ABSTRACT
Epigenetic changes on DNA and chromatin are implicated in cell differentiation and organogenesis. For the heart, distinct histone methylation profiles were recently linked to stage-specific gene expression programs during cardiac differentiation in vitro. However, the enzymes catalyzing these modifications and the genes regulated by them remain poorly defined. We therefore decided to identify the epigenetic enzymes that are potentially involved in cardiomyogenesis by analyzing the expression profile of the 85 genes encoding the epigenetic-related proteins in mouse cardiomyocytes (CMs), and then study how they affect gene expression during differentiation and maturation of this cell type. We show here with gene expression screening of epigenetic enzymes that the highly expressed H3 methyltransferase disruptor of telomeric silencing 1-like (DOT1L) drives a transitional pattern of di-methylation on H3 lysine 79 (H3K79) in CMs at different stages of differentiation in vitro and in vivo. Through a genome-wide chromatin-immunoprecipitation DNA-sequencing approach, we found H3K79me2 enriched at genes expressed during cardiac differentiation. Moreover, knockdown of Dot1L affected the expression of H3K79me2-enriched genes. Our results demonstrate that histone methylation, and in particular DOT1L-mediated H3K79me2 modification, drives cardiomyogenesis through the definition of a specific transcriptional landscape.
Subject(s)
Cell Differentiation , Gene Expression Regulation , Histones/metabolism , Methyltransferases/metabolism , Myocytes, Cardiac/metabolism , Protein Processing, Post-Translational , Animals , Cell Line , Histone-Lysine N-Methyltransferase , Histones/genetics , Methyltransferases/genetics , MiceABSTRACT
By integrating gene profiling and immunohistochemical data with functional experiments in cell lines in this study we show for the first time that doublecortin (DCX) domain containing 2 (DCDC2), a protein belonging to the DCX family and involved in neuronal cell migration, is aberrantly expressed in prostate tumors whereas absent in normal prostate. Furthermore, in patients treated with radical prostatectomy, high levels of DCDC2 RNA were significantly associated with increased biochemical relapse (LogRank Mantel-Cox=0.012). Mechanistically, we found that the ETS transcription factor ESE3/EHF, which is expressed in normal prostate and frequently lost in prostate tumors, maintained DCDC2 repressed by binding to a novel identified ETS binding site in the gene promoter. Consistently, in prostate tumors and in cellular models of gain and loss of ESE3/EHF, the expression of DCDC2 and ESE3/EHF were inversely correlated. In prostate cancer cells, DCDC2 colocalized with microtubules and promoted cell migration and resistance to the microtubule-targeting drug taxol. Collectively, this study establishes DCDC2 as a novel ESE3/EHF oncogenic target in prostate cancer. These findings may be relevant for the clinical management of prostate cancer as DCDC2 may signal tumors more prone to relapse and resistant to taxol treatment.
Subject(s)
Drug Resistance, Neoplasm/genetics , Microtubule-Associated Proteins/genetics , Prostatic Neoplasms/genetics , Prostatic Neoplasms/pathology , Binding Sites , Cell Movement/genetics , Disease Progression , Gene Expression Regulation, Neoplastic , Humans , Male , Microtubule-Associated Proteins/metabolism , Microtubules/metabolism , Paclitaxel/pharmacology , Promoter Regions, Genetic , Prostatic Neoplasms/drug therapy , Prostatic Neoplasms/mortality , Prostatic Neoplasms/surgery , Reference Values , Transcription Factors/genetics , Transcription Factors/metabolism , Tubulin Modulators/pharmacologyABSTRACT
Epigenetic silencing of tumour suppressor genes is an important mechanism involved in cell transformation and tumour progression. The Set and RING-finger-associated domain-containing protein UHRF1 might be an important link between different epigenetic pathways. Here, we report that UHRF1 is frequently overexpressed in human prostate tumours and has an important role in prostate cancer pathogenesis and progression. Analysis of human prostate cancer samples by microarrays and immunohistochemistry showed increased expression of UHRF1 in about half of the cases. Moreover, UHRF1 expression was associated with reduced overall survival after prostatectomy in patients with organ-confined prostate tumours (P < 0.0001). UHRF1 expression was negatively correlated with several tumour suppressor genes and positively with the histone methyltransferase (HMT) EZH2 both in prostate tumours and cell lines. UHRF1 knockdown reduced proliferation, clonogenic capability and anchorage-independent growth of prostate cancer cells. Depletion of UHRF1 resulted in reactivation of several tumour suppressor genes. Gene reactivation upon UHRF1 depletion was associated with changes in histone H3K9 methylation, acetylation and DNA methylation, and impaired binding of the H3K9 HMT Suv39H1 to the promoter of silenced genes. Co-immunoprecipitation experiments showed direct interaction between UHRF1 and Suv39H1. Our data support the notion that UHRF1, along with Suv39H1 and DNA methyltransferases, contributes to epigenetic gene silencing in prostate tumours. This could represent a parallel and convergent pathway to the H3K27 methylation catalyzed by EZH2 to synergistically promote inactivation of tumour suppressor genes. Deregulated expression of UHRF1 is involved in the prostate cancer pathogenesis and might represent a useful marker to distinguish indolent cancer from those at high risk of lethal progression.
