Epigenetic Regulation Of RhoJ Expression And Its Implications In Breast Cancer Metastasis

Objectives:RhoJ/TCL was initially identified as a novel Rho GTPase related to TC10 (RhoQ),belonging to the Cdc42 subfamily of Rho GTPases. RhoJ, highly expressed in endothelial cells,has a critical role in mediating endothelial cell motility and tube formation and modulating actomyosin contractility and focaladhesion numbers. To date, its role in carcinogenesis is largely obscure.Breast cancer is a significant cause of worldwide morbidity and mortality in females. Based on gene expression profiling,6 different molecular subtypes of breast cancer have been established, including luminal A,luminal B,HER2-enriched,basal-like,claudin-low and normal breast-like groups. Among the six subtypes,basal-like breast cancer has drawn particular attention and is associated with poor prognosis.Whether there is a relationship between RhoJ and the malignancy of breast cancer is still unknown so far. RhoJ was identified as a potential target of myocardin-related transcription factor A (MRTF-A, also known as MKL1) using microarray screening. Since MRTF-A is known to regulate cancer cell migration and invasion, we hypothesized that RhoJ might be a target for MRTF-A involved in the regulation of breast cancer cell metastasis.Methods:We chose Luminal-type cells MCF-7, T47D and Basal-like type cells MDA-MB-231,Hs578T. mRNA levels of RhoGTPase were examined by real-time quantitative PCR (qPCR) in these two different subtypes of cells. Transwell and scratch assays were performed to detect the changes of migration and invasion in MDA-MB-231 and Hs578T cells. Luciferase reporter assay was performed to assess the effect of MRTF-A and ERG on RhoJ transcription. Chromatin immunoprecipitation (ChIP) assay was performed to detect occupancies of MRTF-A and histones on the RhoJ promoter.Results:RhoJ expression levels were correlated with malignancies of breast cancer cells. Silencing RhoJ or ERG decreased the migration and invasion abilities of breast cancer cells. MRTF-A and ERG activated RhoJ transcription. TGF-P 1 up-regulated mRNA levels of RhoJ and augmented the accumulation of MRTF-A on RhoJ promoter, which relied on ERG. Breast cancer cells of different malignancies exhibited different occupancies of histones H3K9Me3, H3K27Me3 and H4K20Me3 on RhoJ promoter, which might be mediated by the demethylase Jmjd2b.Conclusion:ERG recruits MRTF-A to activate RhoJ transcription in breast cancer cells. MRTF-A/ERG dependent transactivation of RhoJ might be mediated by Jmjd2b catalyzed histone demethylation of H3K9Me3, H3K27Me3 and/or H4K20Me3, enabling the metastatic process. Therefore, our study provides a potential target for clinical intervention of breast cancer.