PHOTOREGULATED GENE EXPRESSION

这一建议涉及到基因表达的机制 受光调节的。实验生物是植物拟南芥。 柳杉，以其在分子遗传学方面的优点而闻名 分析。拟议的研究既涉及对光的分析- 光调控基因的响应元件(LRE) 一类蛋白质分子(G-box结合)的表征 因子或GBF)与这些LRE的共同元素相结合。在……里面 此外，光照射改变GBF的机制 将会被检查。 高等真核生物的特征是基因系统对 对发展和环境刺激的复杂方式。一个组件 这种复杂性的是常见的基因家族成员 它们可以执行相关但不同的功能。一个这样的例子 复杂性是Fos/Jun转录因子家族的作用 这个bZIP蛋白家族在哺乳动物发育和 致癌作用。GBF家族也是bZIp蛋白和GBF4，与 Fos，与其他家族成员形成异二聚体，但未能 同质二聚化。这两个家族的特征都是家族内遗传 监管体系。有人建议利用分子遗传学的力量 拟南芥中探讨GBF家族特性的分析 蛋白质。 LRE的基本特征将通过研究 启动子-报告基因在转基因植物中的表达 瞬变化验系统。这些构造的表达式将为 与内源拟南芥基因的表达进行比较。大号 部分GBF蛋白存在于细胞质中。的性质 影响这些基因在细胞内分布的GBF序列 蛋白质将被测定。GBF的DNA结合特性如下 受光线影响。显然，这一修改的性质 涉及磷酸化的，将被检查：这项研究将包括 测定这个反应的动力学，光的性质 要求，以及修改的GBF序列。为了这些 研究，就像其他拟议的研究一样，单克隆抗体将 获得并用于区分单个GBF家庭成员。 转录系统以非常特殊的蛋白质-蛋白质为特征。 互动。为了进一步了解GBF家族的作用 光调控基因表达中的蛋白质，相互作用的蛋白质 对于这个bZIP蛋白家族，将通过使用 在酵母中进行基因筛选。将使用两种方法来评估 GBF家庭成员个人的职能。基于直接聚合酶链式反应的筛查 将被用于搜索含有GBF基因标签的拟南芥植物 通过T-DNA插入。此外，利用转基因植物表达 人们正在探索GBF-反义结构。初步结果来自 后一种方法表明存在一个复杂的家庭内部监管网络。 基于这些反义实验的结果，一种新的 提出了互补性制度，从而有可能探索 GBF家族成员的显著特征。

This proposal concerns the mechanism by which gene expression is regulated by light. The experimental organisms is the plant Arabidopsis thaliana, well known for its merits concerning molecular genetic analyses. The proposed studies concern both the analyses of light- response-elements (LREs) that characterize light-regulated genes and the characterization of a family of protein molecules (the G-box-binding factors or GBFs) that bind to a common element of these LREs. In addition, the mechanism by which irradiation with light modifies the GBFs will be examined. Higher eukaryotes are characterized by genetic systems that respond in complex ways to developmental and environmental stimuli. One component of this complexity is the common occurrence of gene families, the members of which may perform related but distinct functions. An example of such complexity is the Fos/Jun family of transcription factors and the role that this bZIP family of proteins plays in mammalian development and oncogenesis. The GBF family are also bZIp proteins and GBF4, as with Fos, forms heterodimers with the other family members but fails to homodimerize. Both families are characterized by intrafamily genetic regulatory systems. It is proposed to use the power of molecular genetic analyses in Arabidopsis to explore the properties of the GBF family of proteins. The essential features of LREs will be examined by studying the expression of promoter-reporter constructs in both transgenic plants and transient assay systems. The expression of these constructs will be compared with the expression of endogenous Arabidopsis genes. A large fraction of the GBF proteins are found in the cytoplasm. The nature of the GBF sequences that affect the intracellular distribution of these proteins will be determined. The DNA binding properties of GBF are influenced by light. The nature of this modification, apparently involving phosphorylation, will be examined: this study will include a determination of the kinetics of this reaction, the nature of the light requirements, and the GBF sequences that are modified. For these studies, as with other proposed studies, monoclonal antibodies will be obtained and used to distinguish the individual GBF family members. Transcription systems are characterized by very specific protein-protein interactions. In order to further understand the role of the GBF family of proteins in light-regulated gene expression, proteins that interact with this family of bZIP proteins will be selected via the use of a genetic screen in yeast. Two approaches will be used to evaluate the function of the individual GBF family members. A direct PCR-based screen will be used to search for Arabidopsis plants containing GBF genes tagged by T-DNA insertion. In addition, the use of transgenic plants expressing GBF-antisense constructs is being explored. Preliminary results from this latter approach indicate a complex intrafamily regulatory network. Based on the results from these antisense experiments, a novel complementation system is proposed whereby it may be possible to explore the distinguishing features of the GBF family members.