Pathogenic specialization in phytopathogenic fungi producing host-specific toxins

在几种植物病原真菌中，宿主特异性毒素是决定宿主特异性致病性或毒力的关键因素。7个变种的互花alternnaria alternata产生宿主特异性毒素，并在不同的植物上引起严重的疾病。这些寄主特异性的形式已被指定为互生假蝇的致病型。为深入了解麻霉致病性专门化的分子基础，本项目重点克隆麻霉毒素生物合成所需基因。我们主要研究了日本梨、草莓和橘子三种致病型，它们分别产生AK-、AF-和act -毒素。这些毒素有一个共同的结构片段，9,10-环氧-8-羟基-9-甲基-十三烯酸。本研究所得结果总结如下。(1)分离了日本梨病原ak -毒素生物合成所需的基因簇，在约80kb的区域内发现了11个假定的开放阅读框（orf）。结构分析还发现该区域内有更多的转座子样序列。orf的功能分析鉴定出8个参与ak毒素生物合成的基因（AKT基因）。(2)通过DNA凝胶印迹分析，观察了AKT基因在7种病原菌和非病原菌中的同源性，并在草莓和柑橘病原菌中发现了6个同源性基因。(3)构建了草莓和橘子的基因组文库，并分离出AKT同源克隆。(4)分离到5个与AKT基因同源的基因（AFT基因）和3个新发现的orf。其中一个orf被鉴定为af毒素生物合成的特异性基因。我们还发现这些基因位于一个可有可无的1.1 mb染色体上。(5)分离到3个与AKT基因同源的基因（ACT基因）和3个新发现的orf。我们通过转化介导的靶向ACT基因（ACTR）产生了ACT毒素缺陷突变体。该突变体通过引入日本梨病原菌的同源基因（AKTR）可以恢复其产毒，显示出ACTR和AKTR的功能同一性。少

Host-specific toxins are critical determinants of host-specific pathogenicity or virulence in several plant pathogenic fungi. Seven variants of Alternaria alternata produce host-specific toxins and cause severe diseases on different plants. These host-specific forms have been designated as pathotypes of A.alternata. To understand the molecular basis of pathogenic specialization in A.alternata, this research project has focused on cloning the genes required for toxin biosynthesis of the pathogens. We have studied mainly three pathotypes, Japanese pear, strawberry and tangerine, which produce AK-, AF- and ACT-toxins, respectively. These toxins have a common structural moiety, 9,10-epoxy-8-hydroxy-9-methyl-decatrienoic acid. Summary of results obtained in this research is as follows.(1) We isolated the gene cluster required for AK-toxin biosynthesis of the Japanese pear pathotype and found 11 putative open reading frames (ORFs) within an about 80-kb region. Structural analysis also found … More several transposon-like sequences within the region. Functional analysis of the ORFs identified eight genes (AKT genes) involved in AK-toxin biosynthesis.(2) We observed distribution of the AKT gene homologues in seven pathotypes and nonpathogenic stains of A.alternata by DNA gel blot analysis and found homologues of six genes in the strawberry and tangerine pathotypes.(3) We constructed cosmid genomic libraries of the strawberry and tangerine pathotypes and isolated clones containing the AKT homologues.(4) We isolated five genes (AFT genes) homologous to AKT genes and newly found three ORFs. One of the ORFs was identified as a gene specific for AF-toxin biosynthesis. We also found that the genes are located on a dispensable 1.1-Mb chromosome.(5) We isolated three genes (ACT genes) homologous to AKT genes and newly found three ORFs. We produced ACT-toxin-deficient mutants by transformation-mediated targeting of an ACT gene (ACTR). Toxin production of the mutant could be restored by introduction of the homologous gene (AKTR) of the Japanese pear pathotype, showing the functional identity of ACTR and AKTR. Less