Department of Physiology, The University of Hong Kong, Hong Kong, China Department of Biochemistry, The University of Hong Kong, Hong Kong, China Nursing, School of Science and Technology, Open University of Hong Kong, Hong Kong, China.
J Pineal Res. 2013 Jan;54(1):69-79. doi: 10.1111/j.1600-079X.2012.01026.x. Epub 2012 Aug 1.
Our laboratory has recently demonstrated a melatonin MT1 receptor-mediated antiproliferative signaling mechanism in androgen receptor (AR)-positive prostate epithelial cells which involves up-regulation of p27(Kip1) through dual activation of Gα(s)/protein kinase A (PKA) and Gα(q)/protein kinase C (PKC) in parallel, and down-regulation of activated AR signaling via PKC stimulation. The aim of the present investigation was to identify the transcription factor that mediates melatonin's up-regulatory effect on p27(Kip1) in LNCaP and 22Rv1 prostate cancer cells. Deletion mapping and reporter assays of the p27(Kip1) promoter revealed that the putative melatonin-responsive transcription factor binds to a 116 base-pair region of the promoter sequence, which contains a potential nuclear factor kappa B (NF-κB) binding site. When the NF-κB binding site was abolished by site-directed mutagenesis, the stimulatory effect of melatonin on p27(Kip1) promoter activity was mitigated. Notably, melatonin inhibited the DNA binding of activated NF-κB via MT1 receptor-induced PKA and PKC stimulation. Furthermore, melatonin's up-regulatory effect on p27(Kip1) transcription and consequent cell antiproliferation were abrogated by NF-κB activator but mimicked by NF-κB inhibitor. The results indicate that inhibition of constitutively active NF-κB via melatonin MT1 receptor-induced dual activation of (Gα(s)) PKA and (Gα(q)) PKC can de-repress the p27(Kip1) promoter leading to transcriptional up-regulation of p27(Kip1). MT1 receptor-mediated inhibition of activated NF-κB signaling provides a novel mechanism supporting the use of melatonin in prostate cancer chemoprevention and therapy.
我们的实验室最近在雄激素受体 (AR) 阳性前列腺上皮细胞中证明了褪黑素 MT1 受体介导的抗增殖信号机制，该机制涉及通过 Gα(s)/蛋白激酶 A (PKA) 和 Gα(q)/蛋白激酶 C (PKC) 的双重激活平行上调 p27(Kip1)，并通过 PKC 刺激下调激活的 AR 信号。本研究的目的是确定介导褪黑素对 LNCaP 和 22Rv1 前列腺癌细胞中 p27(Kip1)上调作用的转录因子。p27(Kip1)启动子的缺失作图和报告基因分析表明，假定的褪黑素反应性转录因子结合到启动子序列的 116 个碱基对区域，其中包含一个潜在的核因子 kappa B (NF-κB) 结合位点。当 NF-κB 结合位点通过定点诱变被消除时，褪黑素对 p27(Kip1)启动子活性的刺激作用减弱。值得注意的是，褪黑素通过 MT1 受体诱导的 PKA 和 PKC 刺激抑制激活的 NF-κB 的 DNA 结合。此外，NF-κB 激活剂阻断了褪黑素对 p27(Kip1)转录的上调作用和随后的细胞增殖抑制作用，而 NF-κB 抑制剂则模拟了这一作用。结果表明，通过褪黑素 MT1 受体诱导的 (Gα(s)) PKA 和 (Gα(q)) PKC 的双重激活抑制固有活性的 NF-κB，可解除 p27(Kip1)启动子的抑制作用，导致 p27(Kip1)转录的上调。MT1 受体介导的抑制激活的 NF-κB 信号提供了支持褪黑素用于前列腺癌化学预防和治疗的新机制。
