低剂量染料木黄酮通过丝裂原活化蛋白激酶和丝裂原及应激激活激酶1非基因组信号通路对转录因子启动子区域进行表观遗传调控。
Epigenetic regulation of transcription factor promoter regions by low-dose genistein through mitogen-activated protein kinase and mitogen-and-stress activated kinase 1 nongenomic signaling.
作者信息
Yu Linda, Ham Kyle, Gao Xiaohua, Castro Lysandra, Yan Yitang, Kissling Grace E, Tucker Charles J, Flagler Norris, Dong Ray, Archer Trevor K, Dixon Darlene
机构信息
Molecular Pathogenesis Group, National Toxicology Program (NTP) Laboratory, Division of the NTP (DNTP), National Institute of Environmental Health Sciences (NIEHS), National Institutes of Health (NIH), U.S. Department of Health and Human Services (HHS), Research Triangle Park, North Carolina, 27709, USA.
Biostatistics and Computational Biology Branch, Division of Intramural Research (DIR), NIEHS, NIH, HHS, Research Triangle Park, North Carolina, 27709, USA.
出版信息
Cell Commun Signal. 2016 Aug 31;14(1):18. doi: 10.1186/s12964-016-0141-2.
BACKGROUND
The phytoestrogen, genistein at low doses nongenomically activates mitogen-activated protein kinase p44/42 (MAPKp44/42) via estrogen receptor alpha (ERα) leading to proliferation of human uterine leiomyoma cells. In this study, we evaluated if MAPKp44/42 could activate downstream effectors such as mitogen- and stress-activated protein kinase 1 (MSK1), which could then epigenetically modify histone H3 by phosphorylation following a low dose (1 μg/ml) of genistein.
RESULTS
Using hormone-responsive immortalized human uterine leiomyoma (ht-UtLM) cells, we found that genistein activated MAPKp44/42 and MSK1, and also increased phosphorylation of histone H3 at serine10 (H3S10ph) in ht-UtLM cells. Colocalization of phosphorylated MSK1 and H3S10ph was evident by confocal microscopy in ht-UtLM cells (r = 0.8533). Phosphorylation of both MSK1and H3S10ph was abrogated by PD98059 (PD), a MEK1 kinase inhibitor, thereby supporting genistein's activation of MSK1 and Histone H3 was downstream of MAPKp44/42. In proliferative (estrogenic) phase human uterine fibroid tissues, phosphorylated MSK1 and H3S10ph showed increased immunoexpression compared to normal myometrial tissues, similar to results observed in in vitro studies following low-dose genistein administration. Real-time RT-PCR arrays showed induction of growth-related transcription factor genes, EGR1, Elk1, ID1, and MYB (cMyb) with confirmation by western blot, downstream of MAPK in response to low-dose genistein in ht-UtLM cells. Additionally, genistein induced associations of promoter regions of the above transcription factors with H3S10ph as evidenced by Chromatin Immunoprecipitation (ChIP) assays, which were inhibited by PD. Therefore, genistein epigenetically modified histone H3 by phosphorylation of serine 10, which was regulated by MSK1 and MAPK activation.
CONCLUSION
Histone H3 phosphorylation possibly represents a mechanism whereby increased transcriptional activation occurs following low-dose genistein exposure.
背景
植物雌激素染料木黄酮在低剂量时可通过雌激素受体α（ERα）非基因组地激活丝裂原活化蛋白激酶p44/42（MAPKp44/42），从而导致人子宫平滑肌瘤细胞增殖。在本研究中，我们评估了MAPKp44/42是否能激活下游效应器，如丝裂原和应激激活蛋白激酶1（MSK1），进而在低剂量（1μg/ml）染料木黄酮作用后通过磷酸化对组蛋白H3进行表观遗传修饰。
结果
使用激素反应性永生化人子宫平滑肌瘤（ht-UtLM）细胞，我们发现染料木黄酮激活了MAPKp44/42和MSK1，并且还增加了ht-UtLM细胞中组蛋白H3丝氨酸10位点（H3S10ph）的磷酸化。通过共聚焦显微镜观察，在ht-UtLM细胞中磷酸化的MSK1和H3S10ph明显共定位（r = 0.8533）。MEK1激酶抑制剂PD98059（PD）消除了MSK1和H3S10ph的磷酸化，从而支持染料木黄酮对MSK1的激活，并且组蛋白H3处于MAPKp44/42的下游。在增殖期（雌激素期）人子宫肌瘤组织中，与正常子宫肌层组织相比，磷酸化的MSK1和H3S10ph的免疫表达增加，这与低剂量染料木黄酮给药后的体外研究结果相似。实时RT-PCR阵列显示，在ht-UtLM细胞中，低剂量染料木黄酮作用后，生长相关转录因子基因EGR1、Elk1、ID1和MYB（cMyb）被诱导，western blot证实了这一点，这些基因处于MAPK下游。此外，染色质免疫沉淀（ChIP）分析表明，染料木黄酮诱导上述转录因子的启动子区域与H3S10ph结合，而PD可抑制这种结合。因此，染料木黄酮通过丝氨酸10的磷酸化对组蛋白H3进行表观遗传修饰，这一过程受MSK1和MAPK激活的调节。
结论
组蛋白H3磷酸化可能代表了低剂量染料木黄酮暴露后转录激活增加的一种机制。
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