Runx2 transcriptome of prostate cancer cells: insights into invasiveness and bone metastasis

Sanjeev K Baniwal,Gerhard A Coetzee,Deepak Mav,Alice E Kohn-Gabet,Yunfan Shi,Yankel Gabet,Daniel J Purcell,Ruchir R Shah,Baruch Frenkel,Omar Khalid

doi:10.1186/1476-4598-9-258

Abstract

BackgroundProstate cancer (PCa) cells preferentially metastasize to bone at least in part by acquiring osteomimetic properties. Runx2, an osteoblast master transcription factor, is aberrantly expressed in PCa cells, and promotes their metastatic phenotype. The transcriptional programs regulated by Runx2 have been extensively studied during osteoblastogenesis, where it activates or represses target genes in a context-dependent manner. However, little is known about the gene regulatory networks influenced by Runx2 in PCa cells. We therefore investigated genome wide mRNA expression changes in PCa cells in response to Runx2.ResultsWe engineered a C4-2B PCa sub-line called C4-2B/Rx2dox, in which Doxycycline (Dox) treatment stimulates Runx2 expression from very low to levels observed in other PCa cells. Transcriptome profiling using whole genome expression array followed by in silico analysis indicated that Runx2 upregulated a multitude of genes with prominent cancer associated functions. They included secreted factors (CSF2, SDF-1), proteolytic enzymes (MMP9, CST7), cytoskeleton modulators (SDC2, Twinfilin, SH3PXD2A), intracellular signaling molecules (DUSP1, SPHK1, RASD1) and transcription factors (Sox9, SNAI2, SMAD3) functioning in epithelium to mesenchyme transition (EMT), tissue invasion, as well as homing and attachment to bone. Consistent with the gene expression data, induction of Runx2 in C4-2B cells enhanced their invasiveness. It also promoted cellular quiescence by blocking the G1/S phase transition during cell cycle progression. Furthermore, the cell cycle block was reversed as Runx2 levels declined after Dox withdrawal.ConclusionsThe effects of Runx2 in C4-2B/Rx2dox cells, as well as similar observations made by employing LNCaP, 22RV1 and PC3 cells, highlight multiple mechanisms by which Runx2 promotes the metastatic phenotype of PCa cells, including tissue invasion, homing to bone and induction of high bone turnover. Runx2 is therefore an attractive target for the development of novel diagnostic, prognostic and therapeutic approaches to PCa management. Targeting Runx2 may prove more effective than focusing on its individual downstream genes and pathways.

Highlights

Runx2 together with Runx1 and Runx3 comprise the Runx class of transcription factors, defined by their highly homologous Runt-related DNA-binding domain
RT-qPCR analysis revealed that the Dox treatment increased Runx2 mRNA by ~20-fold compared to its endogenous levels, and that the induced level was comparable to that observed in the PC3high sub-line (Figure 1C)
Western analysis using anti-Runx2 antibodies indicated that the level of endogenous Runx2 protein was negligible in untreated C4-2B cells, and that Dox induced expression of the exogenous Runx2 to the levels normally found in osteoblasts (Figure 1D) [41]

Summary

Introduction

Runx together with Runx and Runx comprise the Runx class of transcription factors, defined by their highly homologous Runt-related DNA-binding domain. Among the osteomimetic properties of prostate and breast cancer cells are expression of the Runx target genes MMP9 [28], BSP [29] and VEGFA [30], as well as induction of mineralization [25]. Transcriptome profiling using whole genome expression array followed by in silico analysis indicated that Runx upregulated a multitude of genes with prominent cancer associated functions They included secreted factors (CSF2, SDF-1), proteolytic enzymes (MMP9, CST7), cytoskeleton modulators (SDC2, Twinfilin, SH3PXD2A), intracellular signaling molecules (DUSP1, SPHK1, RASD1) and transcription factors (Sox, SNAI2, SMAD3) functioning in epithelium to mesenchyme transition (EMT), tissue invasion, as well as homing and attachment to bone. Consistent with the gene expression data, induction of Runx in C4-2B cells enhanced their invasiveness It promoted cellular quiescence by blocking the G1/S phase transition during cell cycle progression. Targeting Runx may prove more effective than focusing on its individual downstream genes and pathways

Methods

Results

Conclusion