability to become quiescent. Furthering our understanding of how Mre11 regulates this molecular quiescence program may provide new insights into drivers of genomic instability in BLBC and novel therapeutic targets.
Citation Format: Kumar RJ, Xiao HC, Fagan-Solis K, Purvis J, Gupta GP. Mre11 mediates a p53-independent quiescence program in response to mammary oncogene activation [abstract]. In: Proceedings of the 2018 San Antonio Breast Cancer Symposium; 2018 Dec 4-8; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2019;79(4 Suppl):Abstract nr P1-06-05.