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Crucial role for early growth response-1 in the transcriptional regulation of miR-20b in breast cancer.

Li D, Ilnytskyy Y, Kovalchuk A, Khachigian LM, Bronson RT, Wang B, Kovalchuk O - Oncotarget (2013)

Bottom Line: 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.Conversely, suppression of miR-20b increased PTEN and BRCA1 levels.

View Article: PubMed Central - PubMed

Affiliation: Department of Biological Sciences, University of Lethbridge, Lethbridge, Canada.

ABSTRACT
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.

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miR-20b transcriptionally activated by EGR1 directly targets PTEN and BRCA1 in breast cancerSerum-inducible zinc finger transcription factor EGR1 is induced and activated in response to a wide range of extracellular stimuli, including growth factors, cytokines, UV light, ionizing radiation, and mechanical injury. Once activated, EGR1 translocates into nucleus and binds to the consensus motifs at miR-20b promoter, leading to miR-20b transcription. The mature miR-20b assembles with other proteins to form RNA-induced silencing complex (RISC), the later recognizes and binds to PTEN and BRCA1 mRNAs, leading to either translational suppression or degradation of those two molecules, consequently resulting in breast cancer cell proliferation and migration.
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Figure 7: miR-20b transcriptionally activated by EGR1 directly targets PTEN and BRCA1 in breast cancerSerum-inducible zinc finger transcription factor EGR1 is induced and activated in response to a wide range of extracellular stimuli, including growth factors, cytokines, UV light, ionizing radiation, and mechanical injury. Once activated, EGR1 translocates into nucleus and binds to the consensus motifs at miR-20b promoter, leading to miR-20b transcription. The mature miR-20b assembles with other proteins to form RNA-induced silencing complex (RISC), the later recognizes and binds to PTEN and BRCA1 mRNAs, leading to either translational suppression or degradation of those two molecules, consequently resulting in breast cancer cell proliferation and migration.

Mentions: This study for the first time demonstrated that EGR1 regulated miR-20b transcription and provided important clues on the role of miR-20b in breast tumorigenesis (Fig. 7). Breast cancer is a multifactorial and multistage process that involves many environmental and genetic factors. Among the environmental factors that cause breast cancer, IR may be one of the high-risk factors because it has been shown to strongly induce breast cancer in exposed individuals[2].Furthermore, the IR-induced mouse breast cancer model has been widely used in the field of breast cancer research. Unfortunately, the epigenetic mechanisms underlying IR-induced mammary carcinogenesis largely remain unknown. We showed in this paper that IR triggered miR-20b expression in mammary gland tissues in a dose- and time-dependent manner (Fig. 1C). Although IR is a putative inducer of genomic instability, including gene amplification [24], miR-20b gene copy number in our case did not contribute to IR-induced miR-20b expression (Fig. S1 and Fig. 1D).


Crucial role for early growth response-1 in the transcriptional regulation of miR-20b in breast cancer.

Li D, Ilnytskyy Y, Kovalchuk A, Khachigian LM, Bronson RT, Wang B, Kovalchuk O - Oncotarget (2013)

miR-20b transcriptionally activated by EGR1 directly targets PTEN and BRCA1 in breast cancerSerum-inducible zinc finger transcription factor EGR1 is induced and activated in response to a wide range of extracellular stimuli, including growth factors, cytokines, UV light, ionizing radiation, and mechanical injury. Once activated, EGR1 translocates into nucleus and binds to the consensus motifs at miR-20b promoter, leading to miR-20b transcription. The mature miR-20b assembles with other proteins to form RNA-induced silencing complex (RISC), the later recognizes and binds to PTEN and BRCA1 mRNAs, leading to either translational suppression or degradation of those two molecules, consequently resulting in breast cancer cell proliferation and migration.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3824527&req=5

Figure 7: miR-20b transcriptionally activated by EGR1 directly targets PTEN and BRCA1 in breast cancerSerum-inducible zinc finger transcription factor EGR1 is induced and activated in response to a wide range of extracellular stimuli, including growth factors, cytokines, UV light, ionizing radiation, and mechanical injury. Once activated, EGR1 translocates into nucleus and binds to the consensus motifs at miR-20b promoter, leading to miR-20b transcription. The mature miR-20b assembles with other proteins to form RNA-induced silencing complex (RISC), the later recognizes and binds to PTEN and BRCA1 mRNAs, leading to either translational suppression or degradation of those two molecules, consequently resulting in breast cancer cell proliferation and migration.
Mentions: This study for the first time demonstrated that EGR1 regulated miR-20b transcription and provided important clues on the role of miR-20b in breast tumorigenesis (Fig. 7). Breast cancer is a multifactorial and multistage process that involves many environmental and genetic factors. Among the environmental factors that cause breast cancer, IR may be one of the high-risk factors because it has been shown to strongly induce breast cancer in exposed individuals[2].Furthermore, the IR-induced mouse breast cancer model has been widely used in the field of breast cancer research. Unfortunately, the epigenetic mechanisms underlying IR-induced mammary carcinogenesis largely remain unknown. We showed in this paper that IR triggered miR-20b expression in mammary gland tissues in a dose- and time-dependent manner (Fig. 1C). Although IR is a putative inducer of genomic instability, including gene amplification [24], miR-20b gene copy number in our case did not contribute to IR-induced miR-20b expression (Fig. S1 and Fig. 1D).

Bottom Line: 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.Conversely, suppression of miR-20b increased PTEN and BRCA1 levels.

View Article: PubMed Central - PubMed

Affiliation: Department of Biological Sciences, University of Lethbridge, Lethbridge, Canada.

ABSTRACT
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.

Show MeSH
Related in: MedlinePlus