Abstract A20: Excess centrosomes induce p53-dependent senescence in endothelial cells

Abstract

Abstract Excess centrosomes (>2 centrosomes/cell), which are prevalent in both tumor cells and tumor endothelial cells, lead to aneuploidy and chromosome instability. Therefore, understanding how excess centrosomes affect cell behaviors is critical for studying tumor progression and tumor angiogenesis. As demonstrated by previous literature, cells with excess centrosomes routinely undergo bipolar cell division to produce viable progeny. However, we found that endothelial cells were not able to maintain the percentage of cells with excess centrosomes, suggesting that excess centrosomes had a negative impact on endothelial cell cycle. We used a tetracycline-inducible Polo-like kinase 4 (Plk4)-expressing system to induce excess centrosomes in human umbilical vein endothelial cells (HUVEC). Doxycycline treatment induced ~30% centrosome over-duplication, which decreased to <5% in 6 days. We found that HUVEC with excess centrosomes activated p53 by phosphorylating p53 at Ser33. Complete down-regulation of p53 helped maintain the frequency of cells with excess centrosomes. Excess centrosomes-induced p53 activation was independent of DNA damage, as demonstrated by lack of p53 Ser15 phosphorylation or γH2AX foci. Excess centrosomes induced senescence but not apoptosis, as shown by senescence-associated beta-galactosidase (SA-β-gal) activity and Galactosidase Beta 1 (GLB1) antibody staining, and the induced senescence can be blocked by p53 down-regulation. Since endothelial cells maintain different cell shapes and behavior in a 3-dimensional (3D) environment compared to 2D, we asked whether excess centrosomes induce senescence in 3D. We used a sprouting angiogenesis assay and found that centrosome over-duplicated ECs in 3D sprouts showed SA-β-gal activity, suggesting that they underwent senescence. In summary, we show that excess centrosomes induced p53-dependent senescence in endothelial cells in both 2D and 3D. Our work provides a novel association between excess centrosomes and senescence, which will help to better understand regulation of the tumor microenvironment. Citation Format: Zhixian Yu, Erich Kushner, Victoria Bautch. Excess centrosomes induce p53-dependent senescence in endothelial cells. [abstract]. In: Proceedings of the AACR Precision Medicine Series: Cancer Cell Cycle - Tumor Progression and Therapeutic Response; Feb 28-Mar 2, 2016; Orlando, FL. Philadelphia (PA): AACR; Mol Cancer Res 2016;14(11_Suppl):Abstract nr A20.