XRE家族转录因子ProR调控粘质沙雷氏菌合成灵菌红素的分子机制解析及其功能分析

灵菌红素具有抗细菌和抗肿瘤等功能，其主要由粘质沙雷氏菌合成。目前粘质沙雷氏菌合成灵菌红素调控机制的解析依然有限，这制约了高效合成灵菌红素工程菌株的构建。项目前期通过构建转座子插入突变文库筛选获得了灵菌红素合成新型正调控因子ProR，proR突变株合成灵菌红素能力为野生型菌株的25.79%，目前其调控菌株合成灵菌红素的分子机制并不清晰。项目拟在前期证实ProR通过间接调控pig基因簇的表达水平，调控菌株合成灵菌红素基础上，组合比较转录组学分析、ChIP-Seq、RT-qPCR、融合报告基因构建、EMSA和DNaseⅠ足迹试验等技术解析ProR调控粘质沙雷氏菌合成灵菌红素的作用靶点、信号通路和分子机制。组合生物信息学分析和生理学实验解析广泛存在于微生物但功能未知的转录因子ProR的功能。项目将为通过系统代谢工程构建高产灵菌红素菌株和研究其他物种中ProR或ProR-like蛋白的功能提供借鉴。

Prodigiosin, which has important functions such as anti-bacterial and anti-tumor, is mainly synthesized by Serratia marcescens. At present, the regulatory mechanism behind prodigiosin biosynthesis in S. marcescens is still limited, which restricts the construction of efficient prodigiosin biosynthesis engineering strains. In our previous work, a transposon mutant library was constructed to identify the genes related to prodigiosin synthesis, and the proR gene encoding the XRE family regulator ProR was found to positively regulates prodigiosin synthesis in strain JNB5-1. Disruption of proR conferred a remarkably decreased production of prodigiosin, with the prodigiosin synthesis ability of proR mutant is 25.79% of that of wild-type strain. The molecular mechanism how ProR regulates prodigiosin synthesis in S. marcescens is not clear. Based on the results of previous experiments showed that ProR indirectly regulates the expression levels of pig gene cluster thus positively regulates prodigiosin synthesis in S. marcescens, in this project, we will combine comparative transcriptomics analyses, ChIP-Seq analysis, RT-qPCR, fusion reporter gene construction, EMSA, and DNase I footprinting assay to reveal the binding site, signal pathway and molecular mechanism of ProR regulating prodigiosin synthesis in S. marcescens. Bioinformatics analysis and physiological experiments will be used to analyze the function of ProR, a transcription factor widely existing in microorganisms but with unknown function. The project will provides a reference for the construction of high-yield prodigiosin producing strains through systematic metabolic engineering in the future, and provides a new idea to study of the function of ProR or ProR-like proteins in other species.