Aichinger C, Hansson K, Eichhorn H, Lessing F, Mannhaupt G, Mewes W, Kahmann R
Institute for Genetics and Microbiology, Ludwig-Maximilians University Munich, Maria-Ward-Str.1a, 80638 Munich, Germany.
Mol Genet Genomics. 2003 Dec;270(4):303-14. doi: 10.1007/s00438-003-0926-z. Epub 2003 Oct 2.
To identify plant-induced genes in the maize pathogenic fungus Ustilago maydis we have developed a genetic screen that combines REMI (restriction enzyme mediated integration) mutagenesis with enhancer trapping using the gene for Green Fluorescent Protein (GFP) as vital reporter. Of 2,350 insertion mutants isolated, three were shown to express GFP only after the fungus had come into contact with the host maize plant. One of the genes tagged was mfa1, which encodes the pheromone precursor, while the second gene, pig2, codes for a product that showed similarity to protein disulfide isomerase. The third integration event had occurred in a locus which we designated the p -locus. This locus contains 11 genes in a 24-kb stretch. Of these, pig3, 4, 5, 6 and 7 show a plant-regulated expression pattern, while the other genes found at the locus (designated npi) do not. Of the plant-regulated genes only two were found to be similar to database entries: the pig4 product is related to membrane transporters of the major facilitator family, while the pig6 protein shows similarity to multidrug transporters. Detailed expression studies revealed that the five plant-regulated genes at the p -locus differ in their expression profiles. Mutants deleted for each of them showed no apparent phenotype, while the npi1 gene appeared to be essential. A viable deletion encompassing the entire p -locus could be generated when npi1 function was provided ectopically. This deletion mutant also showed no obvious alteration in virulence.
为了鉴定玉米致病真菌玉米黑粉菌中植物诱导基因，我们开发了一种遗传筛选方法，该方法将REMI（限制性内切酶介导整合）诱变与增强子捕获相结合，使用绿色荧光蛋白（GFP）基因作为重要报告基因。在分离出的2350个插入突变体中，有三个被证明仅在真菌与宿主玉米植株接触后才表达GFP。其中一个被标记的基因是mfa1，它编码信息素前体，而第二个基因pig2编码的产物与蛋白质二硫键异构酶相似。第三次整合事件发生在一个我们命名为p位点的基因座上。这个基因座在24kb的区域内包含11个基因。其中，pig3、4、5、6和7呈现出植物调控的表达模式，而在该基因座上发现的其他基因（命名为npi）则没有。在植物调控基因中，只有两个被发现与数据库条目相似：pig4产物与主要易化子家族的膜转运蛋白有关，而pig6蛋白与多药转运蛋白相似。详细的表达研究表明，p位点上的五个植物调控基因在表达谱上存在差异。针对每个基因缺失的突变体没有表现出明显的表型，而npi1基因似乎是必需的。当异位提供npi1功能时，可以产生一个包含整个p位点的可行缺失。这个缺失突变体在毒力方面也没有表现出明显的改变。
