PI3K/AKT signaling modulates transcriptional expression of EWS/FLI1 through specificity protein 1

Abstract

// Chiara Giorgi 1 , Aleksandar Boro 1 , Florian Rechfeld 1 , Laura A. Lopez-Garcia 1 , Maria E. Gierisch 1 , Beat W. Schäfer 1 , Felix K. Niggli 1 1 Department of Oncology and Children's Research Center, University Children's Hospital, 8032 Zurich, Switzerland Correspondence to: Beat W. Schäfer, e-mail: beat.schaefer@kispi.uzh.ch Keywords: Ewing sarcoma, EWS/FLI1, PI3K pathway, promoter analysis Received: March 19, 2015      Accepted: August 12, 2015      Published: August 22, 2015 ABSTRACT Ewing sarcoma (ES) is the second most frequent bone cancer in childhood and is characterized by the presence of the balanced translocation t(11;22)(q24;q12) in more than 85% of cases, generating a dysregulated transcription factor EWS/FLI1. This fusion protein is an essential oncogenic component of ES development which is necessary for tumor cell maintenance and represents an attractive therapeutic target. To search for modulators of EWS/FLI1 activity we screened a library of 153 targeted compounds and identified inhibitors of the PI3K pathway to directly modulate EWS/FLI1 transcription. Surprisingly, treatment of four different ES cell lines with BEZ235 resulted in down regulation of EWS/FLI1 mRNA and protein by ~50% with subsequent modulation of target gene expression. Analysis of the EWS/FLI1 promoter region (−2239/+67) using various deletion constructs identified two 14bp minimal elements as being important for EWS/FLI1 transcription. We identified SP1 as modulator of EWS/FLI1 gene expression and demonstrated direct binding to one of these regions in the EWS/FLI1 promoter by EMSA and ChIP experiments. These results provide the first insights on the transcriptional regulation of EWS/FLI1, an area that has not been investigated so far, and offer an additional molecular explanation for the known sensitivity of ES cell lines to PI3K inhibition.