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. 2017 Apr 21;7:1042. doi: 10.1038/s41598-017-01183-8

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© The Author(s) 2017

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Chromatin modification status of the 5’flanking region of GnIH gene in hypothyroidic and hyperthyroidic female mice. (a) Schematic diagram of the promoter region of GnIH gene. Putative TREs predicted by NHR scan (▼) and AliBaba 2.1 () are indicated at their locations from ORF. (b–d) ChIP-qPCR was performed to scan TR occupancy and chromatin modifications across the GnIH promoter region. Chromatin samples from control, PTU-induced hypothyroidic or T₄-induced hyperthyroidic female mice were precipitated using specific antibodies for TR (b), acetylation of histone H3 (H3Ac, (c)) and histone H3-Lysine 9 tri-methylation (H3K9me3, (d). Normal rabbit IgG was used as a negative control for the specificity of immunoprecipitation. TR occupancy in (b) is expressed as relative enrichment with normal IgG signal. Levels of histone modification in (c,d) are normalized to total histone H3 and expressed as fold increase relative to control. Data represent mean ± SEM ((b), n = 3 per group; n = 1 is 5 hypothalamic samples; (c,d) n = 6 per group). Letters indicate significant differences, p < 0.05 by two-way ANOVA Bonferroni post-test. (c) H3Ac, ANOVA summary; interaction, F _8,75 = 2.809, p = 0.0089; effect for promoter region, F _4,75 = 5.005, p = 0.0012; effect for thyroid status, F _2,75 = 52.48, p < 0.001. (d) H3K9me3, ANOVA summary; interaction, F ₈,₇₅ = 1.628, p = 0.1312; effect for promoter region, F ₄,₇₅ = 4.488, p = 0.0026; effect for thyroid status, F ₂,₇₅ = 95.46, p < 0.001.

Inline graphic — Chromatin modification status of the 5’flanking region of GnIH gene in hypothyroidic and hyperthyroidic female mice. (a) Schematic diagram of the promoter region of GnIH gene. Putative TREs predicted by NHR scan (▼) and AliBaba 2.1 () are indicated at their locations from ORF. (b–d) ChIP-qPCR was performed to scan TR occupancy and chromatin modifications across the GnIH promoter region. Chromatin samples from control, PTU-induced hypothyroidic or T₄-induced hyperthyroidic female mice were precipitated using specific antibodies for TR (b), acetylation of histone H3 (H3Ac, (c)) and histone H3-Lysine 9 tri-methylation (H3K9me3, (d). Normal rabbit IgG was used as a negative control for the specificity of immunoprecipitation. TR occupancy in (b) is expressed as relative enrichment with normal IgG signal. Levels of histone modification in (c,d) are normalized to total histone H3 and expressed as fold increase relative to control. Data represent mean ± SEM ((b), n = 3 per group; n = 1 is 5 hypothalamic samples; (c,d) n = 6 per group). Letters indicate significant differences, p < 0.05 by two-way ANOVA Bonferroni post-test. (c) H3Ac, ANOVA summary; interaction, F _8,75 = 2.809, p = 0.0089; effect for promoter region, F _4,75 = 5.005, p = 0.0012; effect for thyroid status, F _2,75 = 52.48, p < 0.001. (d) H3K9me3, ANOVA summary; interaction, F ₈,₇₅ = 1.628, p = 0.1312; effect for promoter region, F ₄,₇₅ = 4.488, p = 0.0026; effect for thyroid status, F ₂,₇₅ = 95.46, p < 0.001.