To ensure the accuracy of chromosome segregation during cell division, eukaryotic cells have developed a surveillance system, the mitotic checkpoint, which delays anaphase onset until all kinetochores are properly attached to microtubules emanating from opposite spindle poles. One of the core components of this checkpoint is Bub1 protein. Bub1 protein is encoded in the BUB1 gene, which is known to have a low basal levels of transcripts during interphase and increases its transcriptional activity in G2 and mitosis. However, the mechanisms involved in the regulation of BUB1 are not fully understood. Some of the possible components that participate in the transcriptional regulation of BUB1 are the cell cycle-dependent element (CDE) and the cell cycle homology region (CHR) element. The CDE/CHR are DNA sequences of 6 nucleotides, rich in CGs, located at proximal promoter regions of genes, and recruit repressive and activation factors during the initial and late phases of cell cycle, respectively. For instance, the promoters of CDC25C, CCNA2 and CDK1 genes contain the CDE/CHR elements and have high expression in G2 and mitosis. Thus, the aim of this study was to identify the mechanisms involved in the transcriptional regulation of BUB1 in a cell cycle dependent manner, using RT-qPCR, immunoblotting, flow cytometry, luciferase reporter gene assay, Methyl Sensitive-PCR. In this study, we unravel the mechanisms involved in the transcriptional regulation of BUB1 through the cell cycle. We found that it's expressed in a cell cycle-dependent manner in HeLa cells, being highly expressed during G2/M, followed by a rapid decline of both the mRNA and protein levels at the G1 phase. Also, we identified that the CDE/CHR elements are located at the BUB1 promoter and participate in its repression during the early phases of cell cycle, and his activation in G2 and mitosis. Finally, we found that the BUB1 promoter is located inside a CpG island which is mainly unmethylated in cancer cell lines regardless the cell type, indicating that DNA methylation might not be a determinant factor involved in its transcription. However, it's still unclear whether DNA methylation can affect BUB1 transcriptional activity through the cell cycle, particularly the methylation at specific CpG sites, such as those contained in the CDE and CHR elements. In summary, BUB1 transcription is regulated in a cell cycle dependent manner by the CDE/CHR elements located at its promoter. Also, DNA methylation is not a determinant factor implicated in its transcriptional activity.

Nevertheless, whether BUB1 transcriptional deregulation can contribute towards the development of a cancer phenotype is still unclear. Thus, this study contributes to the understanding of the mechanisms implicated on the transcriptional deregulation of cell cycle genes, such as BUB1.

Citation Format: Fernando Luna, Marco A. Andonegui, Fernanda Cisneros, Alfredo Cantu, Luis A. Herrera. Transcription of BUB1 is regulated by the CDE and CHR elements through the cell cycle [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1475.