MicroRNA-mediated loss of PDEF protein expression results in progression to more invasive breast cancer

1569 The purpose of this study is to elucidate possible mechanism(s) of PDEF protein loss during breast cancer progression. Cancer death is due in large part to metastases. One of the more interesting challenges is to understand the cellular changes that occur during progression towards more invasive cancer. PDEF is an Ets transcription factor with expression restricted to tissues with high epithelial content, including breast, prostate and colon. At the mRNA level PDEF overexpression has been proposed as a diagnostic marker of invasive cancer. However, PDEF protein is often reduced in invasive tissue and lost in invasive cancer cells. Indeed, re-expression of PDEF protein in invasive cells lines inhibits cell growth, migration and invasion. To date, the processes involved in the loss of PDEF protein during cancer progression have not been elucidated. The emerging field of microRNAs (miRNAs) has led to the functional discovery of their role in many types of cancer. However, the identification and validation of specific targets has been limited. miRNAs function post-transcriptionally by promoting either mRNA degradation or inhibiting mRNA translation. To explore the potential role for miRNAs in the post-transcriptional regulation of PDEF, a bioinformatics approach identified multiple miRNA recognition sequences within the 3’ untranslated region (UTR) of PDEF. Two miRNAs, miRNA 204 (miR204) and 510 (miR510) were found to be elevated in breast cancer cell lines having high RNA and low PDEF protein levels. Two cell lines were selected as models to test the functional role of these miRNAs in regulation of PDEF protein expression: CAMA-1 (high mRNA, low protein) and MCF7 (high mRNA, high protein). Inhibition of miR204 or miR510 by antisense oligoribonucleotides in CAMA-1 results in an increase in PDEF protein levels without significant affects on PDEF mRNA. Reciprocal overexpression of either miR204 or miR510 in MCF7 cells results in a loss of PDEF protein, without significantly affecting the levels of PDEF mRNA. The functional consequences of miRNA-mediated loss of PDEF include an altered cellular morphology, increased cellular migration and invasion and a more transformed phenotype. Furthermore, we demonstrate that these cellular changes occur through the altered regulation of known PDEF transcriptional targets. In addition, we show for the first time that expression of miR-204 and miR-510 are elevated in human breast tumor samples compared to matched non-tumor samples. Collectively, this study provides for the first time a mechanism for the loss of PDEF protein expression and a unique oncogenic role for these miRNAs during metastatic breast cancer progression.