Function of the cypX and moxY genes in aflatoxin biosynthesis in Aspergillus parasiticus
Function of the cypX and moxY genes in aflatoxin biosynthesis in Aspergillus parasiticus
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DOI：
10.1128/aem.71.6.3192-3198.2005
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发表时间：
2005-06-01
影响因子：
4.4
通讯作者：
Yabe, K
中科院分区：
文献类型：
--
作者：
Wen, Y;Hatabayashi, H;Yabe, K
黄曲霉毒素的生物合成途径有氧阿维菌素(OAVN)->；阿维芬(AVR)->；羟基杂色酮(HVN)->；多聚半缩醛醋酸酯(VHA)，而黄曲霉毒素基因簇中存在的cypX和Moxy基因也参与了黄曲霉毒素的合成。为了更详细地阐明这两个基因的功能，我们中断了产黄曲霉NRRL2999中的基因。缺失cypX的突变体失去了黄曲霉毒素的产量，并在菌丝体中积累了AVR。虽然该突变株在饲养实验中将HVN、杂色酮(VONE)、VHA和杂色酚醋酸酯(VOAc)转化为黄曲霉毒素，但它不能从OAVN或AVR产生黄曲霉毒素。Moxy缺失的突变体也失去了黄曲霉毒素的生产能力，但它新积累了HVN和VONE。在饲养实验中，该突变体将VHA或VOAc转化为黄曲霉毒素，但不能将OAVN、AVR、HVN或VONE转化为黄曲霉毒素。这些结果表明，cypX编码AVR单加氧酶，催化AVR到HVN的反应；Moxy编码HVN单加氧酶，催化从HVN到VHA和从VONE到VOAc的Baeyer-Villiger反应。在这项工作中，我们设计了一种简单快速地从多种真菌中提取DNA的方法，用于PCR分析，其中结合了摇床破碎法和苯酚提取法。
The pathway oxoaverantin (OAVN) -> averufin (AVR) -> hydroxyversicolorone (HVN) -> versiconal hemiacetal acetate (VHA) is involved in aflatoxin biosynthesis, and the cypX and moxY genes, which are present in the aflatoxin gene cluster, have been previously suggested to be involved in this pathway. To clarify the function of these two genes in more detail, we disrupted the genes in aflatoxigenic Aspergillus parasiticus NRRL 2999. The cypX-deleted mutant lost aflatoxin productivity and accumulated AVR in the mycelia. Although this mutant converted HVN, versicolorone (VONE), VHA, and versiconol acetate (VOAc) to aflatoxins in feeding experiments, it could not produce aflatoxins from either OAVN or AVR. The moxY-deleted mutant also lost aflatoxin productivity, whereas it newly accumulated HVN and VONE. In feeding experiments, this mutant converted either VHA or VOAc to aflatoxins but did not convert OAVN, AVR, HVN, or VONE to aflatoxins. These results demonstrated that cypX encodes AVR monooxygenase, catalyzing the reaction from AVR to HVN, and moxY encodes HVN monooxygenase, catalyzing a Baeyer-Villiger reaction from HVN to VHA as well as from VONE to VOAc. In this work, we devised a simple and rapid method to extract DNA from many fungi for PCR analyses in which cell disruption with a shaker and phenol extraction were combined.