Crucial role for early growth response-1 in the transcriptional regulation of miR-20b in breast cancer.
Transcriptional regulation of miRNAs that control the pathogenesis of breast cancer remains largely unknown. Here, we showed that ionizing radiation, a known breast carcinogen, triggered the differential expression of miR-20b in mammary tissues. We identified several GC-rich consensus binding motifs for the zinc finger transcription factor early growth response-1 (EGR1) in miR-20b promoter. miR-20b was upregulated by IR and its upregulation correlated with EGR1 expression in the breast cancer cell line HCC1806. Therefore, we used HCC1806 cells as a model system to explore the role of EGR1 in miR-20b transcription. siRNA knockdown of EGR1 attenuated miR-20b expression. Luciferase assays showed that whereas EGR1 stimulated luciferase activity driven by the wild-type miR-20b promoter, this induction was abolished in the mutant miR-20 promoter construct. We noted significant enrichment of EGR1 at miR-20b promoter in HCC1806 cells compared with normal human mammary epithelial cells. Suppression of miR-20b significantly inhibited HCC1806 cell proliferation and migration, and led to G0/G1 and S phase arrest. In vitro RNA-pull down assays indicated that miR-20b targets numerous tumor suppressors, including PTEN and BRCA1, which were downregulated in HCC1806. Conversely, suppression of miR-20b increased PTEN and BRCA1 levels. Moreover, immunohistochemical and FISH analyses showed that the miR-20b expression correlated significantly with EGR1 levels in breast cancer tissues. Our findings thus demonstrate for the first time that EGR1 is a key player in the transcriptional control of miR-20b, and miR-20b may in turn function as an oncogene by contributing to breast tumorigenesis via tumor suppressor targeting.
Department of Biological Sciences, University of Lethbridge, Lethbridge, Canada.
Dongping
D
Li
Yaroslav
Y
Ilnytskyy
Anna
A
Kovalchuk
Levon M
LM
Khachigian
Roderick T
RT
Bronson
Bo
B
Wang
Olga
O
Kovalchuk
D000818:Animals; D019313:BRCA1 Protein / Q000235:genetics / Q000378:metabolism; D001943:Breast Neoplasms / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D002453:Cell Cycle / Q000235:genetics; D048708:Cell Growth Processes / Q000235:genetics; D045744:Cell Line, Tumor; D002465:Cell Movement / Q000235:genetics; D015536:Down-Regulation; D051766:Early Growth Response Protein 1 / Q000235:genetics* / Q000378:metabolism; D005260:Female; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D007150:Immunohistochemistry; D035683:MicroRNAs / Q000235:genetics* / Q000378:metabolism; D051059:PTEN Phosphohydrolase / Q000096:biosynthesis / Q000235:genetics; D051381:Rats; D020318:Rats, Long-Evans; D015398:Signal Transduction; D014158:Transcription, Genetic; D014162:Transfection
D019313:BRCA1 Protein; C492913:BRCA1 protein, human; C497325:EGR1 protein, human; D051766:Early Growth Response Protein 1; C534451:MIRN20a microRNA, human; D035683:MicroRNAs; D051059:PTEN Phosphohydrolase; C494929:PTEN protein, human