Coordinated epigenetic repression of the miR-200 family and miR-205 in invasive bladder cancer

MicroRNAs (miRNA) are small noncoding RNAs commonly deregulated in cancer. The miR-200 family (miR-200a, -200b, -200c, -141 and -429) and miR-205 are frequently silenced in advanced cancer and have been implicated in epithelial to mesenchymal transition (EMT) and tumor invasion by targeting the transcriptional repressors of E-cadherin, ZEB1 and ZEB2. ZEB1 is also known to repress miR-200c-141 transcription in a negative feedback loop, but otherwise little is known about the transcriptional regulation of the miR-200 family and miR-205. Recently, miR-200 silencing was also reported in cancer stem cells, implying that miR-200 deregulation is a key event in multiple levels of tumor biology. However, what prevents miR-200 expression remains largely unanswered. Here we report concerted transcriptional regulation of the miR-200 and miR-205 loci in bladder tumors and bladder cell lines. Using a combination of miRNA expression arrays, qPCR assays and mass spectrometry DNA methylation analyses, we show that the miR-200 and miR-205 loci are specifically silenced and gain promoter hypermethylation and repressive chromatin marks in muscle invasive bladder tumors and undifferentiated bladder cell lines. Moreover, we report that miR-200c expression is significantly correlated with early stage T1 bladder tumor progression, and propose miR-200 and miR-205 silencing and DNA hypermethylation as possible prognostic markers in bladder cancer. In addition, we observe that the mesoderm transcription factor TWIST1 and miR-200 expression are inversely correlated in bladder tumor samples and cell lines. TWIST1 associates directly with the miR-200 and miR-205 promoters, and may act as a repressor of miR-200 and miR-205 expression.