Abstract
The role of aldosterone has been implicated in the metabolic syndrome and cardiovascular diseases. The biological actions of aldosterone are mediated through mineralocorticoid receptor (MR). Nuclear receptor-mediated gene expression is regulated by dynamic and coordinated recruitment of coactivators and corepressors. To identify new coregulators of the MR, full-length MR was used as bait in yeast two-hybrid screening. Weisolated NF-YC, one of the subunits of heterotrimeric transcription factor NF-Y. Specific interaction between MRand NF-YC was confirmed by yeast two-hybrid, mammalian two-hybrid, coimmunoprecipitation assays, and fluorescence subcellular imaging. Transient transfection experiments in COS-7 cells demonstrated that NF-YC repressed MR transactivation in a hormone-sensitive manner. Moreover, reduction of NF-YC protein levels by small interfering RNA potentiated hormonal activation of endogenous target genes in stably MR-expressing cells, indicating that NF-YC functions as an agonist-dependent MR corepressor. The corepressor function of NF-YC is selective for MR, because overexpression of NF-YC did not affect transcriptional activity mediated by androgen, progesterone, or glucocorticoid receptors. Chromatin immunoprecipitation experiments showed that endogenous MR and steroid receptor coactivator-1 were recruited to an endogenous ENaC gene promoter in a largely aldosterone-dependent manner, and endogenous NF-YC was sequentially recruited to the same element. Immunohistochemistry showed that endogenous MR and NF-YC were colocalized within the mouse kidney. Although aldosterone induces interaction of the N and C termini of MR, NF-YC inhibited the N/C interaction. These findings indicate that NF-YC functions as a new corepressor of agonist-bound MR via alteration of aldosterone-induced MR conformation.
| Original language | English |
|---|---|
| Pages (from-to) | 8084-8093 |
| Number of pages | 10 |
| Journal | Journal of Biological Chemistry |
| Volume | 285 |
| Issue number | 11 |
| DOIs | |
| Publication status | Published - 2010 Mar 12 |
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ASJC Scopus subject areas
- Biochemistry
- Cell Biology
- Molecular Biology
- Medicine(all)
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NF-YC functions as a corepressor of agonist-bound mineralocorticoid receptor. / Takeda, Ayano; Shibata, Hirotaka; Kurihara, Isao; Kobayashi, Sakiko; Yokota, Kenichi; Suda, Noriko; Mitsuishi, Yuko; Jo, Rie; Kitagawa, Hirochika; Kato, Shigeaki; Saruta, Takao; Itoh, Hiroshi.
In: Journal of Biological Chemistry, Vol. 285, No. 11, 12.03.2010, p. 8084-8093.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - NF-YC functions as a corepressor of agonist-bound mineralocorticoid receptor
AU - Takeda, Ayano
AU - Shibata, Hirotaka
AU - Kurihara, Isao
AU - Kobayashi, Sakiko
AU - Yokota, Kenichi
AU - Suda, Noriko
AU - Mitsuishi, Yuko
AU - Jo, Rie
AU - Kitagawa, Hirochika
AU - Kato, Shigeaki
AU - Saruta, Takao
AU - Itoh, Hiroshi
PY - 2010/3/12
Y1 - 2010/3/12
N2 - The role of aldosterone has been implicated in the metabolic syndrome and cardiovascular diseases. The biological actions of aldosterone are mediated through mineralocorticoid receptor (MR). Nuclear receptor-mediated gene expression is regulated by dynamic and coordinated recruitment of coactivators and corepressors. To identify new coregulators of the MR, full-length MR was used as bait in yeast two-hybrid screening. Weisolated NF-YC, one of the subunits of heterotrimeric transcription factor NF-Y. Specific interaction between MRand NF-YC was confirmed by yeast two-hybrid, mammalian two-hybrid, coimmunoprecipitation assays, and fluorescence subcellular imaging. Transient transfection experiments in COS-7 cells demonstrated that NF-YC repressed MR transactivation in a hormone-sensitive manner. Moreover, reduction of NF-YC protein levels by small interfering RNA potentiated hormonal activation of endogenous target genes in stably MR-expressing cells, indicating that NF-YC functions as an agonist-dependent MR corepressor. The corepressor function of NF-YC is selective for MR, because overexpression of NF-YC did not affect transcriptional activity mediated by androgen, progesterone, or glucocorticoid receptors. Chromatin immunoprecipitation experiments showed that endogenous MR and steroid receptor coactivator-1 were recruited to an endogenous ENaC gene promoter in a largely aldosterone-dependent manner, and endogenous NF-YC was sequentially recruited to the same element. Immunohistochemistry showed that endogenous MR and NF-YC were colocalized within the mouse kidney. Although aldosterone induces interaction of the N and C termini of MR, NF-YC inhibited the N/C interaction. These findings indicate that NF-YC functions as a new corepressor of agonist-bound MR via alteration of aldosterone-induced MR conformation.
AB - The role of aldosterone has been implicated in the metabolic syndrome and cardiovascular diseases. The biological actions of aldosterone are mediated through mineralocorticoid receptor (MR). Nuclear receptor-mediated gene expression is regulated by dynamic and coordinated recruitment of coactivators and corepressors. To identify new coregulators of the MR, full-length MR was used as bait in yeast two-hybrid screening. Weisolated NF-YC, one of the subunits of heterotrimeric transcription factor NF-Y. Specific interaction between MRand NF-YC was confirmed by yeast two-hybrid, mammalian two-hybrid, coimmunoprecipitation assays, and fluorescence subcellular imaging. Transient transfection experiments in COS-7 cells demonstrated that NF-YC repressed MR transactivation in a hormone-sensitive manner. Moreover, reduction of NF-YC protein levels by small interfering RNA potentiated hormonal activation of endogenous target genes in stably MR-expressing cells, indicating that NF-YC functions as an agonist-dependent MR corepressor. The corepressor function of NF-YC is selective for MR, because overexpression of NF-YC did not affect transcriptional activity mediated by androgen, progesterone, or glucocorticoid receptors. Chromatin immunoprecipitation experiments showed that endogenous MR and steroid receptor coactivator-1 were recruited to an endogenous ENaC gene promoter in a largely aldosterone-dependent manner, and endogenous NF-YC was sequentially recruited to the same element. Immunohistochemistry showed that endogenous MR and NF-YC were colocalized within the mouse kidney. Although aldosterone induces interaction of the N and C termini of MR, NF-YC inhibited the N/C interaction. These findings indicate that NF-YC functions as a new corepressor of agonist-bound MR via alteration of aldosterone-induced MR conformation.
UR - http://www.scopus.com/inward/record.url?scp=77950871339&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=77950871339&partnerID=8YFLogxK
U2 - 10.1074/jbc.M109.053371
DO - 10.1074/jbc.M109.053371
M3 - Article
C2 - 20054001
AN - SCOPUS:77950871339
VL - 285
SP - 8084
EP - 8093
JO - Journal of Biological Chemistry
JF - Journal of Biological Chemistry
SN - 0021-9258
IS - 11
ER -