Cytokinin Metabolic Genes and Enzymes

细胞分裂素是介导细胞分裂和分化的植物激素。最重要的天然细胞分裂素是玉米素，一种具有高生物活性的化合物。在植物组织中发现了各种各样的玉米素代谢产物，其中一些是活跃的，而另一些则不是，因此，了解玉米素代谢是如何调节的至关重要。这个项目的长期目标是识别和描述施加这种控制的元素。先前的研究已经分离出了玉米蛋白特异性酶，包括o -糖基转移酶，它可以将一个糖基团转移到玉米蛋白上，并克隆了豆类、拟南芥和玉米中与这些酶相对应的基因家族。o糖基化玉米蛋白广泛存在于植物组织中，作为细胞分裂素的一种无活性、可逆（通过水解）的储存形式。与玉米素相比，o -糖基化的玉米素耐酶分解。因此，糖基转移酶基因的活性可能对活性细胞分裂素的水平产生重大影响，从而也影响植物的发育。另一种重要的酶是玉米蛋白还原酶，它将玉米蛋白转化为二氢玉米蛋白，这是一种比玉米蛋白稳定得多的活性细胞分裂素。虽然二氢玉米素存在于所有植物中，并且还原酶已从植物组织中分离出来，但与这种酶相对应的基因尚未被鉴定出来。建议的研究重点是玉米素o -葡萄糖基转移酶和还原酶，包括一系列分子，酶，生理和发育研究。具体目标是：(1)研究玉米素o糖基化在种子萌发、运输和衰老中的作用；2)测定玉米素o -葡萄糖基转移酶在拟南芥中的表达和细胞定位；3)鉴定对底物特异性至关重要的酶上的氨基酸序列；4)分离编码玉米蛋白还原酶的基因。至于更广泛的影响，该项目涉及与捷克科学院的R. Vankova博士的国际合作。该项目将研究与教育相结合，重点是本科培训，其中包括让学生在布拉格R. Vankova实验室获得国际经验。拟议的研究还包括与私营企业合作，这将进一步扩大研究的影响，并促进新发现的流动。

Cytokinins are plant hormones mediating cell division and differentiation. The most important naturally-occurring cytokinin is zeatin, a compound with high biological activity. Various zeatin metabolites are found in plant tissues, some of which are active while others are not, and therefore, it is crucial to understand how zeatin metabolism is regulated. The long-term goal of this project is to identify and characterize elements exerting such controls. Previous research has led to isolation of zeatin-specific enzymes, including O-glycosyltransferases, which transfer a sugar group to zeatin, and cloning of a family of genes corresponding to these enzymes from beans, Arabidopsis, and maize. O-glucosylated zeatin occurs widely in plant tissues where it functions as an inactive, reversible (via hydrolysis) storage form of cytokinin. In contrast to zeatin, O-glucosylated zeatin is resistant to enzymatic breakdown. Consequently, the activities of the glucosyltransferase genes are likely to have significant impact on the level of active cytokinins and, as a consequence, also on plant development. Another important enzyme is zeatin reductase, which converts zeatin to dihydrozeatin, an active cytokinin that is much more stable than zeatin. Although dihydrozeatin occurs in all plants and the reductase has been isolated from plant tissue, genes corresponding to this type of enzyme have not yet been identified. The proposed research focuses on zeatin O-glucosyltransferases and reductases and includes a range of molecular, enzymatic, physiological, and developmental investigations. The specific objectives are to: (1) examine the role of zeatin O-glucosylation in seed germination, transport, and senescence; 2) determine the expression and cytolocalization of zeatin O-glucosyltransferases in Arabidopsis; 3) identify amino acid sequences on the enzymes critical to substrate specificity; and 4) isolate genes encoding zeatin reductases. As to broader impacts, the project involves international collaboration, with Dr. R. Vankova of the Czech Academy of Sciences. The project integrates research with education, with an emphasis on undergraduate training that includes an international experience for students in the laboratory of R. Vankova in Prague. The proposed research also includes collaboration with private industry, which further broadens the impact of the research and enhances the flow of new findings.