Claviceps paspali is used in the pharmaceutical industry for the production of ergot alkaloids. This fungus also biosynthesizes paspalitrems, indole diterpene (IDT) mycotoxins that cause significant economic losses in agriculture and represent safety concerns for ergot alkaloid manufacture. Here, we use Agrobacterium-mediated transformation to replace the idtP and the idtF genes in the IDT biosynthetic gene cluster of C. paspali with a selectable marker gene. We show that the ΔidtP knockout mutant produces paspaline, the first IDT intermediate of the pathway. The ΔidtF strain produces unprenylated IDTs such as paspalinine and paspaline. These experiments validate the function of idtP as the gene encoding the cytochrome P450 monooxygenase that oxidizes and demethylates paspaline to produce 13-desoxypaxilline, and that of idtF as the gene that encodes the α-prenyltransferase that prenylates paspalinine at the C20 or the C21 positions to yield paspalitrems A and C, respectively. In addition, we also show that axenic cultures of the wild type, the ΔidtP and the ΔidtF mutant C. paspali strains fail to produce an assembly of IDTs that are present in C. paspali-Paspalum spp. associations.

中文翻译：

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Caspiceps paspali吲哚二萜生物合成基因簇中idtF和idtP基因的功能表征。

Caspiceps paspali在制药工业中用于生产麦角生物碱。这种真菌还可以生物合成帕斯帕利姆，吲哚二萜（IDT）霉菌毒素，这会在农业上造成重大经济损失，并代表着麦角生物碱生产的安全性问题。在这里，我们使用农杆菌介导的转化，用可选择的标记基因替换了巴氏梭菌IDT生物合成基因簇中的idtP和idtF基因。我们表明，ΔidtP基因敲除突变体产生paspaline，该途径的第一个IDT中间体。ΔidtF菌株产生未异戊二烯化的IDT，例如帕斯帕林宁和帕斯帕林。这些实验验证了idtP作为编码细胞色素P450单加氧酶的基因的功能，该酶氧化和脱甲基帕斯帕林以产生13-脱氧paxilline，和idtF的基因，它是编码在C20或C21位置使帕斯帕林宁异戊二烯化的α-异戊二烯基转移酶的基因，分别产生帕斯帕利坦A和C。此外，我们还表明，野生型，ΔidtP和ΔidtF突变的C. paspali菌株的轴突培养物不能产生在C. paspali-Paspalum spp中存在的IDT的集合。协会。