Analysis of Cytokinin Signaling in Arabidopsis

描述(申请人提供)：细胞分裂素是一组植物激素 它们是强大的有丝分裂原，在许多植物的发育过程中发挥作用 流程。对细胞分裂素作用的分子机制的理解 有助于更好地了解人体细胞增殖的调控 细胞。几乎没有关于细胞分裂素信号转导的确切信息。 高等植物。我们建议使用一种综合的方法来解决这个问题， 结合分子、遗传和生化方法来阐明 参与细胞分裂素作用的元素。这项提议的主要要点是 从我们分离的两个细胞分裂素主要反应基因IBC6和IBC7中， 它属于一个更大的拟南芥基因家族，类似于细菌 信令元素。我们将利用细胞分裂素对绿色荧光蛋白的特异性诱导 IBC6启动子-GFP转基因株系的荧光鉴定 细胞分裂素作用突变体通过识别不适当地展示 GFP表达。初步筛选表明，这种方法将是 并且这些突变体应该有助于剖析细胞分裂素的反应 路径。IBC6和IBC7的快速诱导，加上它们与 细菌信号元件，表明这些基因可能参与了 细胞分裂素信号。我们将测试这一假设和其他可能的函数 通过确定改变的ARR基因表达的效果， 基因中断和受调控的过度表达，对植物发育和 细胞分裂素反应性。我们还将鉴定和表征那些 与IBC6互动，作为识别潜在下游目标和 上游激活剂。我们建议鉴定顺式作用的DNA序列 负责IBC6的细胞分裂素反应作为一种开始 解开细胞分裂素调节基因表达的机制。一次 ，我们将使用该元素来确定其他潜在的细胞分裂素 反应基因和鉴定负责的DNA结合蛋白 此元素的功能。最后，我们将使用Affymetrix基因芯片来 在拟南芥中寻找新的细胞分裂素调控基因。

DESCRIPTION (provided by applicant): Cytokinins are a group of plant hormones that are powerful mitogens and which play a role in many plant developmental processes. An understanding of the molecular mechanisms by which cytokinins act could help to better understand the regulation of cell proliferation in human cells. Almost nothing definitive is known with regard to cytokinin signaling in higher plants. We propose to utilize an integrative approach to this problem, combining molecular, genetic and biochemical approaches to elucidate the elements involved in cytokinin action. The main thrusts of this proposal derive from our isolation of two cytokinin primary response genes, IBC6 and IBC7, which belong to a larger Arabidopsis gene family that is similar to bacterial signaling elements. We will exploit the cytokinin-specific induction of GFP fluorescence in an IBC6 promoter-GFP transgenic line to identify cytokinin-action mutants by identifying seedlings that inappropriately exhibit GFP expression. Preliminary screens indicate that this approach will be successful and that these mutants should help to dissect the cytokinin response pathway. The rapid induction of IBC6 and IBC7, coupled with their similarity to bacterial signaling elements, suggests that these genes may be involved in cytokinin signaling. We will test this hypothesis and other possible functions of this gene family by determining the effect of altered ARR gene expression, both gene disruption and regulated overexpression, on plant development and cytokinin responsiveness. We will also identify and characterize proteins that interact with IBC6 as a means of identifying potential downstream targets and upstream activators. We propose to identify the cis-acting DNA sequences responsible for the cytokinin-response of IBC6 as a means of starting to unravel the mechanisms by which cytokinins regulate gene expression. Once identified, we will use this element to identify additional potential cytokinin response genes and to identify DNA binding proteins responsible for the function of this element. Finally, we will use Affymetrix gene chips to identify novel cytokinin-regulated genes in Arabidopsis.