Production of high-value chemicals in the button mushroom: a synthetic biology toolkit

担子菌门真菌为化学原料和改良疗法的细胞内操作提供了一个易于处理的真核系统。对面包师的酿酒酵母基因组进行了广泛的表征，包括5400个蛋白质编码基因，表明了与人类生化过程的一些相似之处。担子菌目真菌的复杂性增加，有13,000-16,000个蛋白质编码基因，提供了一种具有更大生物处理潜力的生物技术资源，其基本生物化学与酵母相似。担子菌门真菌产生丰富的可用作抗生素和农用化学品的有价值的化学物质，以及可用作生物催化剂的酶。担子菌菌株作为表达其他高等真菌基因的第二宿主尚未开发出来，这阻碍了从这一高产真菌群中发现、研究和生产高价值的化学物质和酶。一种易驯化的担子菌模式真菌，如纽扣蘑菇，是填补这一空白的理想人选，因为它比其他二次寄主，如酿酒酵母和曲霉，具有更多的复杂性和生物处理潜力。该计划的目的是建立一个用于纽扣蘑菇的合成生物学工具包。除了是模式生物和世界上种植最广泛的蘑菇，纽扣蘑菇还能够消化木质纤维素。因此，在这项工作中，将对木质纤维废料中提取的不同碳源对纽扣蘑菇的生长进行优化。基础知识将产生的各种碳源的基因表达。将开发结构性和诱导性启动子，后者由木质纤维素衍生物激活。通过在种子和生产培养之间切换碳源，将实现对目标基因表达的时间控制。这类似于对酵母半乳糖诱导系统所做的工作，在该系统中，葡萄糖的初始生长阶段之后是半乳糖的表达阶段。该工具包将促进培养真菌在发现、研究和生产高价值化学品(如萜类抗菌剂和其他疗法)以及有价值的酶(如脂肪加工和木质纤维素降解酶)方面的新用途。在该计划期间开发的可诱导表达元件将使研究人员能够利用纽扣蘑菇在静止阶段产生高价值的化学物质，如抗生素和抗真菌药物，如果在指数生长期产生这些化学物质可能是有毒的。纽扣蘑菇在木质纤维素上生长的能力将促进可再生碳源的使用和生物产品可持续路线的实施。

Basidiomycota fungi offer a tractable eukaryotic system for the intracellular manipulation of chemical feedstocks and modified therapeutics. Extensive characterisation of the baker's yeast Saccharomyces cerevisiae genome, comprising 5,400 protein-coding genes, has demonstrated some close similarities to human biochemical processes. The increased complexity of Basidiomycota fungi, with 13,000-16,000 protein-coding genes, provides a biotechnological resource of greater bioprocessing potential with similar underlying biochemistry as yeast. Basidiomycota fungi produce a wealth of valuable chemicals that can be used as antibiotics and agrochemicals, as well as enzymes that can be used as biocatalysts. A Basidiomycota strain that serves as a secondary host to express genes from other higher fungi has not yet been developed, and this is hindering the discovery, study and production of high-value chemicals and enzymes from this prolific group of fungi. A model tractable Basidiomycota fungus such as the button mushroom is ideally placed to fill this gap since it has more complexity and bioprocessing potential than other secondary hosts, such as S. cerevisiae and Aspergillus spp.The aim of this programme is to establish a synthetic biology toolkit for use in the button mushroom. As well as being a model organism and the most widely cultivated mushroom in the world, the button mushroom is able to digest lignocellulose. Therefore, in this work, the growth of the button mushroom will be optimised on different carbon sources that may derive from lignocellulosic waste. Fundamental knowledge will be generated of gene expression on the various carbon sources. Constitutive and inducible promoters will be developed, the latter being activated by lignocellulose derivatives. Temporal control of the expression of the genes of interest will be achieved by switching carbon sources between seed and production culture. This is similar to what is being done for the yeast galactose induction system, in which an initial growth phase in glucose is followed by an expression phase in galactose. The toolkit will facilitate novel uses of cultivated fungi in the discovery, study and production of high-value chemicals, such as terpenoid antimicrobials and other therapeutics, and valuable enzymes, such as lipid-processing and lignocellulose-degrading enzymes. The inducible expression elements developed during this programme will enable researchers to use the button mushroom to produce high-value chemicals in stationary phase, such as antibiotics and antifungals, which may be otherwise toxic if produced during the exponential growth phase. The ability of the button mushroom to grow on lignocellulose will facilitate the use of renewable carbon sources and the implementation of sustainable routes to bioproducts.