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

本研究的目的是阐明附着胞发育的调控机制，附着胞是稻瘟病菌Magnaporthe grisea（和其他真菌病原体）成功感染寄主所需的一种特化细胞。以往的研究主要集中在附着胞发育突变体的基因表征和克隆上。这些数据已用于创建附着胞形成模型，以供进一步测试。在这个项目中，将利用迄今为止的进展以及克莱姆森大学基因组学研究所可用的新资源。现在有可能分析许多基因在各种发育条件下的表达。通过差异杂交分析，将尽可能多地确定与附着信息形成有关的基因。各种附着胞缺陷突变体的基因表达将被用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，为了开始定义调节附着胞形成的蛋白质的相互作用，将评估基因产物的相互作用。目标包括:1。确定与附着胞形成调控和发育有关的基因。该目标包括完成正在进行的研究，特别是APP1克隆和通过插入诱变标记的基因鉴定。在新的研究中，参与附着信息形成的基因将通过高密度过滤器上排列的cdna文库的差异杂交来鉴定。鉴定出的cdna将被测序并存入公共数据库。通过基因破坏评价附着胞基因的作用。通过利用2-混合系统，开始定义调控压胞形成的调控网络。本研究结果不仅具有理论意义，而且具有实际意义。对感染发生的必要基因和途径的鉴定可以用于创建基于合理机制的抑制剂。

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.