Abstract
Ewing sarcoma is a bone malignancy of children and young adults, frequently harboring the EWS/FLI chromosomal translocation. The resulting fusion protein is an aberrant transcription factor that uses highly repetitive GGAA-containing elements (microsatellites) to activate and repress thousands of target genes mediating oncogenesis. However, the mechanisms of EWS/FLI interaction with microsatellites and regulation of target gene expression is not clearly understood. Here, we profile genome-wide protein binding and gene expression. Using a combination of unbiased genome-wide computational and experimental analysis, we define GGAA-microsatellites in a Ewing sarcoma context. We identify two distinct classes of GGAA-microsatellites and demonstrate that EWS/FLI responsiveness is dependent on microsatellite length. At close range “promoter-like” microsatellites, EWS/FLI binding and subsequent target gene activation is highly dependent on number of GGAA-motifs. “Enhancer-like” microsatellites demonstrate length-dependent EWS/FLI binding, but minimal correlation for activated and none for repressed targets. Our data suggest EWS/FLI binds to “promoter-like” and “enhancer-like” microsatellites to mediate activation and repression of target genes through different regulatory mechanisms. Such characterization contributes valuable insight to EWS/FLI transcription factor biology and clarifies the role of GGAA-microsatellites on a global genomic scale. This may provide unique perspective on the role of non-coding DNA in cancer susceptibility and therapeutic development.
Highlights
Ewing sarcoma is the second most common pediatric bone malignancy, initiated by a chromosomal translocation t(11;22)(q24;q12), creating the fusion protein and oncogenic driver EWS/ FLI
Gene-associated GGAA-microsatellites serve as DNA response elements for EWS/FLI to bind and mediate transcriptional activation of its up-regulated targets [12,16,35,37]
In this study we describe microsatellites on a global genomic scale, and use Chromatin immunoprecipitation (ChIP)-seq and RNA-seq analysis to computationally investigate EWS/FLI responsiveness at these repetitive elements
Summary
Ewing sarcoma is the second most common pediatric bone malignancy, initiated by a chromosomal translocation t(11;22)(q24;q12), creating the fusion protein and oncogenic driver EWS/ FLI. As an aberrant transcription factor, EWS/FLI plays a critical role in regulating genes involved in tumorigenesis [1]. FLI and other ETS family members bind DNA via their conserved DNA binding domain at the consensus sequence ‘ACCGGAAGTG’ [2,3]. Institute [Grant R01 CA140394 and R01 CA183776], and funds awarded to K.M.J. from the National Cancer Institute [Grant F30 CA21058. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript
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