Dimerization of Floral Organ Identity Proteins in Arabidopsis

相当多的证据支持这样的假设，即不同开花植物物种中控制花发育的基因产物和机制在很大程度上是保守的。许多植物发育控制蛋白含有一个保守的DNA结合域，称为MADS结构域。拟南芥MADS家族由30多个基因组成。MADS蛋白参与植物发育的不同方面，包括开花时间控制、分生组织规格、花器官鉴定和果实发育。尽管这些MADS蛋白在植物发育中很重要，但人们对这些蛋白的功能和相互作用知之甚少。本研究的重点是拟南芥中MADS蛋白APETALA3(AP3)和pI(PI)的花器官识别。AP3和PI在拟南芥花瓣和雄蕊发育过程中起着重要的调控作用。所有的MADS蛋白都以异源二聚体或同源二聚体的形式与DNA结合。拟南芥中的大多数MADS蛋白都能与多种配对蛋白形成同源二聚体和异源二聚体。相比之下，AP3和PI不能均二聚并形成专有的异二聚体。AP3/PI异源二聚体是高度特异的，在进化上是保守的，远亲植物金鱼草的AP3和PI同源物之间的相似相互作用证明了这一点。本项目专注于鉴定AP3和PI蛋白中介导AP3和PI之间高度特异的异二聚体相互作用的氨基酸和亚结构域；这将通过定点突变和利用“反向”酵母双杂交方法来完成。通过这项研究，我们希望能够明确这类重要的植物发育调节因子的二聚化规律。这项研究的第二个重点是PI的一个不寻常的等位基因，称为pi-5。与所有其他ap3和pi突变体不同，pi-5-只在花的一个轮生中表现出表型缺陷。PI-5蛋白与野生型AP3蛋白之间的相互作用存在缺陷。用pi-5进行的遗传实验表明，轮状特异因子调节pi-5的活性。利用遗传和分子方法，将分离和表征影响花器官特性的轮生特有因子。

Considerable evidence supports the hypothesis that the gene products and mechanisms that control flower development in different flowering plant species are largely conserved. Many plant developmental control proteins contain a conserved DNA binding domain called the MADS domain. The MADS family in Arabidopsis consists of more than 30 genes. MADS proteins are involved in diverse aspects of plant development including flowering time control, meristem specification, floral organ identity, and fruit development. Despite the importance of these MADS proteins in plant development, little is known about how these proteins function and interact. This proposal focuses on the floral organ identity MADS proteins APETALA3 (AP3) and PISTILLATA (PI) in Arabidopsis thaliana. AP3 and PI function to direct the development of petal and stamens in the Arabidopsis flower. All MADS proteins bind to DNA either as heterodimers or homodimers. Most MADS proteins in Arabidopsis are capable of forming both homodimers and heterodimers with a variety of partner proteins. AP3 and PI, by contrast, are unable to homodimerize and form an obligate heterodimer. The AP3/PI heterodimer is highly specific and is evolutionarily conserved as evidenced by a similar interaction between the AP3 and PI homologs in the distantly related plant species Antirrhinum majus. This project is focused on identifying the amino acids and subdomains in the AP3 and PI proteins that mediate the highly specific heterodimer interaction between AP3 and PI; this will be done both by site-specific mutagenesis and by utilizing "reverse" yeast two-hybrid approaches. Through this research, we hope to be able to define the rules for dimerization of this importance class of plant developmental regulators. A second focus of this research centers on an unusual allele of PI called pi-5. Unlike all other ap3 and pi mutants, pi-5- exhibits phenotypic defects only in a single whorl of the flower. The PI-5 protein exhibits defects in protein-protein interaction with a wild-type AP3 partner protein. Genetic experiments with pi-5 suggest that whorl-specific factors modulate the activity of PI-5. Using both genetic and molecular approaches, whorl-specific factors that affect floral organ identity will be isolated and characterized.