Identification of Aurora-A as a Direct Target of E2F3 during G2/M Cell Cycle Progression

Yihong Ma,W Douglas Cress,W Jack Pledger,Jin Q Cheng,Lili He,Hua Yang

doi:10.1074/jbc.m803547200

Abstract

Aurora-A is a centrosome kinase and plays a pivotal role in G(2)/M cell cycle progression. Expression of Aurora-A is cell cycle-dependent. Levels of Aurora-A mRNA and protein are low in G(1)/S, accumulate during G(2)/M, and decrease rapidly after mitosis. Previous studies have shown regulation of the Aurora-A protein level during the cell cycle through the ubiquitin-proteasome pathway. However, the mechanism of transcriptional regulation of Aurora-A remains largely unknown. Here, we demonstrated that E2F3 modulates Aurora-A mRNA expression during the cell cycle. Ectopic expression of E2F3 induces Aurora-A expression. Stable knockdown of E2F3 decreases mRNA and protein levels of Aurora-A and delays G(2)/M entry. Further, E2F3 directly binds to Aurora-A promoter and stimulates the promoter activity. Deletion and mutation analyses of the Aurora-A promoter revealed that a region located 96-bp upstream of the transcription initiation site is critical for the activation of the promoter by E2F3. In addition, expression of E2F3 positively correlates with the protein level of Aurora-A in human ovarian cancer examined. These results indicate for the first time that Aurora-A is transcriptionally regulated by E2F3 during the cell cycle and that E2F3 is a causal factor for up-regulation of Aurora-A in a subset of human ovarian cancer. Thus, the E2F3-Aurora-A axis could be an important target for cancer intervention.

Highlights

The frequency of elevated Aurora-A proin vivo and to further define the E2F3 response elements in the tein and/or mRNA is much higher than its change at DNA level promoter, we carried out Chromatin Immunoprecipitation (ChIP) assay, which detects specific, suggesting the mechanism of activating transcripgenomic DNA sequences that are associated with a particular tion and/or translation of Aurora-A in ovarian cancer cells
We investigated the transcriptional regulation of Aurora-A by E2F3
Chromatin immunoprecipitation revealed that E2F3 bound to Aurora-A promoter in vivo, and the interaction is cell cycle-dependent, i.e. it primarily occurred during G2/M phase

Summary

Introduction

Basal level of Aurora-A promoter activity, especially at mitosis, was reduced in E2F3 knockdown cells as compared with pLKO.1 vector-infected HeLa cells (Fig. 4B). The frequency of elevated Aurora-A proin vivo and to further define the E2F3 response elements in the tein and/or mRNA is much higher than its change at DNA level promoter, we carried out ChIP assay, which detects specific (e.g. ϳl5%), suggesting the mechanism of activating transcripgenomic DNA sequences that are associated with a particular tion and/or translation of Aurora-A in ovarian cancer cells.

Results

Conclusion