A Tumor Suppressive Role of CYLD as a Novel Potential DUB of Aurora B in Cervical Cancer.

Abstract

Cervical cancer is a common leading cause of cancer related to women death worldwide. Cylindromatosis (CYLD) is known as an important tumor suppressor in various human cancers, and a deubiquitination enzyme (DUB) as well. Previously, we identified Skp2 as an E3 ligase of Aurora B ubiquitination, but the DUB of Aurora B still remains unknown. Aurora B ubiquitination site is identified through in vivo ubiquitination assay. Activity of Aurora B and CENPA was detected by immunoblotting (IB) and immunofluorescence (IF) assay. Protein-to-protein interaction was investigated by immunoprecipitation (IP). Cell chromosome dynamics was monitored by live-cell time-lapse Imaging. Cancer cell proliferation, colony formation, apoptosis, and cell invasion and migration assays were also performed. Protein level was checked by immunohistochemical (IHC) staining in clinical cervical cancer samples. We identified Lysine 115 (K115) as the main Aurora B ubiquitination site for Skp2. We could also detect an interaction of Aurora B with the DUB CYLD. We found that CYLD promoted deubiquitination of Aurora B, and regulated Aurora B activity and function as well. Compared with control, we found it took more time for the cells to finish cell mitosis with CYLD over-expression. Furthermore, we found that CYLD deficiency promoted cervical cancer cell proliferation, colony formation, cell migration and invasion, and inhibited apoptosis instead, whereas it is just opposite with CYLD over-expression. In clinical cervical cancer samples, we showed a negative correlation of CYLD expression with Aurora B activation and histological cancer cell invasion. Furthermore, there was less CYLD abundance and higher Aurora B activity in advanced cancer samples compared with early stage. Our findings uncover CYLD as a novel potential DUB of Aurora B, which inhibits Aurora B activation and its subsequent function in cell mitosis, and also provide more evidence for its tumor suppressor function in cervical cancer.