Identification of DLX1 as a FOXM1 downstream target in mediating ovarian cancer oncogenesis

Emerging evidences have documented that aberrant expression of FOXM1 is closely associated with human cancers. A recent comprehensive genome analysis has revealed that FOXM1 signaling is one of the major pathways involved in ovarian cancer oncogenesis. However, the regulatory network of FOXM1 in exerting the metastatic phenotypes remains unknown. Therefore, the identification of FOXM1 downstream targets will assist in understanding of its molecular mechanism in ovarian cancer oncogenesis. In this study, by bioinformatics and a series of functional analyses, we identified DLX1 as a novel target of FOXM1. Our results clearly demonstrated that enforced expression of FOXM1 (FOXM1B and FOXM1C) could increase DLX1 in mRNA and protein levels. Conversely, depletion of FOXM1 by Thiostrepton (FOXM1 specific inhibitor) or RNAi knockdown could reduce DLX1 expression. Importantly, we demonstrated that the changes of DLX1 expression were in concomitant with the expression of a positive control gene, Cyclin-D1. Additionally, the luciferase promoter assay further showed that there are two conserved FOXM1 binding sites TFBS1 and TFBS2 which located at -61~-52bp upstream and -737~727bp upstream of the transcription factor binding sites (TSS) of DLX1 promoter respectively. In comparison of two binding sites, the more conserved binding site, TFBS1, seems have higher importance of FOXM1 binding in DLX1 transcriptional activation. Furthermore, our study using immunohistochemical and Q-PCR analyses showed that DLX1 was frequently up-regulated in ovarian cancer samples. Noticeably, clinicopathological analysis revealed that the upregulated DLX1 was significantly associated with not only the overexpressed FOXM1 (P=0.001) but also high grade ovarian cancer (P