The role of RIPP proteins in plant pathogenic fungi.

最近，从曲霉菌中鉴定出一种称为Ustilooxin的小型分泌性毒素。它是真菌中第一个以核糖体合成的翻译后修饰蛋白(RIPP)，其中一个较小的肽序列(以多个串联拷贝的形式存在)是从一个较大的肽中提取出来的，然后被环化和广泛修改，实际上，这些修饰使人们无法将这些产物与NRPS衍生的天然产物区分开来，到目前为止，唯一其他的真菌RIPP是来自Amanita sp(苍蝇木耳、死亡帽等)的鹅膏毒素，这种真菌以具有广泛的生物活性而闻名。小麦病原真菌ZymosepVictoria tritici在其基因组中有一个与Uustilooxin相似的基因簇，RNAseq数据显示，在这种主要小麦病原菌的生物营养生长阶段，Amanita sp.这是效应器分子可能预期的那种表达模式，可能被用来在感染的早期阶段最大限度地减少宿主防御反应。其他研究也表明，这个基因座在不同的菌株之间是可变的，这表明它处于多样化的选择之下，正如在寄主-病原体相互作用中所预期的那样，寄主植物正在经历频繁的遗传变化(由于植物育种者的活动)。核心蛋白的基因被嵌入到一个共表达的基因簇中，同时也存在各种剪裁功能的基因，因此除了它的核心肽序列和类似的涉及内部酪氨酸残留物的循环事件之外，最终成熟化合物的化学结构无法很容易预测。该项目的目的是识别这种真菌产生的最终成熟代谢物，表征其在疾病中的作用(如果有的话)，然后寻求探索这类代谢物在各种宿主-病原体相互作用中的更广泛意义。该项目将涉及小黑曲霉和相关真菌的基因组和基因分析，使用基因中断技术来探索基因簇在这些真菌中的疾病中的作用。它还将涉及基因的异源表达作为一种合成生物学方法来探索这种化合物的生物合成途径，并结合分析化学来确定代谢物的结构和构象。

Recently a small secreted toxin called Ustiloxin has been characterised from Aspergillus sp. It is thefirst Ribosomally synthesised Post-translationally modified Protein (RiPP) of its class in fungi, where asmall peptide sequence (present in multiple tandem copies) is excised from a larger peptide thencyclised and extensively modified, indeed the modifications make it impossible to distinguish suchproducts from NRPS-derived natural products, the only other fungal RiPPs to date are the amatoxinsfrom Amanita sp (fly agaric, deathcap & etc), fungi famous for having wide ranging biological activities.The Wheat-pathogenic fungus Zymoseptoria tritici has a similar gene cluster to ustiloxin within itsgenome, and RNAseq data shows abundant expression during the biotrophic growth stages, peakingat the transition to necrotrophic growth of this major wheat pathogen. This is the sort of expressionprofile that might be expected from an effector molecule, perhaps being used to minimise hostdefence reactions during early stages of infection. Other studies have also shown this locus to bevariable between isolates, suggesting that it is under diversifying selection as might be expected ifinvolved in host-pathogen interactions where the hostplant is undergoing frequent genetic change(due to the activity of plant breeders). The gene for the core protein is embedded in a co-expressedgene cluster with genes for various tailoring functions also present so that the chemical structure ofthe final mature compound cannot be readily predicted - other than its core peptide sequence and alikely cyclisation event involving an internal tyrosine residue.The project will aim to identify the final mature metabolite made by the fungus, to characterise itsrole (if any) during disease and to then seek to explore the wider implications of this class ofmetabolite in various host-pathogen interactions.The project will involve genomic and genetic analysis of Z. tritici and related fungi, using genedisruption techniques to explore the role of the gene cluster in disease in these fungi. It will alsoinvolve heterologous expression of the genes as a synthetic biology approach to explore thebiosynthetic pathway for this compound, coupled with analytical chemistry to determine the structureand conformation of the metabolites.