The Ccr4-Not Complex: Bifunctional roles in Repression of Steroid Receptor-Mediated Transcription and mRNA Decay?

细胞可以通过改变使用的指令来对激素等环境因素做出反应。这些指令或基因包含在可遗传的物质--DNA中。当基因被活跃表达时，在某些细胞中，但不是在所有细胞中，通过转录产生mRNA拷贝。在一个给定的时间，只有一部分基因通过一个高度调控和复杂的过程表达，这个过程需要数千种不同的蛋白质因素。参与转录的因子的确切数量尚不清楚，它们发挥作用的机制通常也不清楚。因为不同的细胞包含不同的转录因子，所以人体内的每种细胞类型都表达了一种特有的基因模式。对转录的激活同样重要的是基因的关闭。这可以通过不同的方式来实现。例如，因子可以与DNA结合并抑制转录。或者，因子可以通过破坏基因的mRNA拷贝来使基因表达失活。CCR4-NOT因子可以通过两种方式潜在地影响基因的表达：通过抑制转录和通过破坏mRNA。最近，我们发现该因子可以与雌激素受体相互作用。雌激素受体是一种转录因子，介导类固醇激素雌激素的细胞效应。激素与雌激素受体的结合会引起受体的结构变化，从而导致受体的激活。结果，数百个基因的表达发生了变化。一些基因被激活，并被活跃地转录，以响应激素，例如几个促进细胞分裂的基因。作为回应，其他基因被关闭并受到抑制，包括抑制细胞分裂的基因。这项拟议的工作将调查CCR4-NOT将如何影响细胞基因表达对雌激素的反应。它将通过专门设计缺乏CCR4亚单位的细胞来做到这一点--不是。利用DNA微阵列技术，将在一个单独的实验中确定3万个基因的表达，这是人类的大多数基因。使用正常细胞和缺乏CCR4-NOT功能的细胞获得的结果大型数据集将使用生物信息学进行比较。这将允许确定CCR4-NOT在细胞对雌激素的反应中有多重要。此外，这项拟议的工作将确定CCR4-NOT在多大程度上利用其抑制转录和破坏mRNA的能力来影响基因表达对雌激素的反应。这将在缺乏转录抑制功能或CCR4-NOT的mRNA破坏功能的细胞系中完成。最后，由于已知雌激素可以刺激细胞增殖，它将评估CCR4-NOT如何对细胞分裂产生影响。这些结果将增加我们对细胞中分子机制的基本理解。此外，它们可能会引起制药行业的兴趣，因为与天然激素雌激素相比，有几种药物可以与雌激素受体结合并以不同的方式影响其活性。这些药物成功地用于治疗骨质疏松症、严重的绝经后症状和癌症，但对有效性和安全性的担忧依然存在。更好地了解雌激素受体的工作原理可能最终导致更安全的药物的开发。

Cells can respond to environmental factors such as hormones by changing the instructions that are used. These instructions, or genes, are contained in the heritable material, the DNA. When genes are actively expressed, mRNA copies are produced by transcription in some, but not all cells. At a given time, only a sub-set of genes is expressed by a highly regulated and complex process that requires thousands of different protein factors. The precise number of factors involved in transcription is unknown, and the mechanism by which they work is often unclear. Because different cells contain different transcription factors, a characteristic pattern of genes is expressed for each cell type in the human body. Equally important to the activation of transcription, is the switching off of genes. This can be achieved in different manners. For example, factors can bind to DNA and repress transcription. Alternatively, factors can inactivate gene expression by destroying the mRNA copies of genes. The Ccr4-Not factor can potentially influence gene expression in both ways: by repressing transcription and by destruction of mRNA. Recently, we showed that this factor can interact with estrogen receptor. Estrogen receptor is a transcription factor that mediates the cellular effects of the steroid hormone estrogen. Binding of the hormone to estrogen receptor causes a structural change of the receptor which leads to activation of the receptor. As a result, the expression of several hundreds of genes is altered. Some genes are switched on and are actively transcribed in response to the hormone, such as several genes that promote cell division. Other genes are switched off in response and become repressed, including genes that inhibit cell division. The proposed work will investigate how Ccr4-Not will influence cellular gene expression in response to estrogen. It will do so by specifically engineer cells that lack subunits of Ccr4-Not. Using DNA microarray technology, the expression of >30,000 genes, the majority of human genes, will be determined in a single experiment. The resulting large data sets obtained using normal cells and cells lacking functional Ccr4-Not will be compared using bioinformatics. This will allow to define how important Ccr4-Not is in the cellular response to estrogen. In addition, the proposed work will determine to what extent Ccr4-Not uses its capacities to inhibit transcription and destroy mRNA to influence gene expression in response to estrogens. This will be done in cell lines that lack either the transcriptional repression function, or the mRNA destruction function of Ccr4-Not. Finally, because it is known that estrogens can stimulate cell proliferation, it will assess how Ccr4-Not can have an effect on cell division. The results will increase our basic understanding of the molecular machinery in cells. Furthermore, they may be of interest to the pharmaceutical industry, because there are several drugs that can bind to estrogen receptor and influence its activity in different ways as compared to the natural hormone estrogen. These drugs are successfully used for the treatment of osteoporosis, severe post-menopausal symptoms and cancer, but concerns about efficacy and safety remain. A better understanding of the workings of estrogen receptor may eventually result in the development of safer drugs.