Jia Xianbo, Liu Fangchen, Zhao Ke, Lin Junjie, Fang Yu, Cai Shouping, Lin Chenqiang, Zhang Hui, Chen Longjun, Chen Jichen
Institute of Soil and Fertilizer, Fujian Academy of Agricultural and Sciences, Fuzhou, China.
College of Resources and Environment, Fujian Agriculture and Forestry University, Fuzhou, China.
Front Microbiol. 2021 Jul 22;12:705853. doi: 10.3389/fmicb.2021.705853. eCollection 2021.
Prodigiosin is a promising secondary metabolite produced mainly by strains. To study the global regulatory mechanism of prodigiosin biosynthesis, a mutagenesis library containing 23,000 mutant clones was constructed with the EZ-Tn5 transposon, and 114 clones in the library showed altered prodigiosin production ability. For 37 of the 114 clones, transposon insertion occurred on the prodigiosin biosynthetic cluster genes; transposon inserted genes of the 77 clones belonged to 33 different outside prodigiosin biosynthetic cluster genes. These 33 genes can be divided into transcription-regulating genes, membrane protein-encoding genes, and metabolism enzyme-encoding genes. Most of the genes were newly reported to be involved in prodigiosin production. Transcriptional levels of the gene were significantly downregulated in 22 mutants with different inserted genes, which was in accordance with the phenotype of decreased prodigiosin production. Functional confirmation of the mutant genes involved in the pyrimidine nucleotide biosynthesis pathway was carried out by adding orotate and uridylate (UMP) into the medium. Gene complementation confirmed the regulatory function of the EnvZ/OmpR two-component regulatory system genes and in prodigiosin production.
灵菌红素是一种主要由菌株产生的有前景的次生代谢产物。为了研究灵菌红素生物合成的全局调控机制，使用EZ-Tn5转座子构建了一个包含23,000个突变克隆的诱变文库，文库中的114个克隆显示出灵菌红素生产能力发生改变。在114个克隆中的37个中，转座子插入发生在灵菌红素生物合成簇基因上；77个克隆的转座子插入基因属于33个不同的灵菌红素生物合成簇外基因。这33个基因可分为转录调控基因、膜蛋白编码基因和代谢酶编码基因。大多数基因是新报道的参与灵菌红素生产的基因。在22个具有不同插入基因的突变体中，该基因的转录水平显著下调，这与灵菌红素产量降低的表型一致。通过向培养基中添加乳清酸和尿苷酸（UMP）对参与嘧啶核苷酸生物合成途径的突变基因进行功能确认。基因互补证实了EnvZ/OmpR双组分调控系统基因和在灵菌红素生产中的调控功能。
