Defosse Tatiana A, Mélin Céline, Clastre Marc, Besseau Sébastien, Lanoue Arnaud, Glévarec Gaëlle, Oudin Audrey, Dugé de Bernonville Thomas, Vandeputte Patrick, Linder Tomas, Bouchara Jean-Philippe, Courdavault Vincent, Giglioli-Guivarc'h Nathalie, Papon Nicolas
Biomolécules et Biotechnologies Végétales, Université François-Rabelais de Tours, EA 2106, F-37200 Tours, France Groupe d'Etude des Interactions Hôte-Pathogène, Université d'Angers, EA 3142, F-49933 Angers, France.
Biomolécules et Biotechnologies Végétales, Université François-Rabelais de Tours, EA 2106, F-37200 Tours, France.
FEMS Yeast Res. 2016 Sep;16(6). doi: 10.1093/femsyr/fow078. Epub 2016 Sep 11.
The fungal CTG clade comprises a number of well-known yeasts that impact human health or with high biotechnological potential. To further extend the set of molecular tools dedicated to these microorganisms, the initial focus of this study was to develop a mycophenolic acid (MPA) resistance cassette. Surprisingly, while we were carrying out preliminary susceptibility testing experiments in a set of yeast species, Meyerozyma guilliermondii, although not being a MPA producer, was found to be primarily resistant toward this drug, whereas a series of nine related species were susceptible to MPA. Using comparative and functional genomic approaches, we demonstrated that all MPA-susceptible CTG clade species display a single gene, referred to as IMH3.1, encoding the MPA target inosine monophosphate dehydrogenase (IMPDH) and that MPA resistance relies on the presence in the M. guilliermondii genome of an additional IMPDH-encoding gene (IMH3.2). The M. guilliermondii IMH3.2 gene displays marked differences compared to IMH3.1 including the lack of intron, a roughly 160-fold higher transcription level and a serine residue at position 251. Placed under the control of the M. guilliermondii actin 1 gene promoter, IMH3.2 was successfully used to transform Lodderomyces elongisporus, Clavispora lusitaniae, Scheffersomyces stipitis and Candida parapsilosis.
真菌CTG进化枝包含许多影响人类健康或具有高生物技术潜力的知名酵母。为了进一步扩展针对这些微生物的分子工具集，本研究最初的重点是开发一种霉酚酸（MPA）抗性盒。令人惊讶的是，当我们在一组酵母物种中进行初步药敏试验时，季也蒙毕赤酵母虽然不是MPA产生菌，但被发现对这种药物具有主要抗性，而一系列九个相关物种对MPA敏感。使用比较基因组学和功能基因组学方法，我们证明所有对MPA敏感的CTG进化枝物种都有一个单一基因，称为IMH3.1，编码MPA靶点肌苷单磷酸脱氢酶（IMPDH），并且MPA抗性依赖于季也蒙毕赤酵母基因组中额外的一个编码IMPDH的基因（IMH3.2）。季也蒙毕赤酵母的IMH3.2基因与IMH3.1相比显示出明显差异，包括缺乏内含子、转录水平高出约160倍以及在第251位有一个丝氨酸残基。在季也蒙毕赤酵母肌动蛋白1基因启动子的控制下，IMH3.2成功用于转化长孢洛德酵母、葡萄牙棒孢酵母、树干毕赤酵母和近平滑念珠菌。
