EWS/FLI1 regulates EYA3 in Ewing sarcoma via modulation of miRNA-708, resulting in increased cell survival and chemoresistance.
Ewing sarcoma is an aggressive pediatric cancer of the bone and soft tissue, in which patients whose tumors have a poor histologic response to initial chemotherapy have a poor overall prognosis. Therefore, it is important to identify molecules involved in resistance to chemotherapy. Herein, we show that the DNA repair protein and transcriptional cofactor, EYA3, is highly expressed in Ewing sarcoma tumor samples and cell lines compared with mesenchymal stem cells, the presumed cell-of-origin of Ewing sarcoma, and that it is regulated by the EWS/FLI1 fusion protein transcription factor. We further show that EWS/FLI1 mediates upregulation of EYA3 via repression of miR-708, a miRNA that targets the EYA3 3'-untranslated region, rather than by binding the EYA3 promoter directly. Importantly, we show that high levels of EYA3 significantly correlate with low levels of miR-708 in Ewing sarcoma samples, suggesting that this miR-mediated mechanism of EYA3 regulation holds true in human cancers. Because EYA proteins are important for cell survival during development, we examine, and show, that loss of EYA3 decreases survival of Ewing sarcoma cells. Most importantly, knockdown of EYA3 in Ewing sarcoma cells leads to sensitization to DNA-damaging chemotherapeutics used in the treatment of Ewing sarcoma, and as expected, after chemotherapeutic treatment, EYA3 knockdown cells repair DNA damage less effectively than their control counterparts. These studies identify EYA3 as a novel mediator of chemoresistance in Ewing sarcoma and define the molecular mechanisms of both EYA3 overexpression and of EYA3-mediated chemoresistance.
Department of Pharmacology, University of Colorado School of Medicine, Aurora, CO 80045, USA.
Tyler P
TP
Robin
Anna
A
Smith
Erin
E
McKinsey
Lisa
L
Reaves
Paul
P
Jedlicka
Heide L
HL
Ford
D001859:Bone Neoplasms / Q000188:drug therapy / Q000378:metabolism / Q000473:pathology; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects / Q000235:genetics; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D004317:Doxorubicin / Q000494:pharmacology; D019008:Drug Resistance, Neoplasm / Q000235:genetics; D005047:Etoposide / Q000494:pharmacology; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D059630:Mesenchymal Stem Cells / Q000378:metabolism; D035683:MicroRNAs / Q000235:genetics / Q000378:metabolism; D015514:Oncogene Proteins, Fusion / Q000235:genetics / Q000378:metabolism; D017027:Protein Tyrosine Phosphatases / Q000235:genetics / Q000378:metabolism; D051789:Proto-Oncogene Protein c-fli-1 / Q000235:genetics / Q000378:metabolism; D034802:RNA-Binding Protein EWS / Q000235:genetics / Q000378:metabolism; D012512:Sarcoma, Ewing / Q000188:drug therapy / Q000378:metabolism / Q000473:pathology
D004268:DNA-Binding Proteins; C106031:EWS-FLI fusion protein; C568528:MIRN708 microRNA, human; D035683:MicroRNAs; D015514:Oncogene Proteins, Fusion; D051789:Proto-Oncogene Protein c-fli-1; D034802:RNA-Binding Protein EWS; D005047:Etoposide; D004317:Doxorubicin; C540714:EYA3 protein, human; D017027:Protein Tyrosine Phosphatases