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Histone Deacetylase 3 and 4 Complex Stimulates the Transcriptional Activity of the Mineralocorticoid Receptor.

Lee HA, Song MJ, Seok YM, Kang SH, Kim SY, Kim I - PLoS ONE (2015)

Bottom Line: The transcriptional activity of MR was significantly decreased by inhibitors of PKA (H89), PP1/2 (calyculin A), class I HDACs (MS-275), but not class II HDACs (MC1568).Interaction between MR and HDAC3 was significantly decreased by H89, calyculin A, and HDAC4 siRNA.A non-genomic effect of MR via PKA and PP1/2 induced nuclear translocation of HDAC4 to facilitate the interaction between MR and HDAC3.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmacology, School of Medicine, Kyungpook National University, Daegu, 41944, Republic of Korea; Cardiovascular Research Institute, School of Medicine, Kyungpook National University, Daegu, 41944, Republic of Korea; Cell and Matrix Research Institute, School of Medicine, Kyungpook National University, Daegu, 41944, Republic of Korea.

ABSTRACT
Histone deacetylases (HDACs) act as corepressors in gene transcription by altering the acetylation of histones, resulting in epigenetic gene silencing. We previously reported that HDAC3 acts as a coactivator of the mineralocorticoid receptor (MR). Although HDAC3 forms complexes with class II HDACs, their potential role in the transcriptional activity of MR is unclear. We hypothesized that HDAC4 of the class II family stimulates the transcriptional activity of MR. The expression of MR target genes was measured by quantitative real-time PCR. MR and RNA polymerase II recruitment to promoters of MR target genes was analyzed by chromatin immunoprecipitation. The association of MR with HDACs was investigated by co-immunoprecipitation. MR acetylation was determined with an anti-acetyl-lysine antibody after immunoprecipitation with an anti-MR antibody. Among the class II HDACs, HDAC4 interacted with both MR and HDAC3 after aldosterone stimulation. The nuclear translocation of HDAC4 was mediated by protein kinase A (PKA) and protein phosphatases (PP). The transcriptional activity of MR was significantly decreased by inhibitors of PKA (H89), PP1/2 (calyculin A), class I HDACs (MS-275), but not class II HDACs (MC1568). MR acetylation was increased by H89, calyculin A, and MS-275, but not by MC1568. Interaction between MR and HDAC3 was significantly decreased by H89, calyculin A, and HDAC4 siRNA. A non-genomic effect of MR via PKA and PP1/2 induced nuclear translocation of HDAC4 to facilitate the interaction between MR and HDAC3. Thus, we have uncovered a crucial role for a class II HDAC in the activation of MR-dependent transcription.

No MeSH data available.


Related in: MedlinePlus

HDAC4 interacts with both the MR and HDAC3.A, HEK293 cells were transfected with MR-HA and either HDAC4-Flag, HDAC5-Flag, or HDAC7-Flag plasmids. After 48 h, the cells were treated with Aldo (10 nM) for 30 min. Whole-cell lysates (WCLs) were prepared with non-denaturing lysis buffer. The MR was precipitated with an anti-HA antibody and MR-interacting HDACs were detected by western blotting with an anti-Flag antibody. HDAC4 and HDAC5 interacted with MR after Aldo treatment. B, HEK293 cells were transfected with HDAC3-Flag and HDAC4-Flag, HDAC5-Flag, or HDAC7-Flag plasmids. HDAC3 was immunoprecipitated with an antibody and HDAC3-interacting class II HDACs were detected by western blotting. HDAC4 interacted with HDAC3 after Aldo treatment. Aldo, aldosterone; HA, hemagglutinin; IP, immunoprecipitation; WB, western blotting.
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pone.0136801.g001: HDAC4 interacts with both the MR and HDAC3.A, HEK293 cells were transfected with MR-HA and either HDAC4-Flag, HDAC5-Flag, or HDAC7-Flag plasmids. After 48 h, the cells were treated with Aldo (10 nM) for 30 min. Whole-cell lysates (WCLs) were prepared with non-denaturing lysis buffer. The MR was precipitated with an anti-HA antibody and MR-interacting HDACs were detected by western blotting with an anti-Flag antibody. HDAC4 and HDAC5 interacted with MR after Aldo treatment. B, HEK293 cells were transfected with HDAC3-Flag and HDAC4-Flag, HDAC5-Flag, or HDAC7-Flag plasmids. HDAC3 was immunoprecipitated with an antibody and HDAC3-interacting class II HDACs were detected by western blotting. HDAC4 interacted with HDAC3 after Aldo treatment. Aldo, aldosterone; HA, hemagglutinin; IP, immunoprecipitation; WB, western blotting.

Mentions: We previously reported that HDAC3 binds to MR. HDAC3 deacetylates MR, which increases its transcriptional activity [1]. Similar to our results, Mihaylova et al. showed that HDAC3 deacetylates the forkhead box O (FOXO) transcription factor, the interaction of which is mediated by class IIa HDACs [17]. To identify the role of class II HDACs in the interaction between MR and HDAC3, HA-tagged MR and Flag-tagged HDAC4, 5, and 7 were co-transfected into HEK293 cells and co-immunoprecipitation (co-IP) was performed to investigate the interaction between MR and class II HDACs. HDAC4 and HDAC5 interacted with MR stimulate by Aldosterone (Aldo) (Fig 1A). To identify a class II HDAC that simultaneously binds to the MR and HDAC3, HDAC3 and each of the class II HDACs were co-expressed in HEK293 cells. Treatment with Aldo promoted an interaction between HDAC3 and HDAC4 but not with the other class II HDACs (Fig 1B). Together, these results indicate that HDAC4 interacts with MR regulators and may also play an important role in MR function.


Histone Deacetylase 3 and 4 Complex Stimulates the Transcriptional Activity of the Mineralocorticoid Receptor.

Lee HA, Song MJ, Seok YM, Kang SH, Kim SY, Kim I - PLoS ONE (2015)

HDAC4 interacts with both the MR and HDAC3.A, HEK293 cells were transfected with MR-HA and either HDAC4-Flag, HDAC5-Flag, or HDAC7-Flag plasmids. After 48 h, the cells were treated with Aldo (10 nM) for 30 min. Whole-cell lysates (WCLs) were prepared with non-denaturing lysis buffer. The MR was precipitated with an anti-HA antibody and MR-interacting HDACs were detected by western blotting with an anti-Flag antibody. HDAC4 and HDAC5 interacted with MR after Aldo treatment. B, HEK293 cells were transfected with HDAC3-Flag and HDAC4-Flag, HDAC5-Flag, or HDAC7-Flag plasmids. HDAC3 was immunoprecipitated with an antibody and HDAC3-interacting class II HDACs were detected by western blotting. HDAC4 interacted with HDAC3 after Aldo treatment. Aldo, aldosterone; HA, hemagglutinin; IP, immunoprecipitation; WB, western blotting.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4549324&req=5

pone.0136801.g001: HDAC4 interacts with both the MR and HDAC3.A, HEK293 cells were transfected with MR-HA and either HDAC4-Flag, HDAC5-Flag, or HDAC7-Flag plasmids. After 48 h, the cells were treated with Aldo (10 nM) for 30 min. Whole-cell lysates (WCLs) were prepared with non-denaturing lysis buffer. The MR was precipitated with an anti-HA antibody and MR-interacting HDACs were detected by western blotting with an anti-Flag antibody. HDAC4 and HDAC5 interacted with MR after Aldo treatment. B, HEK293 cells were transfected with HDAC3-Flag and HDAC4-Flag, HDAC5-Flag, or HDAC7-Flag plasmids. HDAC3 was immunoprecipitated with an antibody and HDAC3-interacting class II HDACs were detected by western blotting. HDAC4 interacted with HDAC3 after Aldo treatment. Aldo, aldosterone; HA, hemagglutinin; IP, immunoprecipitation; WB, western blotting.
Mentions: We previously reported that HDAC3 binds to MR. HDAC3 deacetylates MR, which increases its transcriptional activity [1]. Similar to our results, Mihaylova et al. showed that HDAC3 deacetylates the forkhead box O (FOXO) transcription factor, the interaction of which is mediated by class IIa HDACs [17]. To identify the role of class II HDACs in the interaction between MR and HDAC3, HA-tagged MR and Flag-tagged HDAC4, 5, and 7 were co-transfected into HEK293 cells and co-immunoprecipitation (co-IP) was performed to investigate the interaction between MR and class II HDACs. HDAC4 and HDAC5 interacted with MR stimulate by Aldosterone (Aldo) (Fig 1A). To identify a class II HDAC that simultaneously binds to the MR and HDAC3, HDAC3 and each of the class II HDACs were co-expressed in HEK293 cells. Treatment with Aldo promoted an interaction between HDAC3 and HDAC4 but not with the other class II HDACs (Fig 1B). Together, these results indicate that HDAC4 interacts with MR regulators and may also play an important role in MR function.

Bottom Line: The transcriptional activity of MR was significantly decreased by inhibitors of PKA (H89), PP1/2 (calyculin A), class I HDACs (MS-275), but not class II HDACs (MC1568).Interaction between MR and HDAC3 was significantly decreased by H89, calyculin A, and HDAC4 siRNA.A non-genomic effect of MR via PKA and PP1/2 induced nuclear translocation of HDAC4 to facilitate the interaction between MR and HDAC3.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmacology, School of Medicine, Kyungpook National University, Daegu, 41944, Republic of Korea; Cardiovascular Research Institute, School of Medicine, Kyungpook National University, Daegu, 41944, Republic of Korea; Cell and Matrix Research Institute, School of Medicine, Kyungpook National University, Daegu, 41944, Republic of Korea.

ABSTRACT
Histone deacetylases (HDACs) act as corepressors in gene transcription by altering the acetylation of histones, resulting in epigenetic gene silencing. We previously reported that HDAC3 acts as a coactivator of the mineralocorticoid receptor (MR). Although HDAC3 forms complexes with class II HDACs, their potential role in the transcriptional activity of MR is unclear. We hypothesized that HDAC4 of the class II family stimulates the transcriptional activity of MR. The expression of MR target genes was measured by quantitative real-time PCR. MR and RNA polymerase II recruitment to promoters of MR target genes was analyzed by chromatin immunoprecipitation. The association of MR with HDACs was investigated by co-immunoprecipitation. MR acetylation was determined with an anti-acetyl-lysine antibody after immunoprecipitation with an anti-MR antibody. Among the class II HDACs, HDAC4 interacted with both MR and HDAC3 after aldosterone stimulation. The nuclear translocation of HDAC4 was mediated by protein kinase A (PKA) and protein phosphatases (PP). The transcriptional activity of MR was significantly decreased by inhibitors of PKA (H89), PP1/2 (calyculin A), class I HDACs (MS-275), but not class II HDACs (MC1568). MR acetylation was increased by H89, calyculin A, and MS-275, but not by MC1568. Interaction between MR and HDAC3 was significantly decreased by H89, calyculin A, and HDAC4 siRNA. A non-genomic effect of MR via PKA and PP1/2 induced nuclear translocation of HDAC4 to facilitate the interaction between MR and HDAC3. Thus, we have uncovered a crucial role for a class II HDAC in the activation of MR-dependent transcription.

No MeSH data available.


Related in: MedlinePlus