Journal
JOURNAL OF BIOLOGICAL CHEMISTRY
Volume 285, Issue 18, Pages 13444-13453Publisher
AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC
DOI: 10.1074/jbc.M109.062430
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Funding
- Ministry of Education, Culture, Sports, Science, and Technology [19590553]
- Ministry of Health and Welfare
- Kitamura Memorial Foundation, Japan
- Grants-in-Aid for Scientific Research [19590553] Funding Source: KAKEN
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Pregnant women show a low level of protein S (PS) in plasma, which is known to be a risk for deep venous thrombosis. 17 beta-Estradiol (E-2), an estrogen that increases in concentration in the late stages of pregnancy, regulates the expression of various genes via the estrogen receptor (ER). Here, we investigated the molecular mechanisms behind the reduction in PS levels caused by E-2 in HepG2-ER alpha cells, which stably express ER alpha, and also the genomic ER signaling pathway, which modulates the ligand-dependent repression of the PS alpha gene (PROS1). We observed that E-2 repressed the production of mRNA and antigen of PS. A luciferase reporter assay revealed that E-2 down-regulated PROS1 promoter activity and that this E-2-dependent repression disappeared upon the deletion or mutation of two adjacent GC-rich motifs in the promoter. An electrophoretic mobility shift assay and DNA pulldown assay revealed that the GC-rich motifs were associated with Sp1, Sp3, and ER alpha. In a chromatin immunoprecipitation assay, we found ER alpha-Sp protein-promoter interaction involved in the E-2-dependent repression of PROS1 transcription. Furthermore, we demonstrated that E-2 treatment recruited RIP140 and the NCoR-SMRT-HDAC3 complex to the PROS1 promoter, which hypoacetylated chromatin. Taken together, this suggested that E-2 might repress PROS1 transcription depending upon ER alpha-Sp1 recruiting transcriptional repressors in HepG2-ER alpha cells and, consequently, that high levels of E-2 leading to reduced levels of plasma PS would be a risk for deep venous thrombosis in pregnant women.
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