Characterization of environmental stimuli and regulatory networks involved in the expression of the colibactin polyketide in E. coli

大肠杆菌蛋白是一种非核糖体多肽/多酮基因毒素，在大肠杆菌和其他肠杆菌科中表达，具有引起DNA双链断裂从而阻断真核细胞周期的能力。由于这一特点，大肠杆菌蛋白对人类健康具有重要意义。迄今为止，对大肠杆菌素的研究主要集中在其对宿主细胞的作用上。然而，人们对这种聚酮在大肠杆菌中表达的条件知之甚少。我们假设大肠杆菌蛋白的表达受到环境刺激和细菌生长阶段的严格控制。大肠杆菌蛋白决定因子的表达受到不同的转录和转录后调控回路的影响，以响应宿主细胞因子、细菌中枢代谢和细菌生长阶段或应激条件。为了验证这一假设，我们将：-表征导致菌株特异性大肠杆菌蛋白表达差异的因素-阐明ClbR表达的调控及其作为clb基因表达的转录激活因子的作用-分析“碳储存调节剂”（Csr）系统对clb基因表达的控制-研究应激，饥饿，中枢代谢途径与大肠杆菌蛋白表达之间的假定相关性。-研究大肠杆菌素生物合成与大肠杆菌全球调控网络的整合。全面了解影响大肠杆菌蛋白表达的因素和条件将有助于开发针对大肠杆菌蛋白产生菌的治疗方法。此外，它将支持colibactin过表达和/或纯化，这是对这种有趣但仍未充分研究的化合物进行功能和结构表征的先决条件。

Colibactin is a hybrid non-ribosomal peptide/polyketide genotoxin expressed in E. coli and different Enterobacteriaceae with the ability to cause DNA double strand breaks and thus to block the eukaryotic cell cycle. Due to this characteristics, colibactin can have important implications for human health. So far, studies about colibactin mainly focused on its effect on the host cells. The conditions under which this polyketide is expressed in E. coli are, however, only poorly understood. We postulate that colibactin expression is tightly controlled by environmental stimuli and the bacterial growth stage. Expression of the colibactin determinant is affected by different transcriptional and posttranscriptional regulatory circuits in response to host cell factors, the bacterial central metabolism and the bacterial growth stage or stress conditions. To test this hypothesis, we will:- characterize factors leading to strain-specific differences in colibactin expression- elucidate the regulation of ClbR expression and its role as a transcriptional activator of clb gene expression- analyze the control of clb gene expression by the 'carbon storage regulator' (Csr) system - investigate the putative correlation between stress, starvation, central metabolic pathways and colibactin expression.- study the integration of colibactin biosynthesis into global regulatory networks of E. coli. The comprehensive picture regarding the factors and conditions influencing the expression of colibactin will be very helpful to develop therapeutics against colibactin producing bacteria. Moreover, it will support colibactin overexpression and/or purification, which is a prerequisite for the functional and structural characterization of this interesting and still insufficiently studied compound.