Expression and Functional Analysis of Genes Regulating Appressorium Formation in Magnaporthe Grisea

这项研究的目的是阐明附着胞的发育调控机制，附着胞是稻瘟病菌(和其他真菌病原菌)成功侵染寄主所需的一种特殊细胞。以往的研究主要集中在附着胞发育突变体基因的鉴定和克隆上。这些数据已经被用来创建附着胞形成的模型，用于进一步的测试。在这个项目中，将利用克莱姆森大学基因组研究所的最新进展和可用的新资源。现在可以分析许多基因在各种发育条件下的表达。尽可能多的参与附着体形成的基因将通过差异杂交分析来鉴定。将利用各种附着胞缺失突变体的基因表达来鉴定潜在的关键基因。这些基因的功能将通过基因破坏来研究。最后，为了确定调控附着胞形成的蛋白质之间的相互作用，将对基因产物的相互作用进行评估。目的包括：1.寻找与附着胞形成调控和发育有关的基因。这一目标包括完成正在进行的研究，特别是APP1的克隆和识别插入突变标记的基因。在新的研究中，涉及附着体形成的基因将通过排列在高密度过滤器上的cDNA文库的差异杂交来识别。已识别的DNA将被测序并存放在公共数据库中。通过基因干扰评价附着胞基因的作用。通过利用双杂交系统，开始定义控制附着胞形成的调控网络。本研究结果具有实际意义和概念意义。识别感染发生所必需的基因和途径可以被用来创造合理的基于机制的抑制剂。

The purpose of the proposed research is to elucidate the mechanismsregulating the development of the appressorium, a specialized cell requiredby the rice blast fungus Magnaporthe grisea (and other fungal pathogens)for successful host infection. Previous research focused on characterizingand cloning genes from appressorium developmental mutants. These data havebeen used to create a model of appressorium formation for further testing.In this project, advantage will be taken of progress to date as well as newresources available at the Clemson University Genomics Institute. It isnow possible to analyze the expression of many genes under a variety ofdevelopmental conditions. As many of those genes involved in appressoriumformation as possible will be identified through differential hybridizationanalysis. Gene expression in various appressorium deficient mutants willbe exploited to identify potentially key genes. The function of thesegenes will be investigated by gene disruption. Finally, to begin to definethe interplay of proteins regulating appressorium formation the interactionof gene products will be evaluated. Objectives include:1. Identify genes involved in the regulation and development ofappressorium formation. This objective covers the completion of ongoingresearch, specifically APP1 cloning and identifying genes tagged byinsertional mutagenesis. In new research, genes involved in appressoriumformation will be identified by differential hybridization of cDNAlibraries arrayed on high-density filters. Identified cDNAs will besequenced and deposited in public databases.2. Evaluate role of appressorium genes through gene disruption.3. Begin to define the regulatory networks governing appressorium formationby exploiting the 2-hybrid system.Results from this investigation have practical as well as conceptualsignificance. The identification of genes and pathways essential forinfection to occur can be exploited to create rational mechanism basedinhibitors.