The Role of START Lipid/Sterol Binding Domains in Homeodomain Transcription Factors of Plants

甾醇是一种不溶于水的小分子，在动物和植物的生长和发育中起着关键作用。植物分子生物学领域的研究揭示了甾醇衍生的油菜素甾醇的重要性，这是迄今为止唯一从植物中鉴定出的类固醇激素。除了油菜素甾醇，植物中还含有多种甾醇化合物，其生物学功能尚不清楚。施里克实验室的长期目标是鉴定和表征植物中假定的甾醇信号通路的组分。候选固醇结合蛋白包括具有类固醇生成急性调节（星星）相关脂质转移（START）脂质/固醇结合结构域的同源域（HD）转录因子家族。START结构域以哺乳动物胆固醇结合星星蛋白命名，在真核生物的多种蛋白质中发现，但仅在少数动物中进行了研究。引人注目的是，START结构域与HD DNA结合基序的组合是植物所独有的。该项目解决了HD转录因子中发现的START结构域与脂质/固醇配体结合，并且这种相互作用控制发育过程中的基因表达的假设。来自拟南芥、玉米和水稻的HD-START家族的几个成员在细胞分化中具有关键作用和/或表现出层特异性表达模式。利用拟南芥分生组织层1（ATML 1）和GLABRA 2（GL 2）基因作为模型，本项目的目的将是检查START结构域作为脂质/甾醇结合基序的假定作用。潜在的配体将在一种新的体内脂质/甾醇结合试验中利用酵母酿酒酵母进行实验鉴定。在该测定中，START结构域表达为合成转录因子的一部分，该合成转录因子响应于配体结合而激活报告基因。将实施该测定以测试脂质/固醇化合物的化学文库与来自拟南芥HD-START转录因子的候选START结构域的结合，并且另外将用于所选START结构域的突变分析。该项目的完成将为植物START结构域在配体结合中的假定功能提供概念框架，可能揭示植物中脂质/甾醇代谢与细胞分化之间的联系。此外，这项工作将导致在理解HD-START转录因子在植物发育中的作用方面取得重大进展，并将导致对脂质/固醇信号传导机制的重要见解。这项研究将通过对研究生和本科生的多学科教育产生更广泛的影响，从这项研究中获得的信息将有助于改进植物源物质，以用于生物技术的实际应用。

Sterols are small water-insoluble molecules that play critical roles in growth and development of both animals and plants. Ongoing research in the field of plant molecular biology has revealed the importance of the sterol-derived brassinosteroids, which are the only steroid hormones identified from plants thus far. In addition to brassinosteroids, plants contain a wide variety of sterol compounds whose biological functions are not understood. The long-term goal of the Schrick lab is to identify and characterize the components of putative sterol signaling pathways in plants. Candidate sterol-binding proteins include a family of homeodomain (HD) transcription factors having Steroidogenic Acute Regulatory (StAR)-related lipid transfer (START) lipid/sterol-binding domains. START domains, named after the mammalian cholesterol-binding StAR protein, are found in a variety of proteins across the eukaryotes, but have been studied in only a few cases from animals. Strikingly, START domains in combination with the HD DNA-binding motif are unique to plants. This project addresses the hypothesis that the START domain found in HD transcription factors binds to lipid/sterol ligands and that this interaction controls gene expression during development. Several members of the HD-START family from Arabidopsis, maize, and rice, have crucial roles in cell differentiation and/or exhibit layer-specific expression patterns. Using the Arabidopsis thaliana Meristem Layer 1 (ATML1) and GLABRA2 (GL2) genes as models, the aim of this project will be to examine the putative role of START domains as lipid/sterol binding motifs. Potential ligands will be experimentally identified in a novel in vivo lipid/sterol-binding assay utilizing the yeast Saccharomyces cerevisiae. In the assay the START domain is expressed as part of a synthetic transcription factor that activates a reporter gene in response to ligand binding. This assay will be implemented to test a chemical library of lipid/sterol compounds for binding to candidate START domains from Arabidopsis HD-START transcription factors, and additionally will be used for mutational analysis of selected START domains. Completion of this project will provide a conceptual framework for the presumed function of plant START domains in ligand binding, potentially revealing a link between lipid/sterol metabolism and cell differentiation in plants. Moreover, this work will result in significant advances in understanding of the role of HD-START transcription factors in plant development, and will lead to important insights underlying lipid/sterol signaling mechanisms. This research will have broader impacts through multidisciplinary education of graduate students as well as undergraduates, and the information gained from this research will facilitate improvements in plant-derived substances for practical applications in biotechnology.