Johnston Jonathan D, Schuster Carole, Barrett Perry, Hazlerigg David G
School of Biomedical and Molecular Sciences, University of Surrey, Guildford GU2 7XH, UK.
Mol Cell Endocrinol. 2007 Mar 30;268(1-2):59-66. doi: 10.1016/j.mce.2007.01.015. Epub 2007 Feb 1.
Pineal secretion of melatonin provides a neuroendocrine representation of the light-dark cycle, which is used to synchronise daily and annual rhythms of physiology and behaviour. In mammals, melatonin primarily acts through MT(1) melatonin receptors that exhibit a highly restricted tissue distribution. Expression of MT(1) receptors is subject to developmental and circadian control, which likely modulates the physiological actions of melatonin. To investigate the mechanisms controlling MT(1) expression we cloned the proximal 1.5kb region of the ovine MT(1) promoter. Sequence analysis revealed putative cis-elements for transcription factors involved in pituitary development, namely Pitx-1 and Egr-1, and multiple putative E-boxes, which are involved in both circadian and developmental gene regulation. Nuclear protein from ovine pars tuberalis (PT) cells, a site of high endogenous MT(1) expression, stimulated gene expression from a MT(1) expression construct, indicating the presence of a functional promoter. Pitx-1 was strongly expressed in the ovine PT and stimulated MT(1) promoter activity in transfection assays. Co-transfection with Egr-1 induced promoter-specific effects: Pitx-1-stimulated MT(1) activity was inhibited, whereas betaLH promoter activity was enhanced. In addition to Pitx-1 the circadian clock genes Clock and Bmal1 were also expressed in the PT. However, despite multiple putative E-boxes in the MT(1) promoter, transfected Clock and Bmal1 were unable to regulate either basal or Pitx-1-stimulated MT(1) promoter activity. The current data, in conjunction with our previous study of the rat MT(1) promoter, suggests a general model in which melatonin receptor expression in the mammalian pituitary is determined by the developmentally changing balance between stimulatory and inhibitory transcription factors. Furthermore, our data suggest that circadian variation in MT(1) gene expression does not depend upon the direct action of circadian clock genes on E-box cis-elements.
褪黑素的松果体分泌提供了明暗周期的神经内分泌表征，该表征用于同步生理和行为的每日及年度节律。在哺乳动物中，褪黑素主要通过MT(1)褪黑素受体发挥作用，这些受体呈现出高度受限的组织分布。MT(1)受体的表达受发育和昼夜节律控制，这可能会调节褪黑素的生理作用。为了研究控制MT(1)表达的机制，我们克隆了绵羊MT(1)启动子的近端1.5kb区域。序列分析揭示了参与垂体发育的转录因子的假定顺式元件，即Pitx-1和Egr-1，以及多个假定的E盒，它们参与昼夜节律和发育基因调控。来自绵羊结节部（PT）细胞（内源性MT(1)高表达的部位）的核蛋白刺激了MT(1)表达构建体的基因表达，表明存在功能性启动子。Pitx-1在绵羊PT中强烈表达，并在转染实验中刺激MT(1)启动子活性。与Egr-1共转染诱导了启动子特异性效应：Pitx-1刺激的MT(1)活性受到抑制，而βLH启动子活性增强。除了Pitx-1，昼夜节律时钟基因Clock和Bmal1也在PT中表达。然而，尽管MT(1)启动子中有多个假定的E盒，但转染的Clock和Bmal1无法调节基础或Pitx-1刺激的MT(1)启动子活性。目前的数据，结合我们之前对大鼠MT(1)启动子的研究，提出了一个通用模型，即哺乳动物垂体中褪黑素受体的表达由刺激和抑制转录因子之间发育变化的平衡决定。此外，我们的数据表明MT(1)基因表达的昼夜变化并不依赖于昼夜节律时钟基因对E盒顺式元件的直接作用。
